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The occurrence, chemistry, resolution, and analysis of amino acids published in the literature from 2013 finished with the year of 2016 are reviewed in this Chapter which is arranged in sections similar to previous Volumes in this Specialist Periodical report. Scientific Papers published during 2013–2016 have been sourced mainly from the Web of Science databases and Pubmed on the internet and from scanning a selection of major journals.

Several naturally occurring amino acid derivatives are more and more in the focus as sources for new agrochemicals. The 53 most important natural products of amino acids are presented in a review together with their natural sources, mode of action, and herbicidal, fungicidal, or insecticidal activities.3 

It is known that Arg, the most basic amino acid, occurs less frequently than Lys in proteins. The few important proteins abundant in Arg have important roles in biological systems. A review collects the data of occurrence, functions, and the biological significance of these Arg-rich proteins.4  Leu is a potential signalling molecule to regulate cell growth and metabolism. Structure–activity relationships of Leu derivatives in HeLa S3 cells were investigated for cellular uptake and for the induction of phosphorylation. The results may provide a new insight into therapeutics targeting both l-amino acid transporters 1 and Leu sensor.5 

Phenyl-Gly-type amino acids occur in a wide variety of natural peptides. The biosynthesis of 4-hydroxyphenyl-Gly, 3,5-dihydroxyphenyl-Gly, and phenyl-Gly was investigated. Structures and properties of phenyl-Gly containing natural products, the biosynthetic origin and incorporation of phenylglycines are discussed in a review.6  4-Methyl-Pro (4-mPro) is a rare nonproteinogenic amino acid produced by cyanobacteria. Eight biosynthetic gene clusters were found from available cyanobacteria genomes, showing that 4-mPro is a good marker to discover previously unknown nonribosomal peptides.7 

Amino acids represent a fraction of organic matter in marine and freshwater ecosystems. The occurrence of d-amino acids is usually linked to the presence of bacteria. The distribution of l- and d-amino acids in the lacustrine environment of Terra Nova Bay, Antarctica was investigated.8  Microorganisms that utilize various d-amino acids were successfully isolated from deep-sea sediments. Some of the isolates exhibited high enantioselective degradation activities to various d-amino acids.9  An antialgal compound was isolated from the cultured broth of Streptomyces jiujiangensis JXJ 0074(T). Based on the data of different analytical methods, the active compound was identified as L-Val which showed antialgal activity. This is the first report showing that L-Val is active against cyanobacteria.10  Diverse d-amino acids have been found in mammalian tissues. The physiological functions of these d-amino acids are being gradually clarified. It has been demonstrated that D-Ser, D-Asp, D-Ala, and D-Cys play important roles in the nervous and endocrine systems. The investigations of metabolism and the physiological functions of d-amino acids provide new therapeutic and diagnostic strategies for diseases related to the nervous and endocrine systems.11  D-Ser is a key signalling molecule utilized by neurons and astroglia in the mammalian central nervous system. Alterations in the extracellular levels of D-Ser disrupt cell–cell signalling that leads to many chronic or acute neurological and psychiatric disorders, and are associated with addictive behaviour. Experimental data supports that astroglia and neurons use different pathways to regulate levels of extracellular D-Ser.12  A rare Gln derivative, hemerocallisamine I, was isolated from the flower buds of daylily. It was first reported that a Gln derivative with a pyrrole ring is found in natural plants.13  Increased reactive oxygen species are accompanied by 2-aminobutyric acid accumulation and compensatory maintenance of myocardial glutathione levels. It was demonstrated for the first time that 2-aminobutyric acid modulates glutathione homeostasis in the myocardium.14 

β-Amino acids are components of complex natural products generating significant and unique biological functions. The de novo synthesis of β-amino acids and the mechanisms of β-amino acid incorporation into natural products are summarized.15 

Mycobacterium tuberculosis (Mtb) is responsible for 9 million active tuberculosis cases annually, resulting in 1.5 million death cases per year worldwide. It is known that Mtb uses Arg as a nitrogen source in vitro, but the metabolic pathways have not been identified. It was found that, both nitrogen and carbons from Arg can be incorporated into the central metabolism of Mtb. The highly induced pathway for Arg utilization in Mtb differs from that of other bacteria including non-tuberculous mycobacteria.16 

The increasing interest of modified peptides in chemical engineering of proteins and also as therapeutic agents has refreshed research toward the development of derivatives of new natural and non-natural amino acids.

Oxidative stress plays an important role in the development of atrial fibrillation. Derivatives of Arg including asymmetric dimethyl-Arg (ADMA) are central to nitric oxide metabolism and nitrosative stress. The circulating levels of ADMA, L-Arg, symmetric dimethyl-Arg (SDMA), and the ratio of L-Arg/ADMA to incidence of atrial fibrillation were investigated. The results revealed that circulating ADMA is not strongly associated with new-onset atrial fibrillation, and L-Arg and SDMA are also not predictive of long-term incidence of atrial fibrillation.17  A growing number of studies showed elevated concentrations of circulating ADMA and SDMA leading to mortality and cardiovascular diseases. A systematic review summarizes the evidence from studies of ADMA and SDMA with the risks of all-cause mortality and incident cardiovascular disease in meta-analyses. It was concluded that ADMA and SDMA are independent risk markers for all-cause mortality and cardiovascular disease across different populations and methodological approaches.18  In search for new drugs lowering arterial blood pressure, novel potential renin inhibitors based on human angiotensinogen, were designed and synthesized. All these inhibitors contain unnatural amino acids that are derivatives of N-alkylleucyl-β-hydroxy-γ-amino acids. In vitro renin inhibitory activity of all obtained compounds was within the range 10−6–10−9 M.19 

A series of different amino acid-bearing thieno[2,3-D]pyrimidine moiety and a tricyclic imidazothienopyrimidine of the Gly derivative were synthesized. All the obtained amino acid-derivatives were screened for their post-irradiation protective efficacy. Most of the newly synthesized derivatives showed significant protective effects against injuries induced by γ-irradiation exposure, and they may be promising curative agents against γ-irradiation induced oxidative stress and physiological disturbance in different organs.20 

A novel series of biaryl-based phenylalanines with a carboxyl-benzene (or a carboxyl-thiophene ring) was designed, synthesized, and pharmacologically characterized in vitro. The structural modifications were focused on positions 5- and 4- of the Phe ring. Various combinations of small-sized groups, both polar and lipophilic, hydrogen bond donors and acceptors, were investigated. All the target amino acids were pharmacologically characterized by radioligand binding at native AMPA, kainate, and NMDA receptors, and the structure–activity relationships were also studied.21 

Thiazolides are polypharmacologycal agents with at least three mechanisms of action against a broad spectrum of parasites, bacteria and viruses. New amino-acid ester thiazolide prodrugs were synthesized in order to improve their systemic absorption and tested.22  Oxazolidinone derivatives serve as important drugs among antibiotics, since they act on multidrug-resistant bacteria. A small library of compounds based on isoxazolidinone and dehydro-β-Pro was designed with the aim to obtain antibacterial agents and monoaminooxidase(MAO)-inhibitors.23  Substituted indolizidine and quinolizidine derivatives are readily assembled from cyclic amino acids (Pro or pipecolic acid) and γ-nitroaldehydes by a simple decarboxylative annulation process.24 

Derivatives of aminobenzoic acid were functionalized via a chemoselective carbene insertion manner without implementing protection and deprotection strategy under mild reaction conditions leading to carboxy and hydroxy functionalized α-amino esters (27 examples).25  The neurotrophic effects of L-Glu and β-phenyl-Glu hydrochloride were compared. It was found that β-phenyl-Glu hydrochloride was more potent than L-Glu in neuroprotection and increasing survival rate.26  A new member of the dimethylallyl-Trp synthase superfamily catalyzes prenylations of both Tyr and Trp derivatives. The results enhance the relationship of Tyr O- and Trp C7-prenyltranferases and provide a new enzyme for production of prenylated derivatives.27 

Arctigenin, a traditional medicine with many pharmacological activities, has been restricted due to its poor solubility in water. Five amino acid derivatives of arctigenin have been synthesized using Gly, Ala, Val, Leu, and Ile. The results showed that the amino acid derivatives have better solubility and exhibit higher anti-tumour activity than arctigenin.28 

New amino acid derivatives with carbocycles of adamantine and quinaldic acid were developed; their in vitro antiviral activity against highly pathogenic influenza A virus (H5N1) was evaluated; and found that they suppressed viral replication.29 

New N-(4-substituted phenyl)Gly derivatives have been synthesized. The intermediates (the chalcone and the thiosemicarbazone derivative) were derivatized and/or cyclized into different heterocyclic target derivatives and evaluated as potential anti-inflammatory agents.30  Novel rifamycins containing l-amino acid esters were produced, and their structure–activity relationships in solution were studied. The presence of the rifamycins’ structures influenced antibacterial properties.31  Schiff base compounds of cinnamaldehyde and amino acids have been synthesized and investigated for their antimicrobial activities. A total of 24 Schiff base compounds were synthesized using a simple approach with 3 cinnamaldehyde derivatives and 8 amino acids. Results from the structure–activity relationship suggest that both –p–Cl on benzene ring of cinnamaldehyde and the number of –COOK of amino acid salts significantly contributed to antimicrobial activity.32 

A series of bile acid (cholic acid and deoxycholic acid) aryl/heteroaryl amides linked via α-amino acid were synthesized and tested against 3 human cancer cell-lines. Some of the conjugates showed promising results to be new anticancer agents with good in vitro results. They showed fairly good activity against the breast cancer cell line with respect to Cisplatin and comparable with respect to Doxorubicin and showed better activity against glioblastoma cancer cell line with respect to both Cisplatin and Doxorubicin drugs used as standards.33 

A series of red-shifted azobenzene amino (Scheme 1) acids have been synthesized via a two-step procedure. Derivatives of Tyr were first oxidized to the corresponding quinonoidal spirolactones followed by the unsymmetrical azo formation with the substituted phenylhydrazines in the presence of the ceric ammonium nitrate catalyst.34 

Kojic acid (5-hydroxy-2-hydroxymethyl-4H-pyran-4-one) is a natural product that is produced by many species of fungi. A simple and efficient method for the preparation of dialkylated kojic acid based α-amino acid derivatives was described. The novel derivatives might find application as tyrosinase inhibitors.35 

Oleanolic acid or oleanic acid (3β-hydroxy-olean-12-en-28-oic acid) is a naturally occurring pentacyclic triterpenoid related to betulinic acid. It is widely distributed in food and plants where it exists as a free acid or as an aglycone of triterpenoid saponins and exhibits a wide range of pharmacological and biological activities. Four known and two new amino acid conjugates of oleanolic acid were prepared, and investigated for their cytotoxic effects. The results revealed that two derivatives showed significantly increased inhibition rates than the parent oleanolic acid.36  Novel derivatives of ligustrazine-oleanolic acid were designed, and synthesized by conjugating amino acids to the 3-hydroxy group of ligustrazine-oleanolic acid by ester bonds. Their cytotoxicity was evaluated on four cancer cell lines.37  Hepatic fibrosis is a naturally occurring wound-healing reaction, with an imbalance of extracellular matrix during tissue repair response, which can further deteriorate to hepatocellular carcinoma without timely treatment. It was found that Gly derivative of ligustrazine-oleanolic acid selectively inhibited the proliferation and induced apoptosis indicating that Gly derivative of ligustrazine-oleanolic acid might be a potential antifibrosis agent for the therapy of hepatic fibrosis.38 

Methylation of Lys is one of the important post-translational modifications of histones that produces N(ε)-mono-, di-, or trimethyl Lys residues. Multiple, site-specific Lys methylations of histones are essential to define epigenetic statuses and control heterochromatin formation, DNA repair, and transcription regulation. A new method was developed for preparing histones bearing multiple N(ε)-monomethyl Lys residues at specified positions that enables the installation of authentic N(ε)-monomethyl Lys at multiple positions within a protein for large-scale production.39 

All 20 common natural proteinogenic and 4 other α-amino acid-isosteric α-amino tetrazoles have been synthesized. Since the tetrazole group is bioisosteric to the carboxylic group, these derivatives are widely used in medicinal chemistry and drug design. The synthetic process involves the use of the Ugi tetrazole reaction followed by deprotection (Scheme 2).40 

Amino acids with their variable side chains are ideal candidates for synthesizing biodegradable functional polyesters. The synthetic methods for poly-α-hydroxy esters derived from amino acids were reviewed.41  The efficient tRNA-mediated incorporation of the hydroxamate containing amino acid, N(ε)-acetyl-N(ε)-hydroxy-L-Lys into a transcription factor was reported. The tetrahydrofuranyl and tetrahydropyranyl O-protecting groups can be removed using mild acid conditions. These protecting groups can be used as valuable alternative for O-protection.42  Two new antifouling zwitterionic polymers, poly(lysine methacrylamide) and poly(ornithine methacrylamide) derived from natural amino acids Lys and Orn, respectively were developed and investigated. The super low fouling, biomimetic, and multifunctional properties of poly(lysine methacrylamide) and poly(ornithine methacrylamide) make them promising materials for a wide range of applications, such as implant coating, drug delivery and biosensing.43  Amino-acid-based chiral surfactants with polymerizable moieties are developed for the production of chiral nanoparticles. The synthesized particles are tested for their ability as nucleating agents in the enantioselective crystallization of amino acid conglomerate systems. The results demonstrate that only the chiral nanoparticles made of the polymerizable surfactant are able to act efficiently as nucleation agent in enantioselective crystallization.44 

The preparation by a direct condensation method, characterization, and cytocompatibility of homo- and hetero-polyesters of α-hydroxy amino acid derivatives with or without lactic acid conjugation has been described. It was concluded that overall selective cytocompatibility and bioactivity might render α-hydroxy amino acid polymers useful as extracellular matrix-mimicking materials for tissue engineering.45  An asymmetric decarboxylative Csp(3)–Csp(2) cross-coupling has been achieved via a mild, operationally simple protocol with Ni-catalyst. Variety of naturally abundant α-amino acids and aryl halides are transformed into valuable chiral benzyl amines.46  A highly diastereoselective acid-catalyzed N,O-acetalization/intramolecular transcarbamoylation cascade of reactions between protected α-amino acid derivatives (Ser and Thr) and non-natural α-amino acid derivatives with tetramethoxyalkanes has been reported. The resulting oligocyclic N,O-acetals have been used as excellent chiral building blocks for asymmetric transformations. The complete diastereoselectivity achieved with natural amino acid precursors is completely lost with their non-natural analogues.47  Incorporation of a non-natural Arg analogue (guanidiniocarbonyl pyrrole) into a cyclic peptide is capable of completely altering the self-assembly properties of the peptide. In contrast to the peptide which does not self-assemble, guanidiniocarbonyl pyrrole-containing peptide forms cationic nanofibers of micrometer length that are capable of gene transfection.48 

Several new functions of amino acids have been recently discovered that could result in new applications. E.g. oral stimulation by Glu triggers the cephalic phase response to prepare for food digestion. Branched-chain amino acids are the major components of muscles, and ingestion of branched-chain amino acids has been found to be effective for decreasing muscle pain. Amino acids can be used in a novel clinical diagnostic method: the balance of amino acids in the blood could be an indicator of the risk of diseases such as cancer. The newly discovered functions of amino acids are discussed.49  It was observed that Gln levels in plasma were significantly lower just after cardiac surgery compared to pre-operative levels.50 

The interactions of the side chains of Arg and Lys was investigated with each of the 19 non-Gly amino acids in proteins in the protein data bank. The guanidino group of Arg interacts with non-polar aromatic and aliphatic side chains above and below the guanidinium plane while hydrogen bonding with polar side chains is restricted to in-plane positions. In contrast, non-polar side chains interact with the aliphatic part of the lysine side chain. Molecular dynamics simulations underlie the preference for Arg as a mobile charge carrier in voltage-sensing domains.51  To understand the molecular mechanisms of effect of additives to stabilize proteins, molecular dynamics simulations of the surface residues lysozyme was performed in the presence of three commonly used additives: Arg, Lys, and guanidine that have different effects on stability of proteins and have different structures with some similarities. The investigations revealed that the internal dynamics, as well as the lifetimes of the hydrogen bonds within the protein changes depending on the additives.52 

A three-step process involving Cu-catalyzed allylation of Ser-, Asp-, and Glu-derived organo-Zn reagents have been used for the synthesis of 7-oxo, 8-oxo, and 9-oxo amino acids.53 

Amino acids derived steroidal and nonsteroidal architectures have also been developed. The benzofused, amino acid-derived steroidal and nonsteroidal molecules had promising biological activity in hormonal related disorders.54  Optically pure valinol was prepared via different ω-transaminases from the corresponding prochiral hydroxy ketone. Reductive amination was performed in organic solvent using 2-propyl amine as amine donor, and Ala was applied in aqueous medium.55 

Five simple rules of thumb are developed to summarize the adsorption properties of the proteinogenic α-amino acids as mediated by hydrogen bonding on silicon surface, and they are expected to provide a helpful guide to future studies of larger biomolecules and their potential applications.56  The adsorption of some amino acids by fullerene (C60) and fullerene nanowhiskers have been investigated. The aromatic group of fullerene did not interact with the hydrophobic side chains (alkyl chain, cyclic structures of Trp, Phe and Pro) of amino acids.57 

The role of hydrophobicity of the side chain of amino acids in the formation of hydrogel was studied by using Fmoc-Nle and Fmoc-Met. The results indicate that Fmoc-Met forms reversible hydrogels in water, whereas Fmoc-Nle fails to display any gelation under similar conditions. The difference in the self-association behaviour of Fmoc-Met and Fmoc-Nle emphasise the importance of weak noncovalent interaction between side chains to stabilise supramolecular self-assembly of Fmoc-protected amino acids.58 

Unlike other Fmoc-functionalized amino acid gelators, Fmoc-Lys(Fmoc)-OH exhibits pH-controlled ambidextrous gelation. (hydrogelation at different pH values as well as organogelation). Spectroscopic analyses proved that the self-assembly of Fmoc-Lys(Fmoc)-OH was driven by aromatic π–π stacking and hydrogen bonding interactions in both hydrogels and organogels.59  Fmoc-Lys(Fmoc)-OH was demonstrated as gelators to gelate a variety of alcohols and aromatic solvents under the sonication conditions. The ultrasound-triggered organogel also exhibited thixotropic property. The gels with the fibrous 3D network structure were unravelled into sols. However, after standing, these sols returned to the gels showing a more ordered lamella-like packing structure.60  The nature of π–π interactions in the self- and coassembly of Fmoc-Phe-derived hydrogelators was investigated by systematically varying the electron-donating or electron-withdrawing nature of the side chain substituents and correlating these effects to the emergent assembly and gelation properties of the systems. The findings provide significant insight into the structure–function relationship for Fmoc-Phe-derived hydrogelators.61  A review summarizes the recent progress of self-organization of Fmoc-amino acids and Fmoc-modified di-, tri-, tetra- and pentapeptides.62 

Gemini surfactants have been used for in vitro gene delivery. Amino acid-derived gemini surfactants combine the special aggregation properties with high biocompatibility and biodegradability. Novel Ser-derived gemini surfactants, differing in alkyl chain lengths and in the linker group bridging the spacer to the head groups (amine, amide and ester) were evaluated.63  The effects of the spacer chain length of amino acid-based gemini surfactants on the formation of wormlike micelles in aqueous solutions were investigated. The surfactants were synthesized by reacting dodecanoyl-Glu anhydride with diamine compounds. Formation of cation-rich molecular aggregates was not observed when the longest spacer analogue (8 methylene units) was used.64  A review reports the most important contributions in the structure, synthesis, physicochemical and biological properties (toxicity, antimicrobial activity, and biodegradation) of natural amino acid-based (Arg, Lys, Ser, Ala, Sar, Asp, and Cys) gemini surfactants and some potential applications. Amino acid-based gemini surfactants have some benefits compared with the classical gemini surfactants.65  The development of new antimicrobial agents is very important because of the rapid increase in the number of multiple drug resistant bacteria and fungi. Synthetic amino acid-based surfactants constitute a promising alternative to conventional antimicrobial compounds. In a review, the structural features that promote antimicrobial activity of amino acid-based surfactants and also the synthesis and basic physico-chemical properties are discussed. Cationic surfactants based on amino acids show excellent antimicrobial and antifungal properties.66  Anionic, cationic, and zwitterionic amphiphiles can be prepared as surfactants by using one of the 20 proteinogenic amino acids. A review gives examples of procedures and discusses important physicochemical properties and various applications of amino acid-based surfactants, as well as highlights concepts that are unique to amino acid-based surfactants. E.g. Surfactants based on amino acids with two carboxyl groups are effective chelating agents; a surfactant based on Cys readily oxidizes into the corresponding cystine compound, which can be regarded as a gemini surfactant.67  Another review discusses surfactant-amino acids and surfactant–surfactant interactions in aqueous medium.68 

Threonine aldolases catalyze the pyridoxal phosphate-dependent condensation between small amino acids (Gly) and aldehydes. A review briefly summarizes the reaction mechanism and lists all published synthetic reactions by threonine aldolases.69 

A paper reviews the literature data on amino acid supplementation with Arg, Glu, and beta-hydroxy-beta-methylbutyric acid and wound healing in diabetic foot. A pilot study showed that administration with hydroxyproline (Hyp), a major component of collagen, improved the healing of diabetic foot wounds via increased collagen production.70  Determination of the 4-hydroxy-l-Pro concentration provides information for the diagnosis and prognosis of diseases caused by disorders of collagen metabolism. It was found that LC–MS method would be advantageous for measuring the hydroxyl-Pro concentration and for the diagnosis of diseases associated with abnormalities of collagen metabolism.71  4-Methylproline is a rare nonproteinogenic amino acid produced by cyanobacteria through the action of a zinc-dependent long-chain dehydrogenase. The use of biosynthetic genes of 4-methylproline helped to discover new bioactive compounds from cyanobacteria.7 

Prodrugs in which the hydroxyl moiety is reversibly protected as a carbamate ester linked to the amino group of a natural amino acid (Ile or β-Ala) have been produced. Prodrugs having amino acids with hydrophobic side chains were readily absorbed after intragastric administration.72  Prodrugs of resveratrol in which the OH groups are engaged in an N-monosubstituted carbamate ester (–OC(O)NHR) linkage with a natural amino acid (Leu, Ile, Phe, Thr) to prevent conjugation and modulate the physicochemical properties of the molecule that was synthesized in high yield; characterized; and the stability and in vivo pharmacokinetic behaviour also was determined. It was concluded that prodrugs based on the N-monosubstituted carbamate ester bond have the appropriate stability profile for the systemic delivery of phenol compounds.73 

The protonation constants, the protonation enthalpy changes, and the solubility of six natural α-amino acids (Gly, Ala, Val, Leu, Ser, and Phe) have been performed in NaCl and in (CH3)4NCl media with new potentiometric experiments.74  The protonation equilibrium concentrations of proton–ligand formation as a function of pH were investigated. Biologically active ligands like amino acids, peptides, DNA constituents, and amino acid esters in nonaqueous media have been investigated in a review.75  Complex formation of equilibrium for the binary complexes of Cu(ii) with 1-aminocyclopropane carboxylic acid and 3,3-bis(1-methylimidazol-2-yl)propionic acid were studied. The ternary complexes are formed in a stepwise mechanism.76 

Antibody-drug conjugates as potent antitumour drugs provide increased efficacy, specificity, and tolerability over chemotherapy for the treatment of cancer. A novel method is described that enables site-specific conjugation of toxins to antibodies using chemistry to produce homogeneous, potent, and highly stable conjugates. A cell-based mammalian expression system was used that is capable of site-specific integration of a non-natural amino acid containing an azide moiety. The azide-alkyne linkage has high stability.77  The hybrid molecules of antibody-drug conjugates consist of a tumour antigen-specific antibody coupled to a chemotherapeutic small molecule. A cell-free protein expression system was established for production of antibody drug conjugates through site-specific incorporation of para-azidomethyl-l-Phe. The resultant antibody-drug conjugates were highly potent in in vitro cell cytotoxicity assays.78 

A novel amino acid racemase was discovered through exploration of natural variation in Arabidopsis thaliana. N-malonyl-D-allo-Ile and a novel amino acid racemase that is involved in its biosynthesis was identified. This finding provides the first functional characterization of a eukaryotic member of a large and widely conserved phenazine biosynthesis protein family, and a new d-amino acid racemase gene family is also identified.79 

A simple theoretical model of amino acid similarity matrices, which allows splitting the matrix into two parts, was introduced. The new synthetic amino acid properties are derived from the pairwise similarities and used to reconstruct similarity matrices. The new properties derived from amino acid similarity matrices correlate highly with properties known to be important for molecular evolution such as hydrophobicity, size, shape, and charge of amino acids.80 

A group of five amino acid based zwitterionic vinyl monomers, based on Ser, Lys, Orn, Glu, and Asp were developed for potential antifouling applications and grafted on gold chips by polymerization. Considering multiple applications (e.g. medical devices and drug delivery) of the antifouling materials, the cytotoxicity of monomers and polymer nanogels for all five materials at various concentrations were evaluated. Very low cytotoxicity was observed for all tested amino acid-based monomers and nanogels, which is comparable or even lower than the traditional and some newly developed antifouling materials.81 

Genetically encoded electrophilic unnatural amino acid found to react with His and Lys. In addition to efficient cross-linking of proteins inter- and intramolecularly, the unnatural amino acid permits direct stapling of a protein α-helix and covalent binding of native membrane receptors.82 

A review examines recent advances in substrate-controlled asymmetric reactions induced by the chirality of α-amino acid templates in natural product synthesis and related areas.83 

Small molecules that target different components of the spliceosome represent valuable research tools to investigate this complicated macromolecule. N-palmitoyl-L-Leu was identified as a new splicing inhibitor that blocks a late stage of spliceosome assembly.84 

Amino acid transporters are expressed in the body and form a series of channels to pump nutrients against concentration gradients into cells. Abnormal expression of amino acid transporters is often associated with cancer, addiction, and multiple mental diseases. A new class of amino acid mimics (boramino acids) (Scheme 3) are described that can serve as general imaging probes for amino acid transporters and show strong amino acid transporter specificity, can be labelled easily with 18F-fluorination, and should find wide application in the development of previously unavailable PET imaging probes for clinical diagnosis.85 

The enzyme 3-deoxy-d-arabino-heptulosonate 7-phosphate synthase is the first one which is responsible for the synthesis of aromatic amino acids in bacteria and plants, and a potential target for the development of antibiotics and herbicides. The enzyme from Mycobacterium tuberculosis displays an allosteric regulation with three interdependent allosteric binding sites and a ternary allosteric response to combinations of the aromatic amino acids L-Trp, L-Phe, and L-Tyr. The binding mode of d-amino acids was investigated using X-ray crystallography, site directed mutagenesis, and isothermal titration calorimetry, and the key differences in the binding mode were identified.86 

The syntheses and photophysical characterization of three novel cyanotryptophans (6- and 7-cyano Trp, and N-methyl-7-cyano Trp) and their efficient incorporation into proteins as fluorescent probes are described. 6-Cyano Trp was also incorporated into two DNA binding proteins and an RNA recognition motif. The novel cyanotryptophans can be used for studying conformational changes of proteins and DNA–protein interactions.87 

The controlled assembly of Trpzip β-hairpins was investigated. The results show how sensitive is peptide and protein aggregation to minor sequence variation and that it is possible to use a photolabile non-natural amino acid analogue of Lys to tune the rate of peptide aggregation and to control fibrillar structure.88 

Different functionalized azabicyclo[X.Y.0]alkanone derivatives of amino acids have been developed via electrophilic transannular cyclizations of 8-, 9-, and 10-membered unsaturated macrocycles to form 5,5-, 6,5-, 7,5-, and 6,6-fused bicylic amino acids. Macrocycles were obtained by a sequence featuring peptide coupling of vinyl-, allyl-, homoallyl-, and homohomoallylglycine building blocks followed by ring-closing metathesis. The conformational preferences and the mechanism for the diastereoselective formation of specific azabicycloalkanone amino acids were also studied.89 

Two pyridine-linked bis(β-cyclodextrin) copper(ii) complexes have been reported that enantioselectively hydrolyse chiral esters of amino acids. It was demonstrated that the enantioselective hydrolysis was related to the cooperative roles of the intramolecular flanking chiral β-cyclodextrin cavities with the coordinated copper ion.90  Fluorescent sensors based on semiconductor quantum dots have been investigated for enantioselective molecular recognition. A versatile fluorescent sensor encapsulated with cyclodextrin clicked silica via layer-by-layer modification was reported for chiral selection of amino acids.91 

New stevia amino acid sweeteners, stevia Gly ethyl ester and stevia L-Ala methyl ester were synthesised and characterized. The novel sweeteners were stable in acidic, neutral, or basic aqueous solutions.92 

Mycosporine-like amino acids (MAAs) are produced by organisms that live in environments with high volumes of sunlight, usually in marine environments. They are produced by a large variety of microorganisms including cyanobacteria, lichen, fungi, and marine micro- and macroalgae. They are water soluble, small intracellular secondary metabolites that absorb ultraviolet radiation, but their function is not limited to sun protection. MAAs and their derivatives are natural antioxidants, show immunomodulatory effects and anti-inflammation activities, promote wound healing, and inhibit collagenase.93,94  Over 30 MAAs have been resolved and all contain a central cyclohexenone or cyclohexenimine ring and a wide variety of substitutions. The biosynthesis of MAAs in the cyanobacterium Arthrospira sp. CU255 was investigated. The results indicate that the studied cyanobacterium may protect itself by synthesizing mycosporine-Gly, the UV-absorbing/screening compound as important defense mechanism.95  Identification of novel MAAs is important from a biotechnology perspective, and the identification of the genes responsible for biosynthesis of MAA is vital for future genetic engineering. A study confirmed first a biosynthetic gene cluster for MAA from Gram-positive bacteria. Structure elucidation revealed that the novel MAA is mycosporine-Gly-Ala, which substitutes L-Ala for the L-Ser of shinorine.96 

Although a large number of methods are available for the synthesis of α-amino acids, the development of new, highly efficient, and enantioselective methods for the synthesis of many of the natural and unusual α-amino acids has been a long-standing goal of synthetic chemists.

Much effort has focused on the enantioselective version of Strecker reaction resulting in the development of an assortment of effective metal-based and metal-free catalysts. Derivatives of α-amino acids have been synthesized via enantioselective addition of masked acyl cyanides to imines. This method works for the free amino acids rather than their N- or C-protected derivatives.97  Spontaneous formation of an enantioenriched α-amino nitrile which is a chiral precursor for Strecker amino acid synthesis was produced without addition of any chiral substances.98 

Asymmetric synthesis of a wide variety of α-amino acids was achieved by alkylations of chiral or achiral Ni(ii) complexes of Gly Schiff bases. Origin of diastereo-/enantioselectivity in the alkylations reactions, aspects of practicality, generality, and limitations of this method is critically discussed.99  A broad review discussed the data in the literature on asymmetric synthesis of α-amino acids via Michael addition reactions involving Ni(ii)-complexes and the practical aspects of the methodology.100 

One of the most recent developments in asymmetric catalysis is to employ two or more catalysts under one-pot reaction conditions. A cooperative dual-catalytic protocol relying on the catalytic ability of dirhodium carbenoid (derived from rhodium(ii) tetracarboxylate and a diazo compound) and a chiral spirophosphoric acid in an asymmetric N–H insertion reaction was investigated.101  The development of enantioselective C–H activation reactions by desymmetrization have been progressed in the past decade. Enantioselective C–H olefination of α-hydroxy and α-amino phenylacetic acids was achieved by kinetic resolution with the use of palladium(ii) catalyst.102 N-Aryl glycine esters with terminal alkynes were synthesized via enantioselective cross-dehydrogenative coupling by Cu catalyst.103  Asymmetric synthesis of N-Boc-(R)-silaproline was achieved via Rh-catalyzed intramolecular hydrosilylation of dehydroalanine and continuous flow N-alkylation with excellent yield and enantioselectivity.104 

Due to its pharmaceutical importance, (1R,2S)-1-amino-2-vinylcyclopropane-carboxylic acid was prepared by advanced asymmetric procedure. The target amino acid was achieved via two-step SN2 and SN2′ alkylation of novel axially chiral Ni(ii) complex of Gly Schiff base with excellent yields and diastereoselectivity.105  A chiral amine-catalyzed highly stereoselective vinylogous allylic–allylic alkylation of Morita–Baylis–Hillman carbonates with olefin azlactone gives access to chiral multifunctional acyclic α-amino acid derivatives or protected cyclic quaternary α-amino acids.106 

Despite significant advances in synthetic methodology, the efficient synthesis of enantiopure α-amino acids carrying complex side chains remains challenging. Palladium-catalyzed bidentate auxiliary-directed C–H functionalisation reactions for α-amino acid substrates have been investigated. A variety of α-amino acid precursors can undergo multiple modes of C(sp(3))–H functionalisation, including arylation, alkenylation, alkynylation, alkylation, alkoxylation, and intramolecular aminations at the β, γ, and even δ positions to form new α-amino acids.107  A highly efficient and enantioselective synthesis of γ-lactams and γ-amino acids by Rh-catalyzed asymmetric hydrogenation has been developed from cyano-substituted acrylate esters under mild conditions.108  Highly enantioselective synthesis of 5-substituted γ-lactams was achieved by Ir-catalyzed sp(3) C–H alkylation of γ-butyrolactam with alkenes. 5-substituted γ-lactams were readily converted into chiral 4-substituted γ-amino acids.109 

Enantioselective aza-Diels–Alder reaction of oxodiazenes with α-chloroaldehydes by NHC catalysis leads to derivatives of α-amino acids with excellent yield and enantioselectivity.110  The difficult to access δ(3)-amino acids can be produced by asymmetric cyclopropanation of conjugated cyanosulphones using a novel bifunctional organocatalyst.111 

Phenylalanine ammonia lyases catalyze the synthesis of amino acids by 4-methylideneimidazole-5-one cofactor independent pathway.112  Phenylalanine ammonia-lyase catalyzed the deamination of an acyclic amino acid. Enzyme mechanistic studies were achieved by a novel microreactor filled with magnetic nanoparticles.113 

The simplest and minimal modification of a single amino acid or peptide bonds is represented by N-methylation. It is crucial to provide optically pure N-methyl-amino acids and N-methylated peptides structural and conformational studies. A review focuses on the results obtained in the field of chemical synthetic methodologies for the N-methylation of amino acids in the last decade.114  Another review specifically examines recent advances in substrate-controlled asymmetric reactions induced by the chirality of α-amino acid templates in natural product synthesis research and related areas.83 

Studies on the diastereo- and enantioselective syntheses of anti-β-hydroxy-α-amino acid esters using transition-metal-chiral-bisphosphine catalysts have been summarized. A variety of transition metals (Ru, Rh, Ir, and Ni) in combination with chiral bisphosphines, worked well as catalysts for the direct anti-selective asymmetric hydrogenation of α-amino-β-keto ester hydrochlorides.115  1H-Imidazol-4(5H)-ones are introduced as novel nucleophilic α-amino acid equivalents in asymmetric synthesis. N,C(α),C(α)-trisubstituted α-amino acid derivatives were synthesized by using 1H-imidazol-4(5H)-ones.116  Esters of N-methyl-Pro and N-allyl-Pro were prepared and used for synthesis of chiral triazine based coupling reagents. The coupling reagent can be applied in the enantioselective incorporation of l- and d- amino acids directly from racemic substrate.117 

Amino acids are produced at the multi-million-ton-scale with the economical fermentative production. Productivities of amino acid producing strains, e.g. Corynebacterium glutamicum and Escherichia coli are constantly improved by metabolic engineering. A review summarizes the new pathways for amino acid production as well as fermentative production of non-native compounds derived from amino acids or their metabolic precursors.118  The direct fermentation is an attractive method for production of L-Serine from sugars. However, low L-Ser production and superfluous by-product accumulation limit the industrial production on large scale. Several modifications demonstrated that combined metabolic and bioprocess engineering strategies could minimize by-product accumulation and improve L-Ser productivity.119  The fermentative production of L-Thr and L-Ile with Corynebacterium glutamicum is usually accompanied by the by-production of L-Lys. An effective strategy was developed to reduce L-Lys by-production.120 

A new system is used for continuous and direct production of poly-(γ-Glu) in a hybrid reactor system that integrated conventional fermentative production step with membrane-based downstream separation and purification. This integrated system provides compact, flexible, eco-friendly, and largely fouling-free ensuring steady and continuous production of poly-(γ-Glu) directly from a renewable carbon source.121  A review focuses on the current trends and future perspectives of microbial poly(ɛ-L-Lys) and contributes to the development of this novel homopoly(amino acid) and serve as a basis of studies on other biopolymers.122 

Many α-amino acids and their derivatives are found in addition to the 20 natural α-amino acids in various natural products. Constituting the structural basis of peptides and proteins, α-amino acids continue to be at the forefront of chemical synthesis research.

The Strecker amino acid synthesis is an organic reaction used to convert an aldehyde or ketone and a primary amine or ammonia to an α-amino acid using a metal cyanide, acid catalyst, and water. The Strecker synthesis has long been considered for the synthesis of α-amino acids, but highly enantioenriched α-amino acids through this method remains a puzzle. Since nowadays enantiomeric purity is one of the major issues in α-amino acid synthesis, tremendous efforts have been put into the development of asymmetric versions of Strecker's protocol. Near enantiopure l- and d- amino acids have been produced by asymmetrical Strecker reaction of an achiral imine at a single-crystal face.123  Replication of chiral α-amino acids has been achieved in combination with the asymmetric induction, amplification, and multiplication of their own chiral intermediates (l- and d-aminonitriles) in the solid-phase via Strecker reaction between three achiral components.124 

A novel method for the synthesis of l- and d-amino α-amino acids by a Ni-catalyzed reductive cross-coupling of Ser/homo-Ser-derived iodides with aryl/acyl/alkyl halides is described. This method provides convenient access to varieties of enantiopure and functionalized amino acids.125  The reactivity of Pd-catalysts in the widely used C–H functionalisation strategies were investigated. It was demonstrated that the amino acid ligand plays crucial roles in the ligand-assisted Pd(ii)-catalyzed C–H activation.126 

Novel nonsymmetrically substituted N-protected β,β-diaryl-α-amino acids and esters have been produced through asymmetric hydrogenation of tetrasubstituted olefins by catalytic, asymmetric, and stereodivergent synthesis.127  Non-natural aliphatic α-amino acids were synthesized via asymmetric hydrogenation of β-alkyl (Z)-N-acetyldehydroamino esters employing rhodium-Duanphos complex catalyst with excellent yields and high enantioselectivities. An efficient approach for the synthesis of the intermediate of the orally administered anti-diabetic drugs Alogliptin and Linagliptin in the DPP-4 inhibitor class was also developed.128 

Derivatives of α-amino acids have been prepared via enantioselective additions of masked acyl cyanide reagents to N-Boc-aldimines using modified cinchona alkaloid catalyst in excellent yields and high enantioselectivities.97  Three novel types of regio- and enantioselective multiple amino-functionalisations of terminal alkenes via cascade biocatalysis have been produced to get α-amino acids. The modular approach for engineering multi-step cascade biocatalysis is useful for developing other new types of one-pot biotransformations for chemical synthesis.129 

N-alkyl aminomalonates undergo a fast and selective intramolecular C→N acyl rearrangement reaction in the presence of a strong base leading to N-protected glycinates in excellent yield. The reaction can be applied for the preparation of nonproteinogenic tertiary and quaternary N-alkyl α-amino acids in a very simple and reliable way.130 

Highly functionalised azatricyclononanes were achieved by uncatalysed cycloaddition of substituted diaryldiazo compounds onto bicyclic unsaturated lactams derived from derivatives of pyroglutamic acid (Glp). The products provide rapid access to conformationally constrained amino acids and their analogues.131 

The presence of NH function in “N–H” – Ni(ii) complexes of Gly Schiff bases does not interfere with the homologation of the Gly residue. The practical application of these NH-type complexes were applied for the general synthesis of α-amino acids and in the asymmetric synthesis of various β-substituted Glp via Michael addition reactions with chiral Michael acceptors.132 

Amino acids and their derivatives play a central role in the design of life. The relevance of the 20 proteinogenic l-amino acids as building blocks in peptides and proteins is self-evident, but the class of quaternary α-amino acids also called α,α-disubstituted amino acids has attracted the interest of organic chemists due to their biological importance. They play a crucial role and increasing interest in the development of unnatural peptides and peptidomimetics as therapeutics agents. Quaternary α-amino acids are present in a family of peptide antibiotics of fungal origin that could destabilize the membrane by formation of pores in the bilayer membrane. Quaternary α-amino acid residues such as α-aminoisobutyric acid and isovaline are present in peptaibol and peptaibiotics which constitute a family of peptide antibiotics of fungal origin. Peptaibiotic could destabilize the membrane by formation of pores in the bilayer membrane.

Optically active α,α-disubstituted α-amino acids have generated increasing interest in biology, pharmacology, and in chemistry. Some of these compounds occur naturally or are structural components of natural products that have specific properties. The inclusion of α,α-disubstituted α-amino acids in a peptide may affect its secondary or tertiary structure by inducing a particular conformation.

Biotechnological and biomedical chemical processes often use α,α-disubstituted α-amino acids. These compounds contain a quaternary stereo enter, which is challenging for stereoselective synthesis. The additional alkyl substituent at α-carbon could inhibit the free rotation of the peptide backbone leading to unique folding when incorporated into peptides. They can restrict the conformational freedom of the peptides and provide key information concerning the conformation responsible for biological recognition. Moreover, peptides containing α,α-disubstituted amino acid residues also tend to have an increased hydrophobicity, as well as an increased stability toward both chemical and enzymatic degradations.

The first asymmetric synthesis of α-allyl-α-aryl α-amino acids is reported via a three-component coupling of α-iminoesters, Grignard reagents, and cinnamyl acetate. The α-allyl group offers to generate further α-amino acid structures as exemplified by ring closing metathesis to generate a higher ring homologue of α-aryl-Pro.133  Asymmetric synthesis of α-allyl-α-aryl α-amino acids has also been achieved by tandem N-alkylation/π-allylation. Analogues of homo-Tyr and homo-Glu have been synthesized. This strategy can also be used to the asymmetric synthesis of acyclic and cyclic amino acid derivatives and higher ring homologues of enantioenriched α-substituted Pro.134  Allyl cyanate-to-isocyanate rearrangement (Ichikawa rearrangement) can be the key step to achieve α,α-disubstituted α-amino acids.135  A Brønsted acid accelerated Pd-catalyzed direct asymmetric allylic alkylation of azlactones with simple allylic alcohols leads to quaternery allylic amino acid derivatives under mild reaction conditions in excellent yields and good enantioselectivities.136  Asymmetric allylic alkylation of azlactones with 4-arylvinyl-1,3-dioxolan-2-ones with Pd catalyst was developed leading to “branched” chiral α-amino acids with vicinal tertiary and quaternary stereocenters in high yields. Studies on mechanism revealed that the formation of a hydrogen bond between the Pd–allylic complex and azlactone isomer is responsible for the excellent regioselectivities.137 

An electrophilic alkynylation of azlactones can lead to C(α)-tetrasubstituted α-amino acid derivatives in short reaction times (Scheme 4).138  An enantioselective synthesis of α,α-disubstituted α-amino acids have been developed via direct catalytic asymmetric addition of acetonitrile to α-iminoesters bearing an N-thiophosphinoyl group.139 

(R) or (S)-α-benzyl-β-azido-Ala, α-benzyl-β-(1-pyrrolidinyl)-Ala, α-benzyl-β-(1-piperidinyl)-Ala, and α-benzyl-β-(4-morpholinyl)-Ala have been replaced by Phe in deltorphin I analogues. The potency, selectivity, affinity, and conformational behaviour were investigated.140  Asymmetric synthesis of α,α-disubstituted amino acids using regio- and stereocontrolled 1,3-dipolar cycloaddition reactions with vinyl ethers and α,α-dialkylketonitrones have been reported.141  A series of α,α-disubstituted amino acid derivatives have been synthesized with a novel method using silver carbonate, diaryliodonium bromides as aryl sources, and azlactones.142  A review provides an overview on the synthesis and applications of α-quaternary α-ethynyl α-amino acids from 1977 to 2015.143  Alkylation of deprotonated α-aminonitriles derived by the Strecker reaction from (4S,5S)-5-amino-2,2-dimethyl-4-phenyl-1,3-dioxane leads to a series of α-quaternary arylglycines in high optical purity (Scheme 5).144 

Derivatives of α,α-disubstituted α-amino acid were produced by asymmetric Michael addition of azlactones to α,β-unsaturated trichloromethyl ketones with the use of commercially available quinine-derived thiourea catalyst in high yields and with excellent stereoselectivities.145  Pseudoephedrine-directed asymmetric α-arylation of α-amino acid derivatives leads to quaternary amino acids.146  Enantio-enriched α,α-dialkyl substituted α-nitroacetates were produced by an enantioselective conjugate addition of α-alkyl substituted α-nitroacetates to phenyl vinyl selenone with a use of a novel Cinchona alkaloid catalyst. The Michael adducts were converted to various cyclic and acyclic quaternary α-amino acids.147  All-carbon quaternery stereogenic centres were achieved by asymmetric Michael reaction of nitroalkanes and β,β-disubstituted α,β-unsaturated aldehydes catalyzed by diphenylprolinol silyl ether. The reaction mechanism is also discussed.148  Asymmetric Michael addition of α-fluoro-α-nitro esters to nitroolefins leads to quaternary α-fluoro-α-substituted amino acids in excellent chemical yields and with high enantioselectivities.149  Quaternary α-nitroesters have been achieved by enantioselective Michael addition of tertiary α-nitroesters to β-unsubstituted vinyl ketones in the presence of an L-tert-Leu-derived squaramide as organocatalyst. The products can be transformed to quaternary α-amino acids.150 

Enantioselective C-2- and C-4-selective γ-additions of oxazolones to 2,3-butadienoates catalyzed by phosphine lead to 2-aryl-4-alkyloxazol-5-(4H)-ones that provided rapid access to optically enriched α,α-disubstituted α-amino acid derivatives.151  A chiral phosphine can catalyze the stereoconvergent γ-additions of racemic nucleophiles (racemic heterocycles) to racemic electrophiles. This method enables the synthesis of protected α,α-disubstituted α-amino acid derivatives in good yield, diastereoselectivity, and enantioselectivity.152 

An enantioselective and diastereoselective asymmetric aldol reaction by memory of chirality starting from L-Ala provides a rapid access to enantiopure β-hydroxy quaternary α-amino acids in three steps.153 

A highly stereoselective multicomponent cascade reaction of ketones or unprotected ketohexoses and unprotected amino acids leads to quaternary stereogenic centre.154  The pyridoxal-5′-phosphate-dependent L-serine hydroxymethyltransferase from Streptococcus thermophilus was engineered to achieve the stereoselective synthesis of a broad structural variety of α,α-dialkyl-α-amino acids.155 

Pyrrolidine-2-carboxylate esters substituted in the 3-, 4- or 5-positions were converted to their N′-aryl urea derivatives by intramolecular diastereoselective arylation. After several reaction steps, a range of enantiopure and enantioenriched quaternary α-aryl Pro derivatives have been achieved.156  1,4-Substituted triazole oligomers are made from derivatives of quaternary amino acids that present a conformational behaviour with similarities to that of natural peptides.157 

A new and efficient method has been developed for the synthesis of racemic-protected α-ethynyl Phe starting from DL-2-benzyl-Ser in ten steps. The absolute configurations of the separated enantiomers were also determined.158 

Among the numerous general strategies that have commonly been used for the syntheses of α-amino acids, there are many that apply de novo synthesis focused on enantioselective bond construction around the α carbon and others that consider conversion of existing precursors of α-amino acids carrying suitable functional groups on side chains. A strategy based on the selective functionalisation of side chain C–H bonds of various readily available precursors of α-amino acids may provide a straightforward and broadly applicable synthesis and transformation of α-amino acids. A systematic investigation of palladium-catalyzed bidentate auxiliary-directed C–H functionalisation reactions for α-amino acid substrates was carried out. The palladium-catalyzed auxiliary-directed sp(3) C–H functionalisation reduces the synthetic difficulty for many α-amino acids leading to a wide range of β-monosubstituted α-amino acids that also can undergo further C–H functionalisation at the β-methylene position to generate various β-branched α-amino acids in a stereoselective way.107  Pd-catalyzed alkylation of methylene C(sp(3))–H bonds of aliphatic quinolyl carboxamides with α-haloacetate and methyl iodide have been applied in the stereoselective synthesis of various β-alkylated α-amino acids. This method also provides a convenient and powerful way to site-selectively incorporate isotopes into the carbon scaffolds of amino acids.159 

Cyclopropane α-amino acid esters bearing quaternary carbon centres have been synthesized by cyclopropanation of 2-aminoacrylates with N-tosylhydrazones in high yields and diastereoselectivities.160 

The literature data on asymmetric synthesis of α-amino acids via Michael addition reactions involving Ni(ii)-complexes is reviewed. The discussion is divided into two groups dealing with applications of: (a) Ni(ii)-complexes of Gly as C-nucleophiles and (b) Ni(ii)-complexes of dehydro-Ala as Michael acceptors. The review focused on the practical aspects of the methods.100  Asymmetric synthesis of α-allyl-α-aryl α-amino acid esters was developed. The syntheses of analogues of homo-Tyr and homo-Glu and derivatives of cyclic α-amino acids were completed.134 

Synthesis of γ-amino acids by Rh-catalyzed asymmetric hydrogenation of cyano-substituted acrylate esters has been developed by the use of Rh-(S,S)-f-spiroPhos complex with high yield and enantioselectivity.108 

It is known for more than 40 years that trouble in metabolism of branched-chain amino acids leads to higher levels of Ile, Leu, and Val that is strongly associated with higher risk of type 2 diabetes, but it is not known whether this association is causal. Genome-wide association studies coupled with large-scale metabolomic measurements were used to investigate the etiologic relationship between metabolism of branched-chain amino acid and type 2 diabetes.161  A review on branched-chain amino acids in metabolic signalling and insulin resistance concluded that increased levels of branched-chain amino acids are more likely to be a marker of loss of insulin action and not causative.162  Plasma branched-chain amino acids are positively associated with incident diabetes and insulin resistance163  and underlying metabolic abnormalities.164  It was also shown that high consumption of branched-chain amino acids is associated with an increased risk of type 2 diabetes.165  Recently, a Mendelian randomisation study using genetic variants associated with circulating branched-chain amino acid levels and insulin resistance as instrumental variables suggests that higher levels of branched-chain amino acids do not have a causal effect on insulin resistance while increased insulin resistance drives higher circulating fasting branched-chain amino acid levels.166 

Amino acids in plasma are valuable biomarkers to determine increased risk of mortality in patients with end-stage liver disease. Val concentrations and constellations composed of branched-chain and aromatic amino acids were strongly associated with prognosis.167  Elevated plasma levels of branched-chain amino acids also are associated with a greater than twofold increased risk of future pancreatic cancer diagnosis.168 

Branched chain amino acids (Ile, Leu, Val) participate in protein synthesis in animals and humans and regulate many key signalling pathways that connect many diverse physiological and metabolic roles. Abnormally elevated levels of branched chain amino acids in the blood seem to be a good biomarker for the early detection of obesity, diabetes, and other metabolic diseases. A review provides some insights into these novel metabolic and physiological functions of branched chain amino acids.169 

Conflicting data exist on the impact of dietary and circulating levels of branched chain amino acids on cardiometabolic health and it is unclear to what extent these relations are mediated by genetics. In a cross-sectional study, female twins were examined. The results showed that higher intakes of branched chain amino acids were associated, independently of genetics, with lower insulin resistance, inflammation, blood pressure, and adiposity-related metabolites.170  The associations between intakes of amino acids with known mechanistic links to cardiovascular health and direct measures of arterial stiffness, central blood pressure, and atherosclerosis were examined. It was found that higher intakes of Leu, Glu, and Tyr were most strongly associated with pulse wave velocity. The data provided evidence that intakes of several amino acids are associated with cardiovascular benefits beyond blood pressure reduction.171  A pharmacometabonomic study showed that branched-chain amino acids are predictors for individual differences of cisplatin nephrotoxicity in rats. This study could provide new insight into cisplatin nephrotoxicity and may help expedite personalized medicine of cisplatin or other antitumour drugs in future clinical studies.172 

Production of syn-selective β-branched α-amino acids is based on the alkylation of Gly imine esters with secondary sulphonates. The optimized conditions enabled a straightforward preparation of a number of β-branched α-amino acids.173 

Strategies to protect bioactive peptides from serum proteolytic degradation include incorporation of halogens into natural or nonnatural amino acids. Among the halogen-substituted amino acids, the most important ones have fluorine in the molecule. Fluorine became the “second-favourite heteroatom” after nitrogen in drug design due to the special properties of fluorine–containing compounds.174  It was empirically demonstrated that the presence of fluorine in compounds results in metabolic stability leading to improved bioactivity and bioavailability.175–177  Since compounds with fluorine mimic their parent compounds with hydrogen well, they fit into the same enzyme binding site.178  The highly electronegative fluorine atom forms a highly polarized bond of extraordinary strength with carbon.178 

Combination of the unique physical and chemical properties of fluorine with amino acids represents a new approach for the design of biologically active peptides with improved pharmacological properties.179  Due to their unique electronic properties, fluorinated amino acids have huge effects on protein stability, protein–protein as well as ligand–receptor interactions, and the physical properties of protein- or peptide-based materials.176,178,179 

In order to alter distinct properties of peptides and proteins (e.g. hydrophobicity, acidity/basicity, and conformation), it is an efficient strategy to incorporate amino acids with fluorinated side chains. Proteins and complexes with proteins can be stabilized by highly fluorinated amino acids via enhanced hydrophobicity, and provide novel methods for identification of specific molecular events in complex solutions.178,180  It was shown that specific incorporation of fluorinated amino acids into proteins can experimentally distinguish cation–π interactions between the π systems of aromatic residues and the positively charged portion of phospholipid head groups from membrane insertion of the aromatic side chains of amino acids in proteins which hydrophobically interact with lipid tails. Fluorinated aromatic amino acids destabilize the cation–π interactions by altering electrostatics of the aromatic ring, whereas their increased hydrophobicity enhances membrane insertion.181  In the contrary, the majority of anion–π interacting residues are located in regions with helical secondary structure. The influence of anion–π interactions should not be neglected in supramolecular chemistry.182  Recently, it was found that incorporation of pentafluoro-Phe into human growth hormone releasing hormone analogues suppressed the growth of different human tumours in vitro and in vivo.183 

Although 4-F-Thr is the only naturally occurring fluoro-amino acid to date,178  many synthetic methods are developed for the syntheses of mono- or poly-fluoro and other halogen amino acids and derivatives.

As one of the possible fluorinated analogues of proteinogenic Ile, two diastereoisomers of 5,5,5-trifluoroisoleucine ((2S,3S)-5-F3Ile and (2R,3S)-5-F3-allo-Ile) have been synthesized in enantiomerically pure form and the relationship of their side chain hydrophobicity and α-helix propensity were examined.184  γ-(4S)-Trifluoromethyl Pro was synthesised by a modified literature protocol with improved yield and a multigram scale, and the conformational properties of the amide bond were characterised. Replacement of native Pro by γ-trifluoromethyl Pro in the peptide antibiotic gramicidin S was shown to preserve the overall amphipathic peptide structure.185  The synthesis of (2S,4R)- and (2S,4S)-iodophenyl ethers of hydroxy-Pro has been described. These amino acid derivatives are capable of modification via rapid, specific Suzuki and Sonogashira reactions in water and provide new functionalisation for peptides with great control of conformation.186 

Fluorinated Phe analogues (4-fluoro-, 2,4-difluoro-, or 4-trifluoromethyl-Phe) were incorporated into endomorphin-2-analogues. They showed strong antinociceptive effect indicating that they were able to cross the blood–brain barrier.187  Two all-cis, 2,3,5,6-tetrafluorocyclohexyl (S)-phenylalanines having the 2,3,5,6-tetrafluorocyclohexyl group in meta and para positions at the aromatic ring of Phe have been synthesized as novel fluorinated α-amino acids for peptide synthesis.188  A stereoselective method has been developed for the synthesis of new β-fluorinated 2-aminocyclohexanecarboxylic acid derivatives with fluorine at position 4 of the ring.189  Substituted anti-β-fluorophenylalanines were produced from the corresponding enantiopure α-hydroxy-β-amino esters using a stereospecific XtalFluor-E promoted rearrangement procedure (Scheme 6) in good yield and high diastereoisomeric purity.190 

Optically pure amino acids that have easily replaceable functional groups at the ω-positions are highly important synthetic targets, as the functional group at the ω-position can be modified to the required group, which provide a series of non-natural amino acids. ω,ω-difluoroalkyl amino acid derivatives were synthesized by oxidative desulphurization-fluorination reactions of suitable arylthio-2-phthalimido butanoates and pentanoates. Mainly α,ω-polyfluorinated amino acid derivatives were formed by additional sulphur-assisted α-fluorination.191  Fmoc-perfluoro-tert-butyl-Tyr was synthesized in two steps from commercially available Fmoc-4-NH2-Phe via diazotization followed by diazonium coupling reaction with perfluoro-tert-butanol.192 

Fmoc-, Boc-, and free (2S,4R)- and (2S,4S)-perfluoro-tert-butyl 4-hydroxy-Pro have been synthesized in 2-5 steps. The perfluoro-tert-butyl group was incorporated with perfluoro-tert-butanol in a Mitsunobu reaction. Peptides containing these amino acids were detected by (19)F NMR, suggesting their use in probes and medicinal chemistry.193  Trifluoromethyl-substituted cyclopropane α-amino acids could be obtained with a method that starts with cyclopropanation of trisubstituted olefinic azlactones with a stock solution of CF3CHN2 in CH3CN (Scheme 7). Azlacton rings having trifluoromethyl-substituted cyclopropanes are produced in good to high yields and diastereoselectivities.194 

Fluorinated homoproline derivatives bearing three stereogenic centres are achieved by intramolecular aza-Michael reaction from 2-p-tolylbenzyl carbanions as a source of chiral benzyl nucleophiles. The selectivity of the cyclization process can easily be set by changing the reaction conditions.195  A radical based synthesis of a variety of protected enantiopure fluorinated derivatives of α-amino acids is described that lead to mercapto-α-amino acids (Scheme 8) for native chemical ligation.196 

Pd(ii)-catalyzed fluorination of inactivated methylene C(sp(3))–H bonds led to diastereoselective β-fluorinated α-amino acids and their derivatives.197  Highly site-selective and diastereoselective fluorination on inactivated sp(3) carbons were achieved by the use of palladium acetate as the catalyst leading to derivatives of β-fluorinated amino acids.198  It was shown for the first time that a quinoline-based ligand contributes to β-C(sp(3))–H fluorination. Nonnatural enantiopure fluorinated α-amino acids were obtained through sequential β-C(sp(3))–H arylation and subsequent stereoselective fluorination from L-Ala.199 

Sometimes enzymes are applied as catalysts for the production of fluorinated amino acids or their derivatives. The production of nitro analogues of 4-F-DL-Trp and 5-F-L-Trp in large scale is based on the use of TxtE-based class I cytochrome P450 enzyme;200  this was the first biological nitration procedure. Arabidopsis thaliana (AtPAL2) proved to be a very good catalyst for the formation of (S)-3-F-Phe, (S)-4-F-Phe and (S)-2-Cl-Phe.201  These halogenated amino acids are valuable building blocks for the formation of various drug molecules. Many physiological and pathological processes (e.g. atherosclerosis and tumorigenesis) involve matrix metalloproteinases that belong to a group of zink-dependent endopeptidases located in the extracellular matrix. Fluorinated analogues of α-aminocarboxylic and α-amino hydroxamic acid-based matrix metalloproteinase inhibitors showed comparable or superior inhibiting potencies as compared to their non-fluorinated analogues. The corresponding α-aminocarboxylic acid derivatives are less potent than the α-amino hydroxamic acid analogues or were inactive.202 

Multi-substituted β-lactams can be used as building blocks for the construction of β-amino acids. A review from Tarui summarizes the direct functionalisation of fluoro-β-lactams.203  The introduction of an electronegative fluorine atom to a β-lactam ring gave the corresponding fluoro-β-lactam that can be used for the preparation of electrophilic β-lactams, and these compounds can be converted to fluorinated β-amino acids.203 

α-Fluorinated β-amino thioesters were obtained in high yields and stereoselectivities under mild reaction conditions by organocatalyzed addition reactions of α-fluorinated monothiomalonates to N-Cbz- and N-Boc-protected imines. The addition products were used for coupling-reagent-free peptide synthesis.204  A diastereoselective addition of fluoroacetate or α-alkylated fluoroacetate to N-tert-butylsulfinyl imines results in α-fluoro-β-amino acids containing fluorinated quaternary stereogenic carbon centres (Scheme 9) with very good yields and high diastereoselectivities.205 

A diastereoselective Mannich-type reaction of α-alkyl, α-aryl, and α-vinyl fluoroacetates with N-tert-butylsulfinyl imines provides a broad range of highly functionalized β-amino acids containing α-fluorinated quaternary stereogenic carbon centres. The stereochemical outcome of the present simple reaction is highly dependent on the steric and electronic properties of the fluorocarbon nucleophiles.206  Enantiomerically pure (R)- or (S)-configured 3-amino-4,4,4-trifluorobutanoic acids were obtained by asymmetric Mannich reactions of malonic acid derivatives and (SS)-N-(tert-butanesulfinyl)-3,3,3-trifluoroacetaldimine with the use of inorganic or organic base as catalyst.207 

Despite the growing demand for enantioenriched fluorine containing small molecules, α- and β-fluorinated carbonyl compounds have been neglected in direct enolization chemistry because of the competing and dominating defluorination pathway. Direct and highly stereoselective Mannich-type reactions of α- and β-fluorine-functionalized 7-azaindoline amides guarantee an efficient enolization, while suppressing undesired defluorination provide a series of fluorinated analogues of enantioenriched β-amino acids (Scheme 10).208 

Highly fluorinated non-natural amino acids and derivatives were produced by using oxazolone enolate for a nucleophilic substitution of highly fluorinated (hetero)arenas.209 

Fluorinated aminoanthranilamides, derivatives of non-native aromatic beta-amino acids have been developed by a highly regio-selective nucleophilic aromatic substitution of difluorinated nitrobenzoic acid. The findings opened unexplored routes to novel amino-acid structures.210 

Novel fluorinated analogues of γ-aminobutyric acid (β-polyfluoroalkyl-GABAs) have been synthesized with substituents β-CF3-β-OH, β-CF3; and β-CF2CF2H, these analogues are bioisosteres of Pregabalin (Lyrica®). Biological investigations showed that fluorinated analogues of GABA are structural but not functional analogues of GABA.211 

Chiral α-hydroxy amino acids and their derivatives play important role in preparation of pharmaceuticals and are involved in many biological processes.

β-Hydroxy-α-amino acids are important chiral building blocks in chemical syntheses and serve as precursors to many important medicines. In addition, they are also found in natural products and show anti-microbial or anti-cancer properties or inhibition of β-amyloid peptide release. Derivatives of β-hydroxy-α-amino acids have been produced by aldolization of pseudoephenamine glycinamide with lithium hexamethyldisilazide in the presence of LiCl followed by addition of an aldehyde or ketone. The stereoisomerically pure products can be transformed into β-hydroxy-α-amino acids by mild hydrolysis.212  The Cu-catalyzed asymmetric conjugate hydroboration reaction of β-substituted α-dehydroamino acid derivatives has been accomplished leading to enantioenriched syn- and anti-β-boronate-α-amino acid derivatives with excellent yields and enantioselectivities. The hydroboration products were converted into β-hydroxy-α-amino acid derivatives.213  β-hydroxy-α-amino acid derivatives have been produced by an enantioselective four-component reaction of a diazoketone, water, an aniline and ethyl glyoxylate in the presence of catalytic Rh2(OAc)4 and a chiral Brønsted acid in good yields and high diastereoselectivity and enantioselectivity.214  A review of Zhang Y. also summarizes other methods for the syntheses of β-hydroxy-α-amino acids.215 

An asymmetric aldol reaction by memory of chirality is reported with a substrate control of stereoselectivity by aldehyde. Enantiopure β-hydroxy quaternary α-amino acids have been obtained by starting from L-Ala in three steps.153 

A stereoselective synthesis of anti-β-hydroxy-α-amino acids is based on the palladium-catalyzed sequential C(sp(3))–H functionalisation by 8-aminoquinoline, followed by a previously established monoarylation and/or alkylation of the β-methyl C(sp(3))–H of Ala derivative leading to various anti-β-hydroxy-α-amino acid derivatives. β-mercapto-α-amino acids that are very important for to chemical ligation can also be produced by this practical method.216  Natural l-α-amino acids having aliphatic, aromatic, or heteroaromatic moieties were transformed to the corresponding enantiopure (R)- or (S)-α- hydroxyacids by formal biocatalytic inversion or retention of absolute configuration. The one-pot transformation was carried out by a concurrent oxidation reduction cascade in aqueous media.217  N-hydroxy-Gly, N-hydroxy-Ser, l-homoserine, and α-hydroxy-Gly have been synthesized and tested for their water-holding capacities. Some predictive ‘rules’ for further design and refinement of chemical structures also have been developed.218 

Threonine aldolases are very stereoselective for α-carbon and catalyze the pyridoxal phosphate-dependent condensation between amino acids and aldehydes. A review published the mechanism of reactions by threonine aldolases.69  Threonine aldolases also catalyse the unnatural aldol condensation with Gly to produce valuable β-hydroxy-α-amino acids. A review by Fesko summarizes the techniques and enzyme engineering and mutagenesis studies.219  Natural threonine aldolases were also used for the direct biochemical synthesis of tertiary α-amino acids. The novel l- and d-threonine aldolases catalyze the asymmetric synthesis of β-hydroxy α-methyl- and α-hydroxymethyl-α-amino acids with perfect enantioselectivity at α-carbon.220 

A novel stereoselective synthesis of all four isomers of β- and γ-hydroxy α-amino acids was achieved. The strategy is based on enzymatic kinetic resolution and cyanate-to-isocyanate rearrangement as key steps. The proper choice of the starting hydroxyacid, the course of kinetic resolution, and the stereospecific sigmatropic rearrangement step resulted in stereo control with full chirality transfer.221  Phosphine ligand stabilized air-stable Cu(i) complexes have been successfully used to functionalize the aromatic aminobenzoic acids in a chemoselective manner without protection and deprotection strategy under mild reaction conditions. This chemoselective carbene insertion into –NH bond over –COOH and –OH bonds leads to a wide range of carboxy and hydroxy functionalized α-amino esters.25 

Stereoselective synthesis of analogues of (2S,3R)-α-hydroxy-β-amino acid has been reported by Cu(i)-catalyzed reactions of (R)-glyceraldehyde acetonide and dibenzylamine with terminal alkynes leading to the corresponding (2S,3R)-α-amino alcohols with good-to-excellent diastereoselectivity. Subsequent chemical transformations provided easy access to alkynyl side-chain containing (2S,3R)-α-hydroxy-β-amino acids.222 

Substituted α-hydroxy-β-amino amides have been synthesized by a carbamoyl anion-initiated cascade reaction with acylsilanes. A series of α-aryl-α-hydroxy-β-amino amides has been synthesized in high yields with excellent diastereoselectivities.223  A new method is developed for direct access to derivatives of isoserine from simple imines in a four-step, one-pot reaction. The strategy employs a hypervalent iodine(iii)-catalyzed bromination/rearrangement/cyclization cascade reaction that leads to a broad range of structurally different lactams from cheap and easily available imides. This cascade reaction is furthermore extendable by an in situ ring-opening reaction, giving direct access to α-hydroxy β-amino acids from simple imines in a four-step, one-pot reaction (Scheme 11).224 

Two isomeric forms of Hyp that is found in collagen and few other extracellular proteins (trans-4-hydroxy-L- and trans-3-hydroxy-L-Pro) play an important role in collagen synthesis and thermodynamic stability of the triple-helical conformation of collagen and associated tissues. Elevated level of Hyp is observed in several disorders, while its decreased level is a marker of poor wound-healing. A review summarizes the potential use of Hyp as a biochemical marker.225  An enzymatic method can be used to measure the content of T4LHyp (Hyp 2-epimerase) in the acid-hydrolysate of collagen or blood plasma.226 

To date, there are few general methods that describe the O-arylation of L-Ser. This transformation has been achieved by nucleophilic aromatic substitution, but this protocol is limited by the use of strong bases (NaH and KHMDS) and the need for 1-fluoro-2-nitrobenzene substrates. An effective and practical protocol for O-arylation of β-hydroxy-α-amino acids (Thr and Ser) was achieved via Chan–Lam cross-coupling leading to novel unnatural derivatives of β-aryloxy-α-amino acids. This new Cu(ii)-catalyzed transformation involves mild conditions and is well-tolerated with a variety of protected (Boc-, Cbz-, Tr-, and Fmoc-) Ser and Thr derivatives and various potassium organotrifluoroborates and boronic acids.227  A synthetic approach towards (2S,3S)-3-hydroxyleucine that can be found in an increasing number of bioactive natural products was developed.228  A novel method showed that carbohydrates can be applied as starting materials to prepare amino acids. The synthesis of (2S,5R) and (2S,5S)-5-hydroxy-lysine was reported by using d-galactose as a chiral precursor with stereo retention.229  The mechanism of action of several hydroxy-Trp containing tritrpticin derivatives that has a strong microbial activity against Gram-positive and Gram-negative bacteria as well as fungi was studied. The addition of a hydroxyl group to the indole ring of Trp was able to modify the mechanism of action of the peptides. This study also shows that 5OH-Trp constitutes a new probe to modulate the antimicrobial activity and mechanism of action of other Trp-rich antimicrobial peptides.230 

Gamma-hydroxy norvaline (Scheme 12) was produced in a one-pot organocatalytic Mannich reaction and an enzymatic ketone reduction by using 2-PrOH both as solvent and as reducing agent. Each of the four stereoisomers was achieved in high yield and excellent stereoselectivity.231 

The β-turn inducer cyclic hydroxy-amino acids also are important building blocks for the development of new therapeutic drugs. During the last decades, the stereoselective synthesis of hydroxy-amino acids has been growing since some of their representatives show enzyme inhibitory activities. An enantioselective total synthesis of the amino acid (2S,4R,5R)-4,5-di-hydroxy-pipecolic acid starting from d-glucoheptono-1, 4-lactone is described that involved 12 steps with an overall yield of 19%. The structures of the compounds synthesized were elucidated on the basis of comprehensive spectroscopic (NMR and MS) and computational analysis.232 

Four novel potential renin inhibitors have been synthesized. All these inhibitors contain unnatural moieties that are derivatives of N-methylleucyl-β-hydroxy-γ-amino acids (4-[N-(N-methylleucyl)-amino]-3-hydroxy-7-(3-nitroguanidino)-heptanoic acid, 4-[N-(N-methylleucyl)-amino]-3-hydroxy-5-phenyl-pentanoic acid, or 4-[N-(N-methylleucyl)-amino]-8-benzyloxycarbonylamino-3-hydroxyoctanoic acid).19 

Unsaturated α-amino acids have turned out to be especially important building blocks for incorporation into proteins due to the diverse reactivities of the multiple bonds and their ability to introduce biologically active functionalities. These compounds are also used in peptide chemistry to confer β-turn secondary structure to induce new properties as enzyme inhibitors and peptidomimetics as therapeutic agents. A review of Boibessot T provides an overview of the literature from 1977 to 2015 concerning synthesis and applications of α-quaternary α-ethynyl α-amino acids (Scheme 13).143 

Sterically constrained α-amino acids are important in drug industry. Asymmetric synthesis of (1R,2S)-1-amino-2-vinylcyclopropanecarboxylic acid was achieved via two-step SN2 alkylation of a novel chiral nucleophilic Gly equivalent with excellent yields and diastereoselectivity.105 

Enantiopure α-amino phenylacetic acids (phenylglycines) are important structural motifs found in many pharmaceuticals and biologically active compounds, e.g. antibiotics. Phenylglycine (Phg) derivatives can also be used as catalysts and chiral building blocks in organic syntheses. Kinetic resolution of enantioselective C–H olefination of α-amino phenylacetic acid was developed by a palladium(II)-catalyzed enantioselective C–H activation and C–C bond formation.102  Stereoselective synthesis of unsaturated α-amino acids by asymmetric alkylation was summarized. An enantioselective approach induced by the Corey–Lygo catalyst under chiral phase transfer conditions or the hydroxypinanone chiral auxiliary, both implicating Schiff bases as substrate (Scheme 14). The use of a prochiral Schiff base gave higher enantiomeric excess and yield in the final desired amino acid.233 

A transition metal-free, organocatalytic asymmetric synthesis for β-alkynyl-β-amino acids have been developed via a mild chiral Brønsted base-catalysed asymmetric Mannich-type reaction of in situ generated N-Boc C-alkynyl imines with α-substituted β-keto esters and less-acidic malonate (thio)esters as nucleophiles with high efficiency. The catalytic activation strategy may have broad use in catalysis and synthesis. This methodology could also be applied for the catalytic asymmetric synthesis of biologically important β-alkenyl-β-amino acids that are difficult to prepare by asymmetric catalysis and β-alkyl-β-amino acids.234  Palladium-catalyzed N-quinolyl carboxamide-directed olefination of the inactivated C(sp(3))–H bonds of phthaloyl Ala with vinyl iodides (Scheme 15) led to a wide range of β-vinyl α-amino acids.235 

One-pot synthesis of highly functionalized α-vinylated γ-oxo-β-amino esters from three-components is revealed. The cycloaddition of the 1-alkyne and sulphonyl azidetriazole with the use of Cu(i)-catalyst resulted in 1-sulfonyl-1,2,3-triazole. An α-imino Rh(ii)-carbene generated from an open-chain α-imino diazo derivative of the triazole, reacts with γ-hydroxy α,β-unsaturated esters to form allylic (Z)-amino vinyl ethers. The later one led, after rearrangement, to α-vinyl γ-oxo-β-amino esters in high yields and diastereoselectivity.236 

Photoredox α-vinylation of α-amino acids produced several natural products and a number of pharmacophores.237  Radical decarboxylative allylation of N-protected α-amino acids and esters has been accomplished via a combination of palladium and photoredox catalysis.238  Derivatives of all-carbon quaternary allylic amino acids have been produced by a Brønsted acid accelerated Pd-catalyzed asymmetric allylic alkylation of azlactones with simple allylic alcohols.136  Protected δ,ε-unsaturated α,β-diamino acids as templates have been used for the preparation of 12 new α,β-diamino acids.239  Derivatives of exomethylenic cyclohexane β-amino acid were selectively synthesized from unsaturated bicyclic β-lactams by transformation of the ring olefin bond through three different regio- and stereocontrolled hydroxylation techniques, followed by hydroxy group oxidation and oxo-methylene interconversion with a phosphorane.240 

Dehydroalanines equipped with oxazolidin-2-one chiral auxiliaries have been prepared and applied for stereoselective synthesis of substituted tryptophans [(S)-2-methyltryptophan and (S)-5-fluoro-Trp].241 

Non-covalent interactions involving aromatic amino acids are ubiquitous in nature and facilitate most of the chemical and biological processes. Aromatic amino acids play a diverse role in stabilization of α-helix and β-sheet of soluble and membrane proteins.242  The most often used strategy to increase potency, selectivity, and metabolic stability is constraining the conformation of flexible peptides. Not only constraining the backbone dihedral angles, but the correct orientation of the amino acid side chains is equally important. The applications of cyclized analogues of the aromatic amino acids (Phe, Tyr, Trp, and His) within peptide medicinal chemistry are showed with examples of enzyme inhibitors and ligands for G protein-coupled receptors.243 

Significant advances in C–H bond functionalisations have been achieved with the discovery of new mechanisms in the past decade. Coordinating activation strategy using Nickel-catalysed radical oxidative cross-coupling between C(sp(3))–H bonds and (hetero)arylmethyl free radicals have been developed. Considering that the free radical acts as a real coupling partner, the reported transformation has further enriched the type of the oxidative cross-coupling reactions. This method can use many α-amino acids, (hetero)arylmethanes, arylmethylenes, and arylmethines leading to a large library of both α-tertiary and α-quaternary β-aromatic α-amino acids. In addition, this reaction can proceed well on a larger scale, and the coordinating group can be handled readily and removed easily.244 

Intermolecular catalytic (Pd(ii)) arylation of inactivated β-C(sp3)–H bonds in α-hydroxy aliphatic acid derivatives with aryl iodides was achieved. The feasibility of amino acid auxiliary as a directing group also was demonstrated.245  Direct palladium-catalyzed C(sp(3))–H arylation of amino acid derivatives with aryl iodides bearing different electronic properties resulted in the desired amino acid molecule by the cleavage of the tethered click-triazoles after the catalytic reaction. This provides a practical protocol for the production of both natural and synthetic amino acids.246  Complex β-aryl α-amino acids have been synthesized by palladium-catalyzed β-C(sp(3))–H arylation of phthaloyl Ala derivatives with aryl iodides. A variety of aryl iodides (e.g. bearing alkoxyl, carbonyl, nitro, and halogen groups in ortho position) can react with the 2-(2-pyridyl) ethylamine-coupled phthaloyl Ala (Scheme 16). This method can be used for preparing complex β-aryl α-amino acids.247  A highly efficient monoarylation reaction by Pd-catalyzed β-methyl C(sp(3))–H of an Ala derivative with aryl iodides using an 8-aminoquinoline led to various β-aryl-α-amino acids with high efficiency and retention of chirality.248  The mechanism of amino acid ligand-assisted Pd(ii)-catalyzed C–H activation and the important roles of the amino acid ligand and the CsF base was analyzed in a review.126 

Fmoc-protected aryl/heteroaryl-substituted phenylalanines (Bip derivatives) using the nonaqueous palladium-catalyzed Suzuki–Miyaura cross-coupling reaction of Fmoc-protected bromo- or iodophenylalanines have been reported (Scheme 17). This method allows for the direct formation of a variety of unnatural biaryl-containing amino acids.249  Substituted d-phenylalanines have been synthesized starting from inexpensive cinnamic acids with a novel one-pot approach by coupling phenylalanine ammonia lyase amination with a chemoenzymatic deracemization (based on stereoselective oxidation and nonselective reduction). Multienzymatic cascade increased the yield and optical purity of the D-configured product.250 

Novel nonsymmetrically substituted N-protected β,β-diaryl-α-amino acids and esters have been produced through the asymmetric hydrogenation of tetrasubstituted olefins. A variety of N-acetyl, N-methoxycarbonyl, and N-Boc β,β-diaryldehydroamino acids, containing a diverse and previously unreported series of heterocyclic and aryl substituted groups (24 examples) were obtained with high yields and excellent enantioselectivities.127 

Eighteen substituted α-arylalanines were converted to their (S)-β-aryl and β-heteroaryl-β-amino acids in vivo that showed no toxicity to cells. The synthesized substituted β-aryl-β-amino acids can be used in redox and Stille-coupling reactions to make synthetic building blocks, or as bioisosteres in drug design.251  Aryloxyproline diastereomers can stereospecifically used in molecular design, medicinal chemistry, and catalysis. The synthesis of (2S,4R)- and (2S,4S)-iodophenyl ethers of hydroxy-Pro has been described that can be differentially applied in distinct structural contexts.186  It was reported that a pyridine-type ligand that overcomes the limitation of N-methoxyamide auxiliary leads to a significant improvement of β-arylation of α-amino acids. The arylation used in this method can be applied for syntheses of unnatural amino acids, bioactive molecules, and chiral bis(oxazoline) ligands.252 

Conjugated unnatural α-amino acids bearing a 5-arylpyrazole side-chain has been prepared by Horner–Wadsworth–Emmons reaction of an aspartic acid derived β-keto phosphonate ester with aromatic aldehydes resulting in β-aryl α,β-unsaturated ketones. A further reaction with phenyl hydrazine followed by oxidation gave the regioselective synthesis of pyrazole derived-amino acids.253 

A new method has been developed for the asymmetric synthesis of chiral heterocyclic amino acids via alkylation of the Ni(ii) complex of Gly and alkyl halides (Scheme 18). The intermediate decomposes to form a series of chiral amino acids in high yields and with excellent diastereoselectivity.254 

Enantiopure 1,2,3-triazolyl-β-amino acids were prepared from the corresponding alkynyl-β2-amino acids. The enantiopure products were obtained via diastereomeric salt formation and subsequent separation.255  The synthesis of novel unnatural N(α)-Fmoc pyrimidin-4-one amino acids is based on an aromatic nucleophilic substitution reaction between 4-[4-(benzyloxy)benzyloxy]-2-(benzylsulphonyl)pyrimidine and the nucleophilic side chain of several N(α)-Boc amino esters followed by a series of standard protecting group transformations.256  Indole-substituted (S)-tryptophans have been produced from corresponding indoles with the use of (S)-methylbenzylamine and derivatives. The products utilize a chiral auxiliary-facilitated Strecker amino acid synthesis strategy. Eight optically pure (S)-Trp analogues were synthesized, which were subsequently used for the convergent synthesis of a potent antibacterial agent, argyrin A and its analogues.257  The amino acid His plays a significant role in the structure and function of proteins. Its functions include enzyme catalysis, metal binding activity, and involvement in cation–π, π–π, salt-bridge, and other types of noncovalent interactions. N–H⋯N hydrogen bonds involves imidazole nitrogen atom of His. Along with the predominant occurrence in loop segments, a new structural role for His in protein structures was proposed.258 

Two tetrahydrofuran amino acid derivatives were synthesized to replace Tyr in neurotesin. The new compounds showed substantial neurotensin S2 receptor binding affinity and up to 1000-fold selectivity over neurotensin S1 receptor.259 

All biological systems code strictly 20 canonical amino acids with few exceptions. Given the limited functionalities of 20 amino acids, biochemists have longed for an approached for the production of proteins with unique biochemical and biophysical handles. Pyrrolysine (Pyl), the 22nd proteinogenic amino acid, was restricted until recently to few organisms. Its translational use necessitates the presence of enzymes for synthesizing it from lysine. A review discusses the pyrrolysine (Pyl) incorporating system from its discovery to its applications. In about a decade after the Pyl incorporation mechanism was discovered, the system has been transferred to bacteria, yeast, and mammalian cells for the genetic incorporation of more than 100 non-canonical amino acids or α-hydroxy acids into proteins. With the magnificent tools developed, the field might diverge to focusing on a large variety of applications from basic study to biotechnological development.260  Functionalized Pyl analogues for site specific protein labelling and biochemical studies have been developed. Incorporating a wide array of non-canonical amino acids by the Pyl translational mechanism made possible the synthesis of recombinant proteins. The first use of this technology for the production of branched cyclic proteins have been reported.261 

It was demonstrated how Pyl synthase can be used to oxidize various isopeptides to novel amino acids by combining chemical synthesis with enzyme kinetics and X-ray crystallography. A detailed description of the Pyl synthase reaction is suitable for the biosynthesis of pyrroline and tetrahydropyridine rings as constituents of Pyl analogues.262  The gas-phase conformational potential energy surfaces of Pyl and related derivatives (neutral, deprotonated, and protonated) were extensively searched. The conformational electronic energies and thermochemical properties of proton affinity/dissociation energy and gas-phase acidity/basicity were also determined.263 

The development of an efficient synthesis of N-hydroxy-α-amino acids and derivatives represents a challenging goal in organic synthesis since these compounds are key intermediates in metabolic pathways and can be found in human and animal tumours.

In peptoids the side chain is connected to the nitrogen of the peptide backbone, instead of the α-carbon as in peptides. Like d-peptides and β peptides, peptoids are completely resistant to proteolysis, and are therefore advantageous for therapeutic applications where proteolysis is a major issue. N-alkylated N-arylsulfonylglycines were prepared. The procedures gave access to peptoid monomers bearing a wide variety of functional groups. The synthesized N-substituted N-arylsulfonylglycines were used as monomers in solid-phase synthesis to receive peptoid oligomers and peptide–peptoid hybrids.264  Accelerated submonomer solid-phase synthesis of peptoids was achieved by incorporating multiple substituted N-aryl Gly monomers. This method enables the rapid room temperature synthesis of a wide variety of N-aryl Gly-rich peptoid oligomers in good yields.265 

Catalytic enentioselective synthesis of N,C(α),C(α)-trisubstituted derivatives of α-amino acids was achieved by using 1H-imidazol-4(5H)-ones as novel nucleophilic α-amino acid equivalents. These compounds provide direct access to N-substituted (alkyl, allyl, aryl) α-amino acid derivatives.116  3-[(2-Hydroxyphenyl)amino]butanoic and 3-[(2-hydroxy-5-methyl(chloro)phenyl)amino]butanoic acids were converted to a series of derivatives containing 2-hydroxyphenyl, hydrazide, pyrrole, and chloroquinoxaline moieties. The new compounds were tested for their antimicrobial activities.266  N-Substituted acyclic β-amino acids were synthesized and investigated for their effect as GABA uptake inhibitors.267 N-aralkylpyroglutamic acids of substituted bispidine were prepared and evaluated for their ability to inhibit collagen induced platelet aggregation.268 

A series of dithiocarbamate ligands derived from N-substituted amino acids reacted with different diorganotin dichlorides to give 18 diorganotin complexes. The variations in the molecular conformation, shape, and cavity size of the macrocycles depending on the aliphatic spacer was investigated.269 

The interactions of the side chains of the highly polarizable sulphur containing amino acids Met and Cys have received little attention, in contrast to aromatic–aromatic, aromatic–aliphatic, or/and aliphatic–aliphatic interactions. It was found that sulphur containing amino acids form stronger interactions than aromatic or aliphatic amino acids. Thus, these amino acids may provide additional driving forces for maintaining the structure of membrane proteins and may provide functional specificity.270  The side chain of Met exhibits different degrees of susceptibility to oxidation depending on solvent accessibility. Different sets of oxidation-sensitive and oxidation-resistant methionines contained in human proteins are investigated. The examination of the sequence surrounding of the non-oxidized Met revealed a preference for neighbouring Tyr and Trp residues, but not for Phe. The results showed that the S-aromatic motif, which decreases the reactivity of the involved sulphur towards oxidants is important.271  Pd(ii)-Met methyl ester complex was synthesized and characterized by physicochemical measurements, and interaction with biorelevant ligands was investigated.272 

Amino acids containing sulphur are important in Maillard reaction. The reaction conditions for the blue Maillard reaction products were only studied by few research groups. A study investigates the characteristic colour formation and antioxidant activities in four different Maillard reaction model systems and the optimum reaction conditions for the blue colour formation were investigated in a xylose-Gly model system.273  Glucosamine and Cys were also studied in aqueous model systems to investigate the effects of reaction temperature, initial pH, reaction time, and concentration ratio of glucosamine and Cys. The optimum condition for colour change was also measured.274 

Plasma homocysteine (Hcy) is an important risk factor for various diseases. A novel redox-sensitive fluorescent probe was developed for the selective detection of Hcy.275  The [4-thio]-S-ribosylhomocysteine analogues were synthesized by coupling of 4-thioribose with a thiolate generated from the protected Hcy.276  S-ribosylhomocysteinase enzyme (EC catalyses the cleavage of the thioether linkage of S-ribosyl-Hcy producing Hcy and 4,5-dihydroxy-2,3-pentanedione which is the precursor of the signalling molecules known as autoinducer 2 responsible for the bacterial cell to cell communication. Isomeric analogues of S-ribosyl-Hcy with Hcy unit attached to C2 of ribose ring via C2-sulfur bond have been synthesized.277 

Proteins are often attacked by hydroxyl radicals, that are the most reactive oxygen species, and this oxidation causes diseases. Several oxidized products have been experimentally characterized, but the reaction pathways remain unclear. Sulphur-containing amino acids would be oxidized more easily than OH-containing amino acids. This was proven by the stability of the sulphur radical intermediates.278  Multiple inputs control the synthesis of sulphur-containing amino acids Met and Cys in Saccharomyces cerevisiae.279 

A novel chemoselective and stereochemically defined synthetic method to phosphorylate the side-chain of Cys is reported. This work provides a novel synthetic strategy to incorporate native phosphorylated Cys (pCys) residues into unprotected peptides. The results show that the presented chemical and analytical tools are highly valuable in accessing endogenous pCys peptides and thereby open the possibility to identify new Cys phosphorylation sites from biological samples.280 

Met adenosyltransferase catalyses the metabolism of Met in the liver by converting it to S-adenosylmethionine. The importance of homeostasis in the metabolism of sulphur-containing amino acids with a particular focus on the transsulphuration pathway which could be a promising therapeutic target in liver injury is reviewed.281 

The antioxidative activities of free l-amino acids against Fenton system-mediated hydroxyl radical (HO(˙)) production in aqueous solution was compared, and the relation between amino acids and a set of physicochemical properties was examined. Sulphur-containing free l-amino acids generated different secondary reactive products, which were discriminated by electron paramagnetic resonance spin-trapping spectroscopy. The findings are important for the understanding of oxidation processes in natural and waste waters.282  Sulphur-containing amino acids have dual role in the mechanochemical synthesis of IV–VI semiconductor nanocrystals from lead acetate and l-cystine.283 

Enantiopure fluorinated analogues of Met and Cys (l-trifluoro-Met and l-S-(trifluoromethyl)-Cys) have been synthesized by using a cheap and user-friendly radical trifluoromethylation approach. The protected amino acid derivatives were used in peptide syntheses both liquid- and solid-phase.284  Cinchona alkaloids functionalized with a hydrogen bond donating group at the C6′ position enantioselectively catalyse sulfa-Michael additions to α,β-unsaturated N-acylated oxazolidin-2-ones and related α,β-unsaturated α-amino acid derivatives. The products were subsequently converted in high yields to enantiopure β-functionalized cysteines suitable for native chemical ligation.285  A dithiol amino acid (Dtaa) that can form two disulphide bridges at a single amino acid site have been developed. Application of Dtaas to a serine protease inhibitor and a nicotinic acetylcholine receptor inhibitor, that contain disulphide constraints, enhanced their inhibitory activities.286 

Selenium initially was described as a toxin and was shown to be essential for health and development. By the mid-1990s selenium emerged as one of the most promising cancer chemopreventive agents, but subsequent human clinical trials yielded contradictory results. Selenocysteine (Sec) is a naturally occurring proteogenic amino acid that is encoded in the genomic sequence of relatively abundant proteins in many of the model species commonly used for biomedical research.287  Sec is incorporated into proteins by unique synthetic mechanisms. Regulation of Sec incorporation into the selenium transport protein was investigated. It was revealed that Sec incorporation requires both cis- and trans-acting factors, which are known to be sufficient for Sec incorporation in vitro. It was suggested a role for yet unidentified mammalian-specific processes or factors.288  It was also shown that the subcellular location of most human selenoproteins containing Sec has impact on their function.289  Mammalian selenoproteins contain single Sec residues, with the exception of selenoprotein P that has 7–15 Sec residues depending on species. The regulation of Sec content of human selenoprotein P was investigated.290 

Chiral α-selenoamino acid derivatives were prepared from N-acetoxyphthalimide derivatives of Asp and Glu as visible light photoredox chiral sources and diorganyl diselenides as radical acceptors. This simple method with mild reaction conditions and high efficiency provides an important strategy for the synthesis of chiral molecules.291  2,2′-dithiobis-5-nitropyridine dissolved in trifluoroacetic acid (with or without thioanisole) was used previously for the removal of 4-methoxybenzyl and acetamidomethyl protecting groups from Cys and selenocysteine (Sec). Disadvantage of this method is that excess thiol must be used to drive the reaction to completion and then removed before using the Cys-containing or Sec-containing peptide in further applications. Advancement of this method shows that ascorbate at pH 4.5 and 25 °C reduces the selenosulphide to selenol. This method might find a number of other applications.292 

Malfunctions of selenoproteins can lead to various human disorders including cancer.293  The naturally available Sec exhibits novel anticancer activities against human cancer cell lines. The roles of Sec in cancer, health, and development was summarized.294  Investigations clearly demonstrate that selenoproteins that contain Sec can act as oncosuppressors, but can also favour the formation of malignant tumours.295  In another study, the understanding on the molecular mechanisms of Sec in human glioma treatment was revealed.296  Castration-resistant progression of prostate cancer after androgen deprivation therapy remains a critical challenge in the clinical management of prostate cancer. A study showed the preclinical efficacy of methyl-Sec in delaying castration-resistant progression of prostate cancer.297 

An oxidized Sec (Se–S bond) which has electrophilic character reacts with a nucleophilic arylboronic acid to provide the arylated Sec. The arylated derivatives are more stable under oxidative conditions than the corresponding alkylated Sec. This reaction is unique to Sec, and a wide range of boronic acids can react with different biorelevant functional groups.298 

The synthesis of Sec derivatives which bear TFA-labile sidechain protecting groups was reported from a bis Fmoc-protected Sec precursor. The new compounds (Fmoc-Sec(Xan)-OH and Fmoc-Sec(Trt)-OH) are useful and practical alternatives to the traditional Fmoc-Sec-OH derivatives currently available to the peptide chemist.299  Sec is synthesized from a serine precursor in a series of reactions that require Sec tRNA. It was shown that allosteric regulation might play an important role in regulation of Sec and selenoprotein synthesis.300  Twenty one derivatives of Cys and Sec are synthesized from easily prepared protected dichalcogenide precursors in high yield. The crude form of these derivatives were precipitated from petroleum ether in sufficient purity for direct use for peptide synthesis.301  Site-specific dual antibody conjugation via Cys and Sec was described for the production of homogeneous antibody-drug conjugates with improved therapeutic effects.302 

The important biological redox mediators for two-electron transfers (thioredoxin reductases) contain either 2 cysteines or a Cys and a Sec at the active site. In case of an accidental one-electron transfer to a S–S or a S–Se bond during catalysis, a thiyl or a selanyl radical would be formed, respectively. The thiyl radical can abstract a hydrogen from the protein backbone, which subsequently leads to the inactivation of the protein. But a selanyl radical will not abstract a hydrogen. Therefore, formation of Sec radicals in the active site will less likely result in the destruction of a protein.303 

A variation of native chemical ligation involves the reaction of peptides bearing an N-terminal Sec residue with peptide thioesters, which proceeds through the same mechanism as the parent reaction. It was discovered that Sec within peptides can be chemoselectively deselenized without the concomitant desulfurization of Cys residues. A review summarizes the use of Sec in ligation chemistry and investigations of chemoselective ligation-deselenization chemistry at other selenol-derived amino acids.304  The use of native chemical ligation at selenocysteine residues with peptide thioesters and a one-pot oxidative deselenization chemistry that uses native chemical ligation at Sec residues with peptide thioesters and additive-free selenocystine ligation is described. This simple and rapid method leads to native peptides with Ser in place of Sec at the ligation junction.305 

Trifluoroselenomethionine (TFSeM), a new unnatural amino acid, was synthesized in seven steps from N-(tert-butoxycarbonyl)-l-aspartic acid tert-butyl ester. TFSeM shows enhanced methioninase-induced cytotoxicity against HCT-116 cells derived from human colon cancer. It was shown that transformation of TFSeM into seleno-Met is enzymatically catalysed by E. coli extracts, but TFSeM is not a substrate of E. coli Met adenosyltransferase.306 

Phosphorylation of Ser, Thr, and Tyr is a well-known posttranslational modification of proteins, but phosphorylation of other amino acid side chains is underappreciated and minimally characterized by comparison. In case of the O-phosphorylated amino acids, synthetic constructs were critical to assessing their stability and developing tools for their study. A review summarizes the synthetic chemical approaches of labile amino acid phosphorylation.307 

In recent years, substantial progress has been made in the field of asymmetric phosphine catalysis; many new reactions have been discovered; and numerous enantioselective processes have been reported. New families of powerful amino acid-derived bifunctional phosphines were developed for new modes of phosphine activation, unknown reactions, and more enantioselective transformation.308  Important α-amino phosphonic acids and its derivatives have been synthesized by using copper-catalysed electrophilic α-amination of phosphonates and phosphine oxides with O-acyl hydroxylamines. This amination provides the first example of C–N bond formation which directly introduces acyclic and cyclic amines to the α-position of phosphonates in one step.309  Diphenyl (α-aminoalkyl)phosphonates act as inhibitors against serine proteases by forming a covalent bond with the hydroxy group of Ser in the active centre. The stereochemical effect of the diphenyl phosphonate moiety on the selective chemical modification was evaluated. The results demonstrate that the peptidyl derivatives bearing an optically active diphenyl phosphonate moiety can be used as affinity labelling probes in protein bioconjugation.310  A simple, efficient, and versatile organocatalytic asymmetric 1,2-addition reactions of α-isothiocyanato phosphonate were developed. Through these processes, derivatives of β-hydroxy or β-amino substituted α-amino phosphonic acid and α,β-diamino phosphonic acid derivatives were produced. This new method provides a novel route for the enantioselective functionalization of α-phosphonic acid derivatives.311  A series of phosphonodipeptides containing C-terminal α-aminoalkylphosphonic acids has been synthesized from 2-(N-benzyloxycarbonylamino)alkanamides, aldehydes, and phosphorus trichloride via Mannich-type reaction and subsequent sequential hydrolysis in good yields. The reaction mechanism was proposed and verified.312  Activated carbon-based amino phosphonic acid chelating resin was produced, and its adsorption properties for Ce(iii) removal was investigated. The chelating resin based on activated carbon adsorbent was prepared from activated carbon followed by oxidation, silane coupling, ammoniation and phosphorylation, and characterized.313  Fifty phosphorus-containing amino acids (phosphonic and phosphinic acids) were screened for inhibition of human endoplasmic reticulum aminopeptidases. The SAR studies revealed several potent compounds, particularly among the phosphinic dipeptide analogues, that were strong inhibitors.314 

Highly efficient and chemoselective phosphorylation of amino acid derivatives was achieved with the use of phosphoryl chlorides catalysed by 2-aryl-4-(dimethylamino)pyridine-N-oxides.315  The strength of hydrogen-bond in phosphorylated and sulphated amino acids were investigated. The findings showed that for phospho-Tyr (pTyr), bidentate interactions with Arg are particularly dominant, as has been previously demonstrated for pSer. The interactions of sulfo-Tyr with Arg are significantly weaker, even as compared to the same interactions made by Glu.316  4-Phosphothiophen-2-yl Ala, a novel five-membered ring analogue of pTyr was developed. The new analogue showed high selectivity for pTyr and no cross-reactivity with other phosphorylated amino acids.317  4-Phosphopyrazol-2-yl Ala, a non-hydrolysable analogue of phospho-His, was also synthesized.318  The interactions between O-phosphorylated and standard amino acid side-chain models in water were investigated by theoretical studies.319  Recently, peptides and proteins containing N-phosphorylated amino acids such as phospho-Arg (pArg), phospho-His (pHis), and phospho-Lys (pLys) have gained interest because of their different chemical properties and stability profiles. A direct synthetic approach to incorporate pLys residues in a site-specific manner into peptides in solution was reported by taking advantage of the chemoselectivity of the Staudinger-phosphite reaction.320 

Aminoacyl adenylates constitute essential intermediates of protein biosynthesis. Their polymerization in aqueous solution is a potential route to abiotic peptides despite a highly efficient CO2-promoted pathway of hydrolysis. The efficiency and relevance of this frequently overlooked pathway from model amino acid phosphate mixed anhydrides including aminoacyl adenylates were reported. The evolutionary importance of the intramolecular pathways of hydrolysis of phosphate ester mixed anhydrides with amino acids and peptides was also discussed.321 

Radiolabelled peptides with high specificity and affinity towards receptors that are overexpressed by tumour cells are used in nuclear medicine for diagnosis and therapy of cancer. Since Met is prone to oxidation during radiolabelling procedures and the formation of oxidative side products can affect the purity of the final radiopharmaceutical and affinity towards the corresponding receptor, it is important to substitute Met with oxidation resistant amino acid analogues. Replacement of Met with Nle preserves the length of the side chain of amino acid that is important for hydrophobic interactions, but not its hydrogen-bonding properties. Methoxinine (Mox), a non-canonical amino acid resembles more closely the electronic properties of Met than Nle and represents a suitable, oxidation-stable amino acid substitute of Met in radiolabelled peptide conjugates.322 

A mild and selective photocatalytic synthesis of oncological positron emission tomography (PET) imaging agents was developed by 18F-fluorination of inactivated C–H bonds in branched aliphatic amino acids. The biodistribution and uptake of three 18F-labelled Leu analogues in several cancer cell lines is reported.323  For tumour imaging with PET, 18F-labelled Trp derivatives were synthesized by electrophilic 18F-fluorination or by introducing a [18F]fluoroalkyl group in 3-steps.324  Two simultaneous click reactions in one-pot with a simple solid-phase extraction purification method were developed for the synthesis of new [18F]fluorinated 1,2,3-triazolyl amino acid derivatives. The final product was completely pure in terms of radiochemical (>95%) and chemical purity with high radiochemical yields.325  A novel synthetic method was developed to achieve 6-18F-fluoro-3,4-dihydroxy-L-Phe (18F-DOPA). The method involved the nucleophilic substitution of a diaryliodonium salt precursor with non-carrier-added 18F-fluoride.326  New developments in radiochemistry are summarized in a review. The new methods provide solutions to long standing problems involved in the synthesis of the important but elusive radiotracer [18F]6-fluoro-3,4-dihydroxy-L-Phe ([18F]F-DOPA). Advances in nucleophilic synthesis have been achieved by optimising multistep strategies and using both hypervalent iodine chemistry and transition metal-mediated fluorinations.327  A novel 18F-labelled α,α-disubstituted amino acid-based tracer has been synthesized for brain tumour imaging with a long alkyl side chain to increase brain availability vial-amino acid transport system. Biodistribution and in vitro uptake assays showed that both (R)- and (S) 2-amino-5-[18F]fluoro-2-methylpentanoic acid have good tumour imaging properties with the (S)-enantiomer providing higher tumour uptake and tumour to brain ratios.328  Four structurally related non-natural 18F-labelled amino acids in (R)- and (S)-configuration have been prepared and evaluated for their potential in brain and systemic tumour imaging with PET. All four tracers showed moderate to high levels of uptake by the cancer cell lines tested. The accumulation of these tracers was higher in tumour than most normal tissues.329  New fluoroethoxy Trp analogues were synthesized and evaluated in vivo. Radiosynthesis was accomplished by no-carrier-added nucleophilic 18F-fluorination following either an indirect or a direct approach using a protected mesyl precursor.330  Two 18F-labelled analogues of 5-hydroxy-L-[β-11C]Trp have been synthesized as tracers for tumour imaging.331  Amino acid transport is an attractive target for imaging in oncology. Despite a high demand of cancer cells for cationic amino acids, their potential as PET probes remains unexplored. The synthesis and preliminary biological evaluation of a new cationic tumour imaging agent O-2((2-[18F]fluoroethyl)methylamino)ethyl-Tyr were reported.332 

It was previously reported that high tumour uptake is shown by 11C-labelled 2-amino-2-methyl-butanoic acid (Iva). 11C-Iva is a promising PET probe for non-invasive tumour imaging.333  D-isomers of some radiolabelled amino acids are potential PET tracers for tumour imaging, e.g. the D-isomer of S-11C-methyl-D-Cys that was synthesized and evaluated.334  The radiolabelled serotonin precursor [11C]5-hydroxy-Trp showed significant accumulation in the pancreas of healthy volunteers by dynamic PET. In the contrary, a substantial and highly significant reduction in the pancreatic uptake was seen in type 1 diabetic patients. Retention of [11C]5-hydroxy-Trp seems to be a useful non-invasive surrogate marker for the human endocrine pancreas.335  The 11C-methylation of Schiff-base-activated α-amino acid derivatives has been optimized for the radiosynthesis of various α-11C-methyl amino acids.336 

A convenient alternative to Fluorine-18 and Carbon-11 can be Nitrogen-13. A one-pot, enzymatic and non-carrier-added synthesis of the 13N-labelled amino acids (L-[13N]Ala, [13N]Gly, and L-[13N]Ser) was achieved by using L-Ala dehydrogenase from Bacillus subtilis. After optimization of the experimental conditions, the radiochemical yields were sufficient.337 

The 13C-enrichment measurements were used as a direct calibration to calculate the original 13C/12C ratios of individual amino acids. The number of non-analyte added carbon atoms and assess the non-stoichiometrical recovery due to their incomplete oxidation was determined for the main proteinogenic amino acids.338  L-[4-13C]Gln was prepared by Wittig reaction. This method resulted in fewer steps and higher yield than previously reported methods.339  Leu, Val, and Ile were exclusively 13C labelled on methyl groups. This isotopic labelling strategy represents an easily obtainable unambiguous long-range distance restraints in protein solid-state NMR studies.340 

Aromatic amino acids such as Phe, Trp, 3′,4′-dihydroxy-L-Phe (L-DOPA), and their derivatives play an essential role in human metabolic processes. Incorrect or slow biotransformation of these compounds leads to some metabolic dysfunctions and neurodegenerative diseases. The mechanisms of biotransformation of the above amino acids was investigated by using kinetic and solvent isotope effect methods.341 

Halogen derivatives of L-Trp (4′-F-, 7′-F-, 5′-Cl- and 7′-Br-L-Trp) were specifically labelled with deuterium in α-position of the side chain. The labelled compounds were obtained by enzymatic coupling of the corresponding halogenated derivatives of indole with S-methyl-L-Cys in H2O, catalysed by enzyme tryptophanase. 100% deuterium labelling was observed in the α-position.342  Halogenated derivatives of L-Trp, L-Tyr, and L-Phe were labelled with tritium and doubly with deuterium and tritium. Enzymatic methods were used for the synthesis with tritiated water and heavy water (D2O).343 

Four new 68Ga-labelled 1,4,7,10-cyclododeca-1,4,7,10-tetraacetic acid/1,4,7-triazacyclononane-1,4,7-triacetic acid derived glycine/hippurate conjugates have been produced for PET renography. The 68Ga labelling was achieved by reacting an excess of the non-metallated conjugate with 68Ga14C.344 

Amino acids labelled with 76Br are attractive positron emitters with relatively long half-life, and they could potentially be used as tumour imaging tracers. Two 76Br-labelled L-Phe derivatives were produced and investigated. The findings suggest that 2-76Br-bromo-α-methyl-L-Phe could constitute a potential new PET tracer for tumour imaging.345  The novel compound, (S)-amino-2-methyl-4-76Br]bromo-3-(E)-butenoic acid was synthesized and characterized. The above compound is a promising PET tracer for brain tumour imaging and lead compound for a mixed transport substrate.346 

Fluorescence spectroscopy has become a powerful tool for probing a range of complex biological processes including enzyme mechanisms and protein–protein interactions. A review describes advances in design, properties, and applications of fluorescent amino acids.347  Another review highlights the recent synthetic methods developed for the incorporation of highly conjugated chromophores (e.g. coumarin, flavone, and polyaromatic derived chromophores) into the side-chain of α-amino acids and the application of these compounds as probes for imaging in medicine and biology.348  A fluorescent l-amino acid bearing the 4′-methoxy-3-hydroxyflavone fluorophore that shows dual emission has been synthesized. The fluorescent amino acid undergoes excited state intramolecular proton transfer for site-specific probing and imaging of peptide interactions.349 

Small fluorescence probes are useful for tracking changes in the interior space of proteins. Fluorescence donors of six unnatural amino acids (structurally related to Trp) are described and show how they can be efficiently incorporated into a protein as fluorescence probes.350  The design and synthesis of microenvironment sensitive fluorescent triazolyl unnatural amino acids were reported that are decorated with donor and/or acceptor aromatic chromophores via click chemistry. The synthesized fluorescent amino acids show interesting solvatochromic characteristic and/or intramolecular charge transfer feature. One of the amino acid (triazolyl-perylene amino acid) has been exploited for studying interaction with BSA and found that it is able to sense BSA with an enhancement of fluorescence intensity.351 

New demands for imaging technologies and in vivo diagnostics are increasing. There is a desire for new reporter molecules that can provide strong signals, are non-toxic, and can be tailored to diagnose or monitor the progression of several diseases. Aequorin is a non-toxic photoprotein that can be used as a sensitive marker for bioluminescence in vivo imaging. The sensitivity of aequorin is due to the fact that bioluminescence is a rare phenomenon in nature and, therefore, it does not suffer from autofluorescence, which contributes to background emission.352 

The first purely chemical method has been reported for the dynamic kinetic resolution of unprotected racemic α-amino acids. This method can challenge the economic efficiency of the enzymatic reactions.353  Chemical dynamic thermodynamic resolution and S/R interconversion of unprotected tailor-made α-amino acids was developed through intermediate formation of the corresponding Ni(ii)-chelated Schiff bases. This advanced method works well in convenient conditions and allows the large-scale preparation of target α-amino acids in enantiomerically pure form.354  Highly efficient chiral resolution of D,L-Arg was carried out by cocrystal formation followed by recrystallization under preferential enrichment conditions.355  By utilizing the preferential enrichment technique, an improved enantiomeric resolution of DL-leucine (Leu) was achieved using a 1 : 1 cocrystal of D,L-Leu and oxalic acid.356 

Aqueous two-phase systems based on tropine type chiral ionic liquids and inorganic salt solution were designed and prepared for the enantiomeric separation of racemic Phe. The results indicate that D-enantiomer of Phe interacts more strongly with chiral ionic liquids and Cu2+.357  A new, recyclable solid–liquid resolution system was developed based on tropin ionic liquids for the enantiomeric resolution of several racemic amino acids (Phe, Trp, Tyr, and phenyl Gly). The system provides high resolution without organic solvent and recycle of all chemical materials.358  Tropine-type chiral ionic liquid with proline anion was immobilized on silica gel as adsorbent for separation by chemical modification method for the first time. The static experiment showed that adsorption rate of two enantiomers of the racemic amino acids was different.359  An aqueous two-phase system was designed and prepared for the enantiomeric separation of racemic amino acids. The system used task-specific hydrophilic ionic liquid and inorganic salt solution. The mechanism of separation was also studied.360  A novel solid-liquid two-phase system was developed for the chiral separation of racemic Phe with new dication imidazolium-based chiral ionic liquids in a solid-liquid two-phase system where copper ions represented the solid phase. The results indicated that L-enantiomer of Phe interacts more strongly with chiral ionic liquids and Cu2+.361  Diasteromerically pure γ-oxyfunctionalized α-amino acids were produced by a multi-enzymatic cascade reaction. Racemic N-acetylmethionine was quantitatively converted into L-methionine-(S)-sulfoxide by the cascade reaction under optimized conditions.362 

Research on application of amino acids and amino acid amides as chiral auxiliaries in cyanuric chloride based chiral derivatizing agents for enantioseparation was discussed in a review. The reagents gain advantage as chiral derivatizing agents (in terms of mild derivatization conditions for synthesis), stability of derivative, and high resolution over several other reagents.363  A new and efficient method was developed for the synthesis of racemic protected α-ethynyl Phe from D,L-2-benzyl Ser, in ten steps. Resolution was performed by HPLC using a chiral stationary phase.158 

Structurally simple, recyclable, and inexpensive chiral tridentate ligands were used for non-enzymatic dynamic kinetic resolution of unprotected racemic tailor-made α-amino acids.364  Enzymatic dynamic kinetic resolution of different racemic N-formyl- and N-carbamoyl-amino acids using a dynamic kinetic resolution approach was achieved. The enzymes (LN-carbamoylase and N-succinyl-amino acid racemase) were immobilized on two different solid supports.365  Since racemases allow racemization in one reaction step, using enzymatic catalysis for racemization can be very beneficial. The natural roles of racemases and their occurrence, the applications, and the biochemistry and engineering of this promising class of biocatalysts was summarized.366 

Pyridine-linked bis(β-cyclodextrin) copper(ii) complexes were reported that enantioselectively hydrolyse chiral amino acid esters.90  Monoterpene nitroso chlorides with α-amino acid derivatives form terpene-amino acid hybrids. Reaction with an excess of racemic dl-amino acids and their derivatives induces partial resolution of the amino acid components and formation of the diastereomeric mixtures of the terpene-amino acids hybrids.367 

Resolution of N,C-unprotected β-amino acids was accomplished through enantioselective formation and disassembly of nickel(ii) complexes under convenient conditions in good yields and excellent enantioselectivity. The method can be used for resolution of β-aryl, β-heteroaryl, and β-alkyl-derived β-amino acids.368 

Peptide-catalysed conversion of racemic oxazol-5(4H)-ones into enantiomerically enriched α-amino acid derivatives was reported.369  Racemic amino acids can be enantiomerically resolved by materials composed from polysaccharide containing cellulose and chitosan.370 

Sugar amino acids represent an important class of building blocks for the generation of peptide scaffolds and constrained peptidomimetics, owing to the presence of a relatively rigid furanoid or pyranoid ring decorated with space-oriented substituents. Glycosylation is one type of posttranslational modification of proteins. Understanding on the molecular level of the structural features of glycoproteins that are recognized by various enzymes and receptors would be valuable in developing inhibitor-based strategies to control carbohydrate-mediated cellular processes. This fundamental understanding leads to new therapeutic strategies for conditions that are characterized by abnormal glycosylation.

Amino acids and sugars are involved in complex biological processes such as catalysis and highly selective molecular recognition. Four ranunculins, novel hybrids of amino acids and sugars from Ranunculus ternatus Thunb, a plant used in traditional Chinese Medicine was reported. These four sugar amino acids possess a γ-aminobutyric acid (GABA) moiety and monosaccharide or disaccharide moieties and have potential tail to tail ether-connected (6,6-ether-connected) bonds. This represents an unprecedented structural phenomenon for the connections of sugars with the participation of γ-amino acids.371 

Branched-chain and aromatic amino acids are transformed into higher alcohols with yeast Saccharomyces cerevisiae in the Ehrlich pathway. Five specific metabolites of glycated amino acids were synthesized and characterized. It was shown for the first time that S. cerevisiae can use glycated amino acids as a nitrogen source and transform them into new metabolites, provided that the substances can be transported across the cell membrane.372  Cyclic functionalised sugar amino acids and their 3- to 6-membered nitrogen heterocyclic and carbocyclic analogues are used in the synthesis of peptidomimetics and glycomimetics. These derivatives of sugar amino acids provide access to hydrophilic and hydrophobic peptide isosteres.373 

A new synthetic route, useful derivatives, and coupling strategies were published for obtaining C-3 epimers of sugar amino acids as foldameric building blocks.374  A systematic conformational search was carried out for monomers and homohexamers of furanoid β-amino acids. The results show that hexamers of cis-furanoid β-amino acids show great variability, while hexamers of hydrophobic aminocyclopentane carboxylic acid and hydrophilic xylose sugar amino acid foldamers favour two different zigzagged conformations, the backbone fold turns into a helix in case of hexamer of β-aminotetrahydrofurancarboxylic acid.375 

The synthesis, purification, and characterization of a new lactosyl-derivative, i.e. a lactosyl thiophenyl-substituted triazolyl-thione L-Ala was described. This amino acid-sugar conjugate was prepared by solution synthesis analogue to the natural fructosyl-amino acids, tested oncologically, and showed significantly higher anticancer effect.376 

C-2 Deoxy glycosides and amino acid conjugates were synthesized by mediation of hypervalent iodine. E.g. 2-iodo serinyl glycosides was produced by this simple, efficient, and practical method (Scheme 19).377 

S-Glycosylated β(2,2)-amino acids were achieved with a sulphur-containing nucleophile by using 1-thio-β-d-glucopyranose derivatives. The glycosylation occurred with inversion of configuration at the quaternary centre.378 

Enantiopure β-amino acids represent interesting scaffolds for peptidomimetics, foldamers, and bioactive compounds. Because of their high biological potential, cyclic β-amino acids are of importance in medicinal chemistry. These compounds are both elements of bioactive products and building blocks in peptide research. Since β-amino acids are components of complex natural products; produce structural diversity in natural products; and provide characteristic architectures beyond those of α-l-amino acids, they have significant and unique biological functions in nature. The known bioactive β-amino acid-containing natural products (nonribosomal peptides, macrolactam polyketides, and nucleoside-β-amino acid hybrids), the biosynthetic enzymes that form β-amino acids from α-amino acids, the de novo synthesis of β-amino acids, and the mechanisms of β-amino acid incorporation into natural products are summarized in a review.15 

A mild organocatalyzed Mannich reaction provides direct and highly stereoselective access to acyclic β(2)- and β(2,3,3)-amino thioesters with adjacent tertiary and quaternary stereocenters. Mechanistic studies provided insight into the high stereochemical differentiation between the two ester moieties and showed that the stereochemistry can be controlled by the choice of the substrate. The β-amino thioesters can be used in coupling-reagent-free peptide synthesis.379  Highly stereoselective Mannich-type addition reactions of α-fluorinated monothiomalonates with N-Cbz- and N-Boc-protected imines under mild organocatalytic conditions resulted in α-fluorinated β-amino thioesters. Optimization of the stereoelectronic properties of the thioester moiety allowed to tune the reactivity of the α-fluoro-β-amino thioesters and enabled their coupling-reagent-free incorporation into peptides.204  Enantioselective Mannich reactions of aldehydes with in situ generated N-carbamoyl imines followed by a Horner–Wadsworth–Emmons reaction lead to chiral vinylogous β-amino acids that are useful building blocks for the construction of combinatorial libraries of peptidomimetic compounds.380  A broad range of highly functionalized β-amino acids containing α-fluorinated quaternary stereogenic carbon centres have been synthesized via a diastereoselective Mannich-type reaction of α-alkyl, α-aryl, and α-vinyl fluoroacetates with N-tert-butylsulfinyl imines. The stereochemical outcome of the reaction is highly dependent on the steric and electronic properties of the fluorocarbon nucleophiles.206  A diastereoselective addition reaction of fluoroacetate and α-alkylated fluoroacetate to N-tert-butylsulfinyl imines provides α-fluoro-β-amino acids.205 

β- and β(3)-amino acid analogues have been prepared by a method using copper(i)-catalyst. The substituted dihydropyrimidin-4-ones from propargyl amides led to the intermediate ketenimine under mild reaction conditions. The obtained substituted dihydropyrimidin-4-ones were elegantly transformed into the corresponding β- and β(3)-amino acid analogues.381 

The challenging intermolecular amination of inactivated C(sp(3))–H bonds has been achieved via Pd(ii) – catalysed intermolecular amination leading to a variety of unnatural β(2)-amino acid analogues. The reaction works without using of phosphine ligand or external oxidant.382  Rhodium-catalysed enantioselective hydrogenation of tetrasubstitued α-acetoxy β-enamido esters leads to chiral derivatives of α-hydroxyl-β-amino acid in excellent enantioselectivities.383 

An organocatalytic asymmetric aminomethylation of α,β-unsaturated aldehydes by N-heterocyclic carbene and Brønsted acid cooperative catalysis resulted in β-amino esters in good yields and high enantioselectivities. This redox-neutral strategy is conducted under mild reaction conditions and is suitable for the scalable production of enantiomerically enriched β-amino acids bearing various substituents.384  Asymmetric Friedel–Crafts alkylation of α-substituted β-nitroacrylates led to β(2,2)-amino acids bearing indolic all-carbon quaternary stereocenters.385  Novel stereoisomerically pure β′-hydroxy-β-amino acid derivatives have been produced by an easy, mild, and efficient method using dibutylboron triflate-mediated aldol reaction of suitably protected β-amino acids bearing chiral oxazolidinone. Both α,β-syn and α,β-anti isomers are accessible through the choice of the oxazolidinone chirality.386 

It was reported a new method for the synthesis of conformationally restricted β-amino acids involving a Vilsmeier–Haack reaction with nonaromatic carbon nucleophiles. The tertiary and all-carbon quaternary centres was successfully used to generate several β(2,2,3)-amino esters (e.g. derivatives of homoproline, homoalanine, and homopipecolinic esters).387 

A new type of arylboronic acid equipped with chiral aminothiourea was used for the first intermolecular asymmetric Michael addition of nitrogen-nucleophiles to α,β-unsaturated carboxylic acids. BnONH2 as a nucleophile gives a range of enantioenriched β-(benzyloxy)amino acid derivatives in good yields and with high enantioselectivity.388  Stereoselective synthesis of enantioenriched α-(silyloxy)-β-amino amides was developed by a silyllithium-initiated coupling of α-ketoamides with tert-butanesulfinylimines. The use of α-ketoamides is critical for achieving high yields and diastereoselectivities in the resulting α-hydroxy-β-amino acid derivatives.389  Four structures of oxoindolyl α-hydroxy-β-amino acid derivatives have been synthesized. The diastereoselectivity of the chemical reaction involving α-diazoesters and isatin imines in the presence of benzyl alcohol is confirmed through the relative configuration of the two stereogenic centres.390  The selective preparation of 1,4-disubstituted 1,2,3-triazoles attached to β-amino acids has been summarized from the corresponding alkynyl-β2-amino acids according to Huisgen's copper-catalysed 1,3-dipolar cycloaddition under mild conditions and with very high efficiency.255  Spontaneous rearrangement of 4-carboxy-2-oxoazepane α,α-amino acids resulted in 2’-oxopiperidine-containing β(2,3,3)-amino acids, upon basic or acid hydrolysis of the 2-oxoazepane α,α-amino acid ester. The reordering process involved the spontaneous breakdown of an amide bond, which typically requires strong conditions, and the formation of a new bond leading to the six-membered heterocycle.391 

N-substituted-β-amino acid derivatives containing 2-hydroxyphenyl, benzo[b]phenoxazine and quinoxaline moieties have been synthesized from 3-[(2-hydroxyphenyl)amino]butanoic and 3-[(2-hydroxy-5-methyl(chloro)phenyl)amino]butanoic acids. The novel compounds were tested for their antimicrobial and antifungal activities.266 

Cyclic β-amino acids are of importance in medicinal chemistry since they are elements of bioactive products and building blocks in peptide research. Catalytic cycloaddition of enecarbamates with electrophilic metalloenolcarbenes yields chiral cyclopentyl β-amino esters in excellent enantio- and diastereocontrol.392  Protected Orn-derived (3S,4S)-β-lactam was used as intermediate in the preparation of conformationally constrained (3S,4S)-2-oxoazepane α,α- and (2S,3S)-2-oxopiperidine-β(2,3,3)-amino acid derivatives. In the synthesis of these heterocyclic amino acids, the incorporation of a p-methoxyphenyl moiety is crucial for the excellent stereochemical purity.393  Diastereoselective synthesis of β-heteroaryl syn-α-methyl-β-amino acid derivatives (esters and amides) have been achieved via an addition reaction to five- and six-membered heterocyclic tert-butyl sulfinimines.394 

The non-proteinogenic β2,2-amino acid was produced via an enantioselective Rh(I)-catalyzed conjugate addition reaction of α-substituted β-nitroacrylates with various arylboronic acids by using chiral Rh(I) diene catalysts under mild conditions in a range of common organic solvents.395 

The research in the field of design and synthesis of unnatural amino acids is growing fast because of the increasing demand of proteins of potential therapeutics. Molecules of non-proteinogenic amino acids (e.g. β-N-methylamino-L-Ala) participate in various physiological processes and can produce adverse ecological effects. It is known that accumulation of β-N-methylamino-L-Ala via the food chain can lead to development of neurodegenerative diseases in humans. Natural sources of β-N-methylamino-L-Ala, methods for its detection, and possible mechanisms of toxicity in different living organisms are discussed in a review.396 

Although the asymmetric synthesis provides some efficient protocols, it is attractive to make unnatural chiral α-amino acids from available natural α-amino acids through keeping of the existing chiral α-carbon. 83 unnatural chiral α-amino acids were prepared at room temperature under visible-light assistance from derivatives of L-Asp and L-Glu.397  Efficient asymmetric synthesis of unnatural alkenyl amino acids has been achieved using alkylation of a fluorine-modified Ni(ii) Schiff base complex as the key step.398  New methods for the synthesis of unusual amino acids using copper(i)-catalyzed click reactions was summarized. This method is efficient for the synthesis of peptides and amino acids conjugated with carbohydrates, thymidine, and ferrocene.399  An improved second-generation synthesis of the unnatural amino acids AHMOD and AMD, components of the anticancer peptaibol culicinin D has been developed. A novel Wittig reagent for one-carbon homologation of aldehydes is also reported.400  The effect of replacing Trp with the non-natural analogue β-(1-azulenyl)-L-Ala was investigated in the well-known bee-venom peptide melittin. The results showed that β-(1-azulenyl)-L-Ala can serve as a solvent insensitive alternative to Trp that does not have significant impacts on structure or function of membrane interacting peptides.401  The non-proteinogenic amino acids phenyl-Gly and hydroxyphenyl-Gly are crucial components of certain peptidic natural products and are important for the preparation of various medicines. The conformational behaviour of these two non-proteinogenic amino acids was investigated that might help in designing of bioactive peptides and peptide based drugs.402 

Effects (on structure and function) of incorporation of non-natural amino acids into core region of enzyme was investigated.403  An iron-catalyzed diastereoselective synthesis of unnatural chiral (S)-α-amino acids with γ-quaternary carbon centres has been developed. The method shows some advantages: simple and wide substrates, mild conditions, high diastereoselectivity, and easy workup procedures.404  The synthesis of non-natural amino acid 2-amino-3,3,4-trimethyl-pentanoic acid has been developed via Cu(i) chloride Michael addition, followed by a Curtius rearrangement.405  Non-natural aryl Gly amino acids are achieved in three steps from aromatic aldehydes with N-(trimethylsilyl)imines as enantioselective catalyst.406  Ligand-controlled C(sp(3))–H bond arylation and olefination was developed for the synthesis of unnatural chiral α-amino acids with transition metal catalysts.407  Novel hydrophilic trans-cyclooctenenylated noncanonical amino acids have been designed, synthesized, tested, and incorporated into proteins.408 

Silicon-containing unnatural amino acids are becoming an interesting new class of building blocks. A review summarizes the different methods used to prepare silicon-containing amino acids and their implications on conformational structures and biological applications.409 

The dengue virus and West Nile Virus proteases are attractive targets for the development of dual-acting therapeutics against these viral pathogens. The synthesis and biological evaluation of inhibitors that contain benzyl ethers of 4-hydroxyphenyl-Gly as non-natural peptidic building blocks was published.410 

Nitrogen isotopic composition of amino acids has been widely applied to biochemical, ecological, archaeological, and biogeochemical studies to trace nitrogen source and transformation processes. For accurate isotope analysis of individual amino acids, a preparative method was validated involving the isolation of underivatized amino acids by ion-pair chromatographic separation and confirmed the consistency of nitrogen isotope composition. Ion-pair reversed-phase liquid chromatography coupled with electrospray ionization mass spectrometry (LC–ESI/MS) and gas chromatography/combustion coupled with isotope ratio mass spectrometry (GC/C/IRMS) were conducted for the purpose of separation of underivatized amino acids and nitrogen isotopic analysis, respectively.411 

An extra-facile chiral liquid chromatography–time of flight mass spectrometry (LC–TOF/MS) analytical method of amino acid enantiomers has been developed without a derivatization process. The enantioseparation of eighteen proteinogenic amino acids (except for Pro) was simultaneously performed using a combination of a chiral column (CROWNPAK CR-I(+)) and TOF/MS. An isocratic condition of a simple mobile phase comprising acetonitrile/water/trifluoroacetic acid gave baseline separation of all underivatized amino acid enantiomers on the chiral column.412 

Amino acids are an important and highly dynamic fraction of organic N in soils and their determination in soil without derivatization is challenging due to the difficulties in separation and detection of trace amounts of these polar analytes. An analytical method to quantify 20 free amino acids in aqueous soil extracts without derivatization was developed.413 

The quantitation of free amino acids from physiologic samples is essential for diagnosing and monitoring patients with inherited metabolic disorders. Current methods are hindered by long preparative and/or analysis times, expensive reagents, and often suboptimal performance characteristics. An improved method for amino acid analysis using liquid chromatography–tandem mass spectrometry (LC–MS/MS) has been developed and validated. Chromatographic separation of amino acids occurred using two columns. Eluted compounds were detected by selective reaction monitoring (SRM). Results showed excellent correlation with the Biochrom 30 amino acid analyser. The results show that the method is extremely sensitive, specific, reproducible and represents an improvement over other currently available technologies.414  A procedure was developed for the direct determination of dissolved free amino acids (DFAAs) in freshwater samples employing ion-pairing liquid chromatography and mass spectrometry. The approach allowed accurate quantification of subnanomolar concentrations of DFAAs without derivatization or sample clean-up steps. Ala, Ser, Glu, Arg, and Gly constituted 65% of the total pool, while Met and Trp occurred at sub-nM concentrations only. The composition of DFAAs significantly differed at all examined spatial scales, and this could be mainly attributed to Ala, Asp, and Gly. The method equals or outperforms existing ones in terms of sensitivity and reproducibility, and it is superior for the high-throughput analysis of freshwater samples.415 

A novel triazine-type chiral derivatization reagent, i.e., (S)-2,5-dioxopyrrolidin-1-yl-1-(4,6-dimethoxy-1,3,5-triazin-2-yl) pyrrolidine-2-carboxylate (DMT-(S)-Pro-OSu), was developed for the highly sensitive and selective detection of chiral amines and amino acids by ultra high performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) analysis. The enantiomers of amino acids were easily labelled with the reagents at room temperature. The diastereomers derived from proteolytic amino acids, except Ser, were well separated under isocratic elution conditions by reversed-phase chromatography. A highly sensitive detection was obtained from the SRM chromatogram. The chiral amines (e.g., adrenaline and noradrenaline) labelled with DMT-(S)-Pro-OSu were also well separated. Furthermore, D-Ala and D-Pro were also detected in relatively high concentrations. Consequently, DMT-(S)-Pro-OSu seems to be a useful chiral derivatization reagent for the determination of amines and amino acids in biological samples.416  An improved analytical procedure was developed for the resolution and quantification of amino acid enantiomers by multidimensional GC. The procedure contains a derivatization step, by which amino acids were transformed into N(O,S)-ethoxycarbonylheptafluorobutyl esters. This highly sensitive method was tested on a sample of the Murchison meteorite, for which obtained chromatograms show excellent peak resolution, minimal co-elution and peak overlap. The comprehensive two dimensional chromatography, in combination with the optimized derivatization method is a highly suitable technique for the analysis of samples with very limited quantities.417  Stereoisomers (enantiomers and diastereoisomers) of synthetic, non-protein amino acids comprising α-, β-, and γ-amino acids, including α,α-dialkyl amino acids, were converted into the respective N-trifluoroacetyl-O-methyl esters, analysed and resolved by GC on a commercial fused silica capillary column coated with the chiral stationary phase octakis(3-O-butyryl-2,6-di-O-pentyl)-γ-cyclodextrin. The chromatographic method presented is highly suitable for the elucidation of the stereochemistry of non-proteinogenic amino acids.418 

Rapid, easy, and reliable quantification of amino acids is crucial in research on plant amino acid metabolism and nutritional improvement of crops via enrichment of essential amino acids. A recently reported analytical method, based on solid phase extraction (SPE), derivatization with methyl chloroformate and gas chromatography–mass spectrometry (GC-MS) was optimized and tested on three-week-old Arabidopsis thaliana leaf tissues. Of the 16 selected amino acids, 14 were quantified successfully.419 

Recently, the demand for d-amino acid profiling has been drastically increasing. The present methodologies for d-amino acid profiling are still unsatisfactory. A novel method for d-amino acid profiling was developed by using a combination of a chiral column and TOF/MS. Based on the literature this method has the best performance for d-amino acid analysis that also includes the shortest analytical time, the highest enantioseparability without derivatization, and the largest coverage for analytical targets.420 

Most chiral amino acid separation techniques require complicated derivatization procedures to achieve the desirable chromatographic behaviour and detectability. A highly sensitive analytical method for the enantioseparation of chiral amino acids without any derivatization process using LC–MS/MS was developed. The results demonstrated the applicability and feasibility of the LC–MS/MS method as a novel, effective tool for d-amino acid measurement in various biological samples.421 

Ion-exchange chromatography (IEX) is a historical technique widely used for the detailed characterization of therapeutic proteins and can be considered as a reference and powerful method for the qualitative and quantitative evaluation of charge heterogeneity. Several analytical techniques have been used in amino acid geochronology to measure the relative abundances of D- and L-enantiomers. During the past two decades, reverse-phase (RP) liquid chromatography has become most common, whereas ion-exchange (IE) liquid chromatography was widely used prior to the mid-1990s. A new method was proposed based on intra-lab paired analyses of RP and IE liquid chromatography to mathematically convert A/I (allo-isoleucine: isoleucine) values determined with IE to corresponding D/L values for comparison with new RP results. Regression equations are provided to convert A/I to an equivalent D/L value for five amino acids, thereby enabling the large literature base of AAR results from IE chromatography to be compared and integrated with AAR results from RP chromatography.422 

The natural Cinchona alkaloid quinidine as chiral selector in chiral ligand-exchange chromatography was systematically studied. Chromatographic conditions for enantioseparation of twenty α-amino acids were first time studied by changing mobile phase parameters such as pH, concentration of organic solvent, type of salt, ligand to metal ratio and column temperature. Maximum retention and enantioselectivity factors were observed at the region close to pH=8, since the tertiary amine (the quinuclidinic nitrogen) of the quinidine is protonated only in a small degree, and therefore is available for the chelate formation.423 

Specific food proteins, peptides, and amino acids released by enzymatic hydrolysis have demonstrated several biological activities, therefore they represent an interesting perspective in agri-food, biopharmaceutical, and nutraceutical industries. However, these biomolecules are present at significantly lower concentration in a complex mixture of different sizes, conformations, and net charges. The conventional pressure-driven membrane processes are widely used as a first choice for the separation and purification of proteins, peptides, and amino acids. Innovative electro membrane processes such as EMF and EDFM are recently gaining much attentions and are regarded as favourable alternatives to the conventional methods. These methods could be potentially used for the efficient separation of charged compounds like proteins, peptides, and amino acids.424 

A conventional nonchiral column was used for the enantioseparation of several racemic α-amino acids (native and derivatized) using Cinchona alkaloids as chiral selectors along with Cu(ii) ions in chiral ligand-exchange chromatography. The mobile phase composition modulated retention and enantioselectivity. Good enantioseparation of many amino acids was obtained using equimolar amounts of Cu(ii) and either cinchonidine, quinine, or quinidine as chiral selectors in the mobile phase. Good correlations were obtained between experimental enantioselectivity factors and calculated energetic differences.425  The enantiomeric purity of N-α-Fmoc-protected amino acids is crucial from the viewpoint of peptide synthesis; therefore, a sensitive HPLC protocol was developed for the identification and quantification of enantiomeric impurities of commercially available N-α-Fmoc-protected amino acids on Cinchona alkaloid quinine-and quinidine-based weak anion exchanger-type chiral stationary phases. During the evaluation of the chiral chromatographic method, the effect of the mobile phase composition, nature, and concentration of different additives were optimized. The method permits detection of less than 0.01% enantiomeric impurity in the presence of the major enantiomer.426  Stereoselective HPLC and subcritical fluid chromatographic separations of 19 N-Fmoc proteinogenic amino acid enantiomers were carried out by using Quinidine-based zwitterionic and anion-exchanger-type chiral stationary phases Chiralpak ZWIX(-) and QD-AX. The effect of column temperature on the enantioseparation was investigated and thermodynamic parameters were calculated. The thermodynamic parameters revealed that the enantioseparations were enthalpy-driven. The elution sequence was determined in all cases and with the exception of Fmoc-Cys(Trt)-OH, it was identical on both chiral stationary phases whereby the L-enantiomers eluted before the D-enantiomers.427 

Thin-layer chromatography (TLC) is a simple and inexpensive technique permitting several samples to be handled simultaneously, thus yielding a higher precision than sequential analysis. The inert character of the thin-layer material makes it ideally suitable for use with strong corrosive reagents. Certain groups of interest can be chemically bonded to the reactive groups of support material, for example silanization for reversed-phase studies. Impregnation of the adsorbent with a variety of reagents adds an additional feature for influencing the adsorption characteristics without covalently affecting the inert character of the adsorbent. TLC is also successful in providing direct resolution of enantiomers of a variety of compounds by the proper manipulation of the support material.428 

High performance thin layer chromatography (HPTLC) is becoming increasingly popular amongst analysts for its simplicity of operation and capability of running several samples simultaneously. The introduction of newer spectroscopic detection methods and the combination of HPTLC with mass spectrometry (HPTLC-MS) has made this method useful for analysts. That is why HPTLC finds numerous applications in the pharmaceutical field. All analyses reported with HPLC are now being performed by HPTLC with a saving of time and cost.429  Impregnation of a stationary phase by organic and inorganic agents in HPTLC may result in higher separation selectivity and resolution. The influence of polymer structure in the stationary phase and a method of modification of the stationary phase on the efficiency of vitamin and amino acid determination and on the enantioselectivity factors of beta-blockers separation were investigated. It was established that such polymers can be used as modifying agents of HPTLC systems for on-line preconcentration of vitamins (B2) and amino acids (Lys, Trp and Glu). These polymers can also be recommended as chiral selectors for the effective TLC separation of beta-blockers.430 

In recent years, protein chemistry tends toward the analysis of more complex proteins, proteoforms, and posttranslational protein modifications. Although MS developed quite fast, sample preparation and separation of these analytes is still a major issue. For many years, electrophoresis seemed to be the method of choice. Similarly, two-dimensional separation can also be performed with TLC. As the revival of TLC developed enormously in the last decade, it seems to be also an alternative to use HPTLC for the separation of proteins. An HPTLC system was established, that allows a separation of protein mixtures over a broad polarity range, or if necessary allowing to modify the conditions with only few steps to improve the separation for a specific scope. Several layers and solvent systems have been evaluated to reach a fully utilized and optimized separation system.431 

A new analytical method for the analysis of 18 amino acids in natural waters using SPE followed by LC–MS/MS operated in multiple reaction monitoring mode was developed. Two different preconcentration methods, SPE, and concentration under reduced pressure were tested. SPE was a suitable extraction method for real samples due to the lower matrix effects. The SPE method still incorporates a broad sample clean-up and decreased endogenous matrix effects by reducing interferences originating from real water samples. The method limits of quantification (MLQ) for the SPE LC–MS/MS method in ultrapure water ranged from 0.1 to 100 μg L−1 as N for the different amino acids. The SPE LC–MS/MS method was successfully applied to the analysis of amino acids in 3 different drinking water source.432 

CHIRALPAK ZWIX(+) and ZWIX(−) are cinchona alkaloid-derived zwitterionic chiral stationary phases (CSPs) containing a chiral sulfonic acid motif which serves as negatively charged interaction site. An extensive experimental work aimed at developing schemes for an efficient generic screening and proposing straightforward approaches for method optimization on these ZWIX columns. Various chromatographic parameters were investigated using a large series of diverse amino acids and analogues. The involvement of acetonitrile (ACN) or tetrahydrofuran (THF) can help for adjusting retention time and selectivity. The presence of water in a low percentage is beneficial for peak shape, resolution, analysis speed, sample solubility, and MS detection performance.433 

Unusual amino acids play fundamental roles in many scientific fields. Since the single enantiomeric form can cause different and often serious response of organisms, chiral separations of unusual amino acids are irreplaceable tools. Two types of chiral stationary phases, two teicoplanin-based and four polysaccharide-based columns, were used. Separation conditions of RP mode, polar organic mode, and hydrophilic interaction chromatography were evaluated and compared. Teicoplanin-based chiral stationary phases, especially Chirobiotic T column, were able to separate almost all enantiomers tested, with the exception of Z-D,L-4-F-Phe ethyl ester.434  The successful enantioseparation of axially chiral amino acid derivatives containing a cyclohexylidene moiety on an analytical and semipreparative scale was achieved by HPLC using polysaccharide-based chiral stationary phases. Racemic methyl N-benzoylamino esters, easily obtained by methanolysis of the corresponding 5(4H)-oxazolones, were subjected to chiral HPLC resolution using chiral stationary phases based on immobilized 3,5-dimethylphenylcarbamate derivatives of amylase or cellulose. The behaviour of both selectors under different elution conditions was evaluated and compared.435  Trp and its eight derivatives are biologically important compounds. Since their enantiomers can exhibit different behaviour, efficient enantioselective separation methods are needed for both analytical and preparative purposes. In capillary electrophoresis (CE), cyclodextrins and their derivatives were proved to be suitable chiral selectors. Two pH values of background electrolytes were tested to affect ionization of the analytes and consequently their enantioseparation. Despite of the CE method was suitable for the enantioseparation, different separation systems/conditions were required. In HPLC, various separation modes and columns were used. The best results of enantioseparation of Trp and its amphoteric derivatives were achieved with teicoplanin based chiral stationary phases and methanol as a mobile phase.436 

The growing scientific attention in the biological function of d-amino acids leads to an increasing analytical interest for enantiomeric amino acid separation, which is challenging due to the lack of sufficiently sensitive, high-throughput analytical methods. A reversed phase ultra high performance liquid chromatography coupled with quadrupole–quadrupole time of flight mass spectrometry (RP-UHPLC-QqToF/MS) method using pre-column derivatization for very precise discrimination of amino acids enantiomers was developed. The method shows a superb sensitivity with limits of detection in the range of several pmol/l.437  The intrinsic D-amino acid profile of mouse macrophages was analysed using HPLC. Six d-amino acids (D-Ser, D-Asp, D-Leu, D-Ala, D-Lys, and D-Gln) were detected in cell lysates of mouse macrophages. The d-amino acid composition of RAW 264.7 cells, which is a model macrophage cell line, was like that of the mouse macrophage. The results suggest that macrophages and RAW 264.7 cells with macrophage-like functions have a similar D-amino acid profile.438 

A fully automated two-dimensional HPLC (2D-HPLC) was established by using silica-based monolithic ODS column as the first-dimension column with acetonitrile-trifluoro acetic acid–water as the mobile phase, micro Chiralpak QD-1-AX column as the enantiomer separation column with 10 mM citric acid in methanol-acetonitrile as the mobile phase for the second-dimension separation, and 4-fluoro-7-nitro-2,1,3- benzoxadiazole as the fluorometrical derivative reagent. The method has a higher separation efficiency and a higher detection sensitivity than that of existing methods in the determination of acidic amino acid enantiomers.439  A highly sensitive analytical method for the enantioseparation of chiral amino acids without any derivatization process using LC-MS/MS was developed. By optimizing MS/MS parameters, a quantification method that allowed the simultaneous analysis of 18 d-amino acids with high sensitivity and reproducibility was established.421 

A chromatographic analytical method for the direct determination of amino acids by hydrophilic interaction liquid chromatography (HILIC) was developed. A dual gradient simultaneously varying the pH 3.2 ammonium formate buffer concentration and level of acetonitrile in the mobile phase was employed. Analyte chromatographic parameters such as the sensitivity of retention to the water fraction in the mobile phase values HILIC were determined as part of method development. A degradation product of Gln (5-pyrrolidone-2-carboxylic acid) was observed and resolved chromatographically with no method modifications. The separation was used to quantitate amino acid content in acid hydrolysates of various protein samples.440  Rapid and simple quantitative analysis of intracellular metabolites is a critical tool for monitoring the alteration of biologically significant metabolites. An UHPLC method was established, equipped with HILIC column coupled to MS/MS. 19 amino acids and 2 related derivatives in human cell lines were simultaneously determined. Chromatographic separation was achieved within 20 min using a BEH amide column, with ammonium acetate and ammonium hydroxide as aqueous mobile phase, and acetonitrile as the organic mobile phase. Amino acids were analysed in positive ion multiple reaction monitoring (MRM) mode without the need of derivatization. The method was successfully applied to simultaneously detect the 21 compounds in a human colon cancer cell line DLD1.441 

The analysis of human plasma free amino acids is important for diagnosing the health of individuals, because their concentrations vary with various diseases. An amino acid analytical method was developed based on HPLC–ESI/MS. The method was also validated for 21 major types of free amino acids in human plasma samples. The results of the specificity, linearity, accuracy, repeatability, intermediate precision, reproducibility, and limits of detections were sufficient for the measurement of amino acids in human plasma samples and should be suitable for use in clinical fields.442 

As a common derivatization reagent, phenyl isothiocyanate (PITC) is widely used in the field of amino acid analysis. However, researchers have long faced the problem of coupling efficient separation with simultaneous, sensitive MS detection. A novel HPLC–ESI/MS method based on PITC derivatization was established for detecting amino acids. Complete separation of 15 amino acid derivatives was achieved. Therefore, the proposed method is a simple alternative to current methods of detecting amino acids.443 

A reference measurement procedure for amino acid quantification in blood samples was established based on deproteinization with 5-sulfosalicylic acid (SSA) and an isotope dilution-ultra performance LC–MS method. Five model amino acids (Val, Ile, Leu, Tyr, and Phe) in the SSA-treated samples showed ionization enhancement as well as stable background signals without significant ion suppression effects. Five analytes were clearly separated within 3 min using gradient elution and ion-pair chromatography. The method was applied to various blood samples including serum, whole blood, and plasma.444 

Ion-exchange HPLC generally fails as a method to determine low levels of free amino acids in body fluids. A modified RP-HPLC protocol for the determination of amino acids in body fluids and its application in mood disorder patients were developed. A previous research protocol was improved. The combination of the modifications, together with fluorescence detection (FLD), allows sensitive and practical determination of free amino acid levels in body fluids of depressive patients.445 

A sensitive RP liquid chromatography method for the determination and investigation of amino acids in wolfberry fruit (Lycium barbarum) after solid-phase extraction-derivatization was established. The investigation illustrated that each tested wolfberry fruit contained at least 16 amino acids and the main amino acids were Glu, Asp, Pro, Ala, Ser, Gly, Lys, and Tyr.446  An RP-HPLC method was developed for determining free amino acids in burley tobacco. The test was done with OPA/3-mercaptopropionic acid as the pre-column derivatizing reagent. The results of the determination show that seventeen kinds of free amino acids in burley leaves were produced.447  An analytical method for the simultaneous determination of free amino acids and biogenic amino acids (BA) in Cannonau and Vermentino wines was developed by using selective derivatization with dansyl chloride followed by HPLC with fluorescence detection. Thirty-two compounds were identified in the wines analysed. High levels of AA were found, with Pro being the most abundant. His was never detected in any Vermentino wines. γ-Aminobutyric acid, 4-hydroxy-Pro, Gly, Leu, Ile, and putrescine proved to be useful for differentiating Cannonau wines from Vermentino wines.448  A simple analytical method was proposed and validated for determining Trp in some cereal and legume samples. In the method, alkaline hydrolysis of proteins was used due to the destruction of Trp structure during acid hydrolysis. Separation and detection of Trp were performed on a RP column with fluorescence detection by using a mobile phase of acetonitrile and acetate buffer of pH 6.3.449 

The analysis of amino acids has become a central task in many aspects. While amino acid analysis has traditionally mainly been carried out using either GC in combination with flame ionization detection or LC with either post-column derivatization using ninhydrin or pre-column derivatization using o-phthalaldehyde, many of today's analysis platforms are based on chromatography in combination with MS. While derivatization is mandatory for the GC-based analysis of amino acids, several LC platforms have emerged, e.g. HILIC coupled to MS, allowing the analysis of underivatized amino acids. Among the numerous analytical methods available for amino acid analysis today, three prominent approaches, GC–MS, LC–MS, and HILIC–MS were compared.450 

A novel method was developed for the direct detection of amino acids in biological fluids by extractive electrospray ionization (EESI) tandem MS using minimal sample pretreatment. The EESI/MS conditions were optimized using representative amino acid standards. The methanol–water solvent was electrosprayed at positive ion detection mode. The temperature of the heated capillary was optimized. Collision induced dissociation (CID) experiments were performed by applying 17%–25% of the collision energy. The limit of detection (LOD) for the amino acids was in the range of 0.14-26.2 μg L−1. The average time for a single amino acid analysis was less than 0.5 min. The results showed that EESI/MS is a powerful tool for the rapid, sensitive, and quantitative detection of amino acids in complex biological samples.451 

A LC–MS analytical procedure has been developed for the detection and quantitative determination of underivatized amino acids at low concentrations in a standard reference material-urban dust. An accelerated solvent extraction followed by a solid phase extraction was applied prior to LC–MS/MS. Fourteen amino acids were separated by high resolution LC, detected and quantified by MRM on a triple quadrupole. This methodology avoids derivatization and allows the amino acid quantification in a complex matrix, and represent a good method suitable to analyse this class of compounds in atmospheric aerosol.452  A simple capillary electrophoresis (CE)-MS/MS method was developed for the analysis of free amino acids in commercial royal jelly (RJ) products. All 16 amino acids were determined without derivatization. The CE separation was achieved in an uncoated fused-silica capillary using a 1 M formic acid solution as the electrolyte, followed by MS/MS detection.453 

A preliminary investigation was undertaken to assess the performance of a new chromatography column technology in applications involving LC coupled to MS. The new column design allows mobile phase and solute to be extracted from the radial central region of the column, which reduces the solvent load to the mass spectrometer and improves separation efficiency.454 

A novel method based on the strategy of N-phosphorylation labelling was developed for quantification of twenty natural amino acids in human serum by RP-LC/ESI/MS/MS. Twenty N-phosphoryl amino acids were separated on an RP-C18 column within 20 min by isocratic elution. At the same time, MRM/MS enabled quantitation of twenty natural amino acids in human serum. All twenty amino acids were successfully detected in human serum samples.455 

Nitric oxide (NO) is a regulatory molecule involved in many biological processes. NO is produced by NO synthase by conversion of L-Arg to L-citrulline. Methylated derivatives of L-Arg, ADMA and SDMA, regulate L-Arg availability and the activity of NO synthase. A new multistep analytical methodology based on LC combined with MS was established for the accurate identification of the above compounds that were measured as stable 2,3,4,5,6-pentafluorobenzoyl chloride derivatives, which allows for simultaneous analysis of all compounds through chromatographic separation of ADMA and SDMA using a reverse phase column. This robust and fast LC-ESI/MS method may be a useful tool in quantitative analysis of L-Arg, ADMA, SDMA, and L-citrulline.456 

Trp is an essential amino-acid and the precursor of many biologically active substances such as kynurenine (KYN) and serotonin (5HT). Its metabolism is involved in different physiopathological cases, such as cardiovascular diseases, cancer, immunomodulation, or depression. A simple method was developed for quantification of Trp and 8 of its metabolites, involved in both KYN and 5HT pathways, using LC coupled to tandem MS. The method was also validated in human plasma samples. The method enables the detailed analysis of these metabolic pathways, which are thought to be involved in a number of pathological conditions.457 

Phthalic acid was introduced as a mobile phase additive to quantify free amino acids by HILIC coupled to ESI/MS/MS. The addition of phthalic acid significantly increased the signal intensity of protonated amino acid ions. Meanwhile, the shapes of chromatographic peaks of amino acids were optimized. This simple method was validated and successfully applied to the analysis of twenty-four free amino acids in human thyroid carcinoma and para-carcinoma tissues.458 

A new analytical method for the analysis of 18 amino acids in natural waters using SPE followed by LC–MS/MS was performed. The SPE LC–MS/MS method was successfully applied to the analysis of amino acids in 3 different drinking water source waters. Among the 18 amino acids analysed, the most abundant amino acids were found to be Tyr, Leu and Ile.432  An improved method for free amino acid analysis using LC–MS/MS has been developed and validated. Chromatographic separation of amino acids occurred using two columns, and the eluted compounds were detected by SRM, and quantitated by relating peak areas of amino acids to externally run standards.414 

Regular and accurate monitoring the levels of Phe and Tyr in blood is prerequisite for a successful management of patients with hyperphenylalaninemia (HPA). The MS/MS and the amino acid analyser (AAA) as methods to measure blood Phe and Tyr levels and Phe/Tyr ratio was compared. Venous blood samples were collected for the AAA analysis. Capillary blood was spotted directly on filter paper for the MS/MS analysis. 207 pairs of measurements were performed. The Phe and Tyr levels obtained by the MS/MS were on average 26.1% and 15.5% lower, respectively compared to those obtained by the AAA. The Phe/Tyr ratio by the MS/MS was on average 10.6% lower. Due to the considerable inter-assay variability, a single method is preferable for long-term follow-up of patients.459 

An RP-UHPLC-QqToF/MS method using pre-column o-phthalaldehyde/isobutyryl-l-cysteine (OPA/IBLC) derivatization for very precise discrimination of amino acid enantiomers was developed. The method shows a superb sensitivity with limits of detection in the range of several pmol/l.437 

A procedure for the direct determination of DFAAs in freshwater samples was developed employing ion-pairing LC–MS. The method approach accurate quantification of subnanomolar concentrations of DFAAs without prior concentration, derivatization or sample clean-up steps. DFAAs were separated on a C-18 resin using tridecafluoroheptanoic acid as an ion-pairing agent.415 

The sensitivity of coupled enantioselective capillary electrophoresis mass spectrometry (CE–MS) of amino acids is often blocked by the chiral selectors in the background electrolyte (BGE). A new CE–MS method is presented in which the use of a chiral selector is circumvented by involving (+)-1-(9-fluorenyl)ethyl chloroformate as chiral amino acid derivatizing agent and ammonium perfluorooctanoate (APFO) as a volatile pseudostationary phase for separation of the formed diastereomers.460 

An automated precolumn derivatization amino acid analytical method was developed based on HPLC–ESI/MS. This method enabled the separation of at least 38 types of physiological amino acids within 8 min. The method was also validated for 21 major types of free amino acids in human plasma samples.442 

A novel method for d-amino acid profiling using a combination of a chiral column and TOF/MS was developed. Based on the literature this method has the best performance for d-amino acid analysis.420 

During the ESI process, ions move through a heated capillary aperture to be detected on arrival at a mass analyser. However, the ESI process an ion cloud with an area larger than that of the heated capillary aperture, significantly contributing to an ion loss of 50% due to coulombic repulsion. To improve ion transmission, a novel method was proposed using a home-made golf ball positioned between the ion source and the inlet of the mass analyser to hydrodynamically focus the ions passing through the golf ball. The ion plume produced by the ESI that passes through the golf ball will reduce the size of the ion cloud then be focused and most of them flowed into the mass analyser. 20 trace amino acids in complex samples, including tea, urine and serum were determined. The results showed that the analytical performance of the determination of the 20 amino acids in samples using the home-made golf ball-assisted ESI source is better than that of a commercial ESI source.461 

Accurate assessment of mass isotopomer distributions (MIDs) of intracellular metabolites, such as free amino acids, is crucial for quantifying in vivo fluxes. A high-throughput LC–MS/MS method allowing the quantification of the levels and labelling of free amino acids was developed and validated. Sensitivity in the order of the femtomol was achieved using MRM mode. The method was applied to the determination of the 13C-labelling abundance in free amino acids extracted from maize embryos cultured with 13C-Gln or 13C-glucose. Although Cys was below the limit of detection in these biological samples, the MIDs of 18 free amino acids were successfully determined. This novel method will enable the assessment of more complete and accurate labelling information of intracellular amino acids.462 

Quantitative structure-retention relationships (QSRR) were constructed based on data obtained by a LC–ESI/QTOF/MS/MS method for the determination of amino acid analogues, following their derivatization via chloroformate esters. Chromatographic separation is based on gradient elution using methanol/water mixtures. The group of examined molecules was diverse, including mainly α-amino acids, but also β- and γ-amino acids and their analogues, decarboxylated and phosphorylated analogues, and dipeptides. Through stratified random sampling procedures, 57 compounds were split to a training set and a test set. Based on models, simplified equivalent was then created using a multi-linear regression (MLR) method. The suggested models are considered useful for the estimation of retention times of amino acid analogues for a series of applications.463 

An analytical method to quantify 20 free amino acids in aqueous soil extracts without derivatization was developed. The method employed HILIC–MS/MS technique combined with a cation exchange SPE. Good separation of 20 underivatized amino acids was achieved within 12 min. The method with high throughout and high analyte specificity shows great promise for consistent analysis of free amino acids.413  An UHPLC–HILIC–MS/MS method has been developed and validated for the quantification of 21 amino acids (20 proteinogenic amino acids and Cys) in their free form (FAA) and as protein constituents (total amino acids, TAA) in a rich protein food matrix such as lyophilized mussels samples. FAAs were analysed after suspending the samples in the presence of trichloroacetic acid, while TAAs were determined after acid hydrolysis with 6 M HCl. In hydrolysed samples 17 amino acids could be determined since Trp, Cys, cystine, and Asn were degraded during acid hydrolysis. The method proved to be a fast and reliable tool for acquiring information on free and total amino acids profile in high protein content foodstuffs such as mussels.464 

Cisplatin (cis-[PtCl2(NH3)2]) is an important platinum-containing anticancer drug for treatment of malignancies. Probing the interactions between cisplatin and free amino acids are beneficial for investigation of its pharmacology and side effects. The interactions of twenty amino acids with cisplatin in water, acidic, neutral and basic solutions were studied using ESI/MS. Adducts of cisplatin and amino acids were recognized. Multiple tandem mass spectrometry was employed to probe the affinities of nucleophilic functional groups, such as side chains, α-NH2 groups and α-COOH groups on amino acids for cisplatin in water. The results indicated that the chelated complexes formed were stable in aqueous solution.465 

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease that affects both lower and upper motor neurons, leading to muscle atrophy, paralysis, and death caused by respiratory failure or infectious complications. Altered levels of hCy, Cys, Met, and Glu have been observed in plasma of ALS patients. A method for determination of these potential biomarkers in plasma by CE–MS/MS was established. The validated method was applied to the analysis of plasma samples from a group of healthy individuals and patients with ALS, showing the potential of Glu and homocysteine metabolites as biomarkers for ALS.466 

Capillary array electrophoresis (CAE) is a promising technique for multiple enantiomeric separations. Carboxytetramethylrhodamine succinimidyl ester (TAMRA SE), a rhodamine-core fluorescent probe, has rarely been applied as an original precolumn derivatization reagent for chiral amino acid analysis so far. High-throughput enantiomeric separations of 12 TAMRA SE-AAs by a home-made 532 nm CAE-LIF scanner are presented. The effect of cyclodextrins and a variety of organic modifiers was quickly investigated. Multiple determination of the enantiomeric excess in non-racemic mixtures of Ala is successfully presented.467 

A new CE–MS method is developed in which the use of a chiral selector is circumvented by involving (+)-1-(9-fluorenyl)ethyl chloroformate as chiral amino acid derivatizing agent and ammonium perfluorooctanoate as a volatile pseudostationary phase for separation of the formed diastereomers. Selective detection and quantification of 14 chiral proteinogenic amino acids were achieved with chiral resolution. Asp and Glu were detected, but not enantioseparated.460 

New kinds of amino acid ionic liquids (AAILs) with pyridinium as cations and l-Lys as anion have been developed as the available chiral ligands coordinated with Zn(ii) in chiral ligand-exchange CE (CLE-CE). Four kinds of AAILs, including [1-ethylpyridinium][l-lysine], 1-butylpyridinium][l-lysine], [1-hexylpyridinium][l-lysine] and 1-[octylpyridinium][l-lysine], were successfully synthesized and characterized by nuclear magnetic resonance (NMR) and MS. Compared with other AAILs, the best chiral separation of l-amino acids could be achieved when [1-ethylpyridinium][l-lysine] was chosen as the chiral ligand.468  A CLE-CE method using Zn(ii) as the central ion and l-4-hydroxy-Pro as the chiral ligand coordinating with γ-cyclodextrin was developed for the enantioseparation of amino acids and dipeptides. After optimization, it has been found that eight pairs of labelled amino acids and six pairs of labelled dipeptides could be baseline-separated. Furthermore, the proposed method was applied in determining the enantiomeric purity of amino acids and dipeptides.469 

CE with ultraviolet detection was applied to determine underivatized amino acids in beer, based on the coordination interaction of Cu ions and amino acids. Using the United Nations Food Agriculture Organization/World Health Organization model of essential amino acid pattern and flavour of amino acids, the quality and taste in three kinds of beer were evaluated. It was found that the content of Phe, Pro, Ser and Ile was relatively large in all three kinds of beers with a great influence on beer flavour. This method was shown to be applicable to the separation of amino acids in beer and to perform quantitative analysis directly without derivatization.470 

A novel CE–MS/MS was established for the enantioseparation of enantiomers without derivatization for clinical purposes. Vancomycin chloride was used as an efficient chiral selector for the discrimination of the enantiomers by capillary electrophoresis employed with complete capillary filling method. Hyphenation of CE with MS/MS allows a reliable identification of separated enantiomers as well as their quantification.471 

Simple and inexpensive CE with UV-detection method (CE-UV) was optimized and validated for determination of six amino acids (Ala, Asn, Gln, Pro, Ser, and Val) for Sudanese food. Amino acids in the samples were derivatized with 4-chloro-7-nitro-2,1,3-benzoxadiazole (NBD-Cl) prior to CE-UV analysis. The validated method was successfully applied for the determination of amino acids for Sudanese food samples.472 

A method for determination of hCy, Cys, Met, and Glu as potential biomarkers of ALS in plasma by CE–MS/MS was established. All amino acids were separated within 25 min.466 

A method using 1H NMR spectroscopy has been developed to quantify simultaneously thirteen analytes in honeys without previous separation or pre-concentration steps. The method has been successfully applied to determine carboxylic acids, amino acids (Ala, Phe, Pro and Tyr), carbohydrates, ethanol, and hydroxymethylfurfural in eucalyptus, heather, lavender, orange blossom, thyme, and rosemary honeys. Quantification was performed by using the area of the signal of each analyte in the honey spectra, together with external standards. Good precision, with relative standard deviations over the range of 0.78–5.21% is obtained.473 

The amino acid composition of Nephila clavipes dragline silk fiber was determined by conducting 1H NMR spectroscopy experiments on acid-hydrolyzed material. This silk was found to consist of Gly, Ala, Glx, Leu, Tyr, Ser, Pro, Arg, Asx, Val, Thr, Phe, and Ile. Compared with standard chromatography-based AAA, the chemical resolution of NMR allows for an amino acid solution to be characterized without separation. In general, this 1H NMR AAA technique is applicable to a large range of proteins and peptides for precise composition characterization, especially when the precise content of a minor component is critical and relatively large amounts of sample are available (microgram to milligram quantities).474 

Amino acids, fully or partially neutralized with amines, are a new class of CO2 absorbents that have the possibility to combine the ionic character of amino acids with the better CO2 absorbing properties of the amines to form a more energy efficient solvent with improved environmental footprint. Equimolar mixtures of the amino acids, Gly, L-Ala, L-Pro, taurine, and L-Ser, with alkanolamines, monoethanolamine (MEA) and 2-amino-2-methyl-1-propanol (AMP), were quantitatively studied by 13C NMR at 25 °C. It is observed that in amino acid-MEA blended systems, carbamate formation from both MEA and the amino acids occur. The experimental data show that not only the pKa value of the amino acid is important but that also steric effects play a role in determining the preferred carbamate forming position. In the amino acid-AMP systems almost all carbamate is formed on the amino acid amine group due to the steric hindrance in AMP. Generally, addition of amine to an aqueous amino acid solution increases the amino acid solubility. NMR spectroscopy analysis showed that the precipitate consists mainly of amino acid and bicarbonate/carbonate with very small amounts of amine.475 

Amino acid ionic liquids (AAILs) have attracted significant attention in the recent literature owing to their ubiquitous applications in diversifying areas of modern chemistry, materials science, and biosciences. Noncovalent interactions accompanying Phe, Trp, and Tyr AAILs composed of 1-methyl-3-butyl-imidazole and its methyl-substituted derivative as cations have been analysed employing the dispersion corrected density functional theory. It has been shown that cation–anion binding in these bioionic ILs is primarily facilitated through hydrogen bonding in addition to lp—π and CH—π interactions those arising from aromatic moieties which can be probed through (1)H and (13)C NMR spectra calculated from the gauge independent atomic orbital method. It has been demonstrated that indirect spin–spin coupling constants across the hydrogen bonds correlate linearly with hydrogen bond distances. Besides the direction of frequency shifts of characteristic CO and NH stretching vibrations in the calculated vibrational spectra has been rationalized.476 

Binding interactions between twisted cucurbit[14]uril (tQ[14]) and twenty standard amino acids have been investigated by NMR spectroscopy and isothermal titration calorimetry (ITC) in aqueous HCl solutions and in DMSO. The results showed that tQ[14] displays weaker binding affinity for amino acids with hydrophobic or polar side chains and clear binding affinity for amino acids with a positively charged side chain or containing an aromatic ring, with the binding mode depending on the type of side chain present in the amino acids.477 

(2S,4R)- and (2S,4S)-perfluoro-tert-butyl 4-hydroxy-Pro were synthesized (as Fmoc-, Boc-, and free amino acids) in 2-5 steps. The key step of each synthesis was the incorporation of perfluoro-tert-butyl group, with nine chemically equivalent fluorines. Both amino acids were incorporated in model α-helical and poly-Pro helix peptides. Each amino acid exhibited distinct conformational preferences, with (2S,4R)-perfluoro-tert-butyl 4-hydroxyproline promoting poly-Pro helix. Peptides containing these amino acids were sensitively detected by (19)F NMR, suggesting their use in probes and medicinal chemistry.193 

The genetically encoded amino acid Sec and its dimeric form, selenocystine, are both utilized by nature. They are found in active sites of selenoproteins, enzymes that facilitate a diverse range of reactions, including the detoxification of reactive oxygen species and regulation of redox pathways. Due to Sec and selenocystine's specialized biological roles, it is of interest to examine their (77)Se NMR properties and how those can in turn be employed to study biological systems. A solid-state (77)Se NMR measurements was established of the L-selenocystine chemical shift tensor, which provides the first experimental chemical shift tensor information on Sec-containing systems. Quantum chemical calculations of L-Sec models were performed to help understand various structural effects on (77)Se L-selenocystine's chemical shift tensor. These results suggest that the dihedral information may be deduced for a protein with appropriate structural models. These first-time experimental and theoretical results will facilitate future NMR studies of selenium-containing compounds and proteins.478 


Amino acid analyser


Asymmetric dimethyl-Arg


Amyotrophic lateral sclerosis


2-Amino-3-(3-hydroxy-5-methyl-isoxazol-4-yl)propanoic acid


Capillary electrophoresis


Chiral ligand-exchange


Dissolved free amino acid


Dithiol amino acid


Extractive electrospray ionization


γ-Aminobutyric acid


Gas chromatography


Hydrophilic interaction liquid chromatography


High performance thin layer chromatography


Ion-exchange chromatography


Liquid chromatography–electrospray ionization mass spectrometry


Liquid chromatography-tandem mass spectrometry


Liquid chromatography–time of flight mass spectrometry


Mycosporin-like amino acid








Multiple reaction monitoring


Mycobacterium tuberculosis


N-Methyl d-aspartic acid




Positron emission tomography








Reversed phase ultra-high performance liquid chromatography–quadrupole–quadrupole time of flight mass spectrometry


Structure–activity relationship


Symmetric dimethylarginine




Selected reaction monitoring


solid phase extraction




Thin layer chromatography–mass spectrometry


Ultra-high performance liquid chromatography–tandem mass spectrometry.


Papers published during 2013–2016 have been sourced mainly from the Web of Science databases1  and Pubmed2  on the internet and from scanning a selection of major journals.

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