Hyphenated Analytical Techniques Research Articles

Analytical Insights: Ce-Ms Unveiled

Hyphenated techniques in the field of analytical chemistry involve combining different analytical methods to enhance the analysis of samples. One common approach is to combine chromatographic techniques with spectroscopic techniques. This makes it possible to comprehend the component included in a mixture on a more thorough level. It contains the various techniques such as GC-MS, LC-MS, LC-NMR, CE-MS, and LC-IR etc. Overall hyphenated technique offer improved analysis of samples in terms of specificity, accuracy, and precisions by combining multiple analytical methods, scientists can obtain more comprehensive information about the components. Capillary electrophoresis-mass spectrometry (CE-MS) has emerged as a powerful analytical tool for the comprehensive analysis of complex biological samples. This review provides a comprehensive overview of the latest advances in sophisticated hyphenated analytical techniques within the realm of CE-MS. It highlights the synergistic combination of capillary electrophoresis with mass spectrometry, showcasing its applications in proteomics, metabolomics, pharmaceutical analysis, and clinical diagnostics.

Assessing bioenergy prospects of algal biomass and yard waste using an integrated hydrothermal carbonization and pyrolysis (HTC–PY): A detailed emission–to–ash characterization via diverse hyphenated analytical techniques and modelling strategies

Hydrothermal carbonization (HTC) presents a promising method for converting carbonaceous waste into renewable fuels, facilitating energy recovery. This study focuses on the HTC pretreatment of microalgae (Chlorella sorokiniana) and yard waste, examining their physicochemical structural changes, component migration, and potential for valorization via integrated HTC and pyrolysis processes. HTC notably enhances the carbon content while reducing oxygen and nitrogen levels in these feedstocks, producing a clean solid fuel with high energy density. The study investigates the pyrolytic behavior of hydrochars and their blends using different analytical techniques, kinetic modeling, and fixed–bed reactor experiments. The blends with 90% and 70% of microalgae hydrochar (CSH) exhibited the most significant synergistic effects during co–pyrolysis, evident in its devolatilization behaviors and reduced activation energies. Integrating artificial neural networks and response surface methodology models facilitated optimization of pyrolysis parameters and accurate prediction of bio–oil, biochar, and gas yields. The analytical techniques enabled characterization of pyrolysis products, revealing notable changes in bio–oil and biochar compositions, suggesting potential enhancements. The study comprehensively evaluates the HTC–pyrolysis approach’s potential for converting microalgae and lignocellulosic biomass into eco–friendly biofuels for industrial applications.

Chemometric Approach for Profiling of Metabolites of Potential Antioxidant Activity in Apiaceae Species Based on LC-PDA-ESI-MS/MS and FT-NIR

Chemometrics is a tool for data mining and unlocking the door for solving big data queries. Apiaceae is a family species which is commonly cultivated worldwide. Although members of this species are widely used as antioxidant, antibacterial, antifungal, and anti-inflammatory agents, their metabolites profiling remains ambiguous. Based on WHO support, chemometrics has been used in evaluating the quality and authenticity of the herbal products. The objective of this study is to profile and characterize phenolic metabolites in nine species from Egyptian cultivars and three different species of German cultivars from the Apiaceae family using multivariate analysis after LC-PDA-ESI-MS/MS and near infrared spectroscopy data are generated. Principal component analysis was successfully applied to distinguish between the nine Egyptian cultivars and the three German cultivars, and hierarchical cluster analysis also confirmed this distinctive clustering. Partial least square regression (PLS-R) models showed a relationship between phytochemicals and antioxidant activities. The metabolites responsible for the clustering pattern and variables important for projection (VIP) were identified, being twelve amongst nine Egyptian cultivar samples and thirteen amongst the Egyptian cultivar and the German cultivar comparison. The identified VIPs were also correlated with the antioxidant activity using PLS-R. In conclusion, the study showed novelty in the application of hyphenated analytical techniques and chemometrics that assist in quality control of herbal medicine.

Structural Elucidation of Relevant Gibberellic Acid Impurities and In Silico Investigation of Their Interaction with Soluble Gibberellin Receptor GID1.

Gibberellin derivatives are a family of tetracyclic diterpenoid plant hormones used in agriculture as plant growth regulators included in the European Directive 91/414. In the pesticide peer review process and to assess their toxicological relevance and product chemical equivalence, the European Food Safety Authority (EFSA) highlighted data gaps such as the identification of hydrolysis products and unknown impurities. The aspect of impurity characterization and quantitation is challenging and requires the use of hyphenated analytical techniques. In this regard, we used an LC-QTOF/MS and NMR analysis for the characterization of gibberellic acid impurities found in technical products. Gibberellic acid impurities such as gibberellin A1 (GA 1 ), 3-isolactone gibberellic acid (iso-GA 3 ), gibberellenic acid, 1α,2α-epoxygibberellin A3 (2-epoxy- GA 3 ), and (1α,2β,3α,4bβ,10β)-2,3,7-trihydroxy-1-methyl-8-methylenegibb-4-ene-1,10-dicarboxylic acid were identified and successfully characterized. Moreover, an in silico investigation on selected gibberellic acid impurities and derivatives and their interactions with a gibberellin insensitive dwarf1 (GID1) receptor has been carried out by means of induced fit docking (IFD), generalized-Born surface area (MM-GBSA), and metadynamics (MTD) experiments. A direct HPLC method with DAD and MS for the detection of gibberellic acid and its impurities in a technical sample has been developed. Moreover, by means of the in silico characterization of the GID1 receptor-binding pocket, we investigated the receptor affinity of the selected gibberellins, identifying compounds (2) and (4) as the most promising hit to lead compounds.

Use of hyphenated analytical techniques to identify the bioactive constituents of Gunnera perpensa L., a South African medicinal plant, which potently inhibit SARS-CoV-2 spike glycoprotein-host ACE2 binding.

SARS-CoV-2, the causative agent of COVID-19, continues to cause global morbidity and mortality despite the increasing availability of vaccines. Alongside vaccines, antivirals are urgently needed to combat SARS-CoV-2 infection and spread, particularly in resource-limited regions which lack access to existing therapeutics. Small molecules isolated from medicinal plants may be able to block cellular entry by SARS-CoV-2 by antagonising the interaction of the viral spike glycoprotein receptor-binding domain (RBD) with the host angiotensin-converting enzyme II (ACE2) receptor. As the medicinal plant Gunnera perpensa L. is being used by some South African traditional healers for SARS-CoV-2/COVID-19 management, we hypothesised that it may contain chemical constituents that inhibit the RBD-ACE2 interaction. Using a previously described AlphaScreen-based protein interaction assay, we show here that the DCM:MeOH extract of G. perpensa readily disrupts RBD (USA-WA1/2020)-ACE2 interactions with a half-maximal inhibition concentration (IC50) of < 0.001 µg/mL, compared to an IC50 of 0.025 µg/mL for the control neutralising antibody REGN10987. Employing hyphenated analytical techniques like UPLC-IMS-HRMS (method developed and validated as per the International Conference on Harmonization guidelines), we identified two ellagitannins, punicalin (2.12% w/w) and punicalagin (1.51% w/w), as plant constituents in the DCM:MeOH extract of G. perpensa which antagonised RBD-ACE2 binding with respective IC50s of 9 and 29 nM. This good potency makes both compounds promising leads for development of future entry-based SARS-CoV-2 antivirals. The results also highlight the advantages of combining reverse pharmacology (based on medicinal plant use) with hyphenated analytical techniques to expedite identification of urgently needed antivirals.Supplementary InformationThe online version contains supplementary material available at 10.1007/s00216-022-04041-3.

Current and potential applications of simultaneous DSC-FTIR microspectroscopy for pharmaceutical analysis.

Quality control (QC) is the most important key issue in the pharmaceutical industry to ensure the quality of drug products. Many analytical instruments and techniques in pharmaceutical analysis are applied to assess the quality and quantity of the drugs. In the current and future trends, a combination of digitization, automation and hyphenation with high throughput on-line performance will be the topics for the future of pharmaceutical QC. The hyphenated analytical techniques have recently received great attention as unique means to solve complex analytical problems in a short period of time. This review article is an update on the recent potential applications of hyphenated technique developed from the coupling of a rapid separation or induction technique (differential scanning calorimetry; DSC) and an on-line spectroscopic (Fourier transform infrared; FTIR) detection technology to carry out an one-step solid-state analysis in pharmaceutical formulation developments, including (1) intramolecular condensation of pharmaceutical polymers, (2) intramolecular cyclization of drugs and sweetener, (3) polymorphic transformation of drugs and excipients, (4) drug-polymer (excipient) interaction, (5) fast cocrystal screening and formation. This simultaneous DSC-FTIR microspectroscopy can also provide an easy and direct method for one-step screening and qualitative detection of drug stability in real time.

Automated hyphenation of in-tube boronate affinity solid-phase microextraction with high performance liquid chromatography for the determination of cis-diol compounds in tea beverages

Automatic hyphenated analytical techniques hold great promise for reducing labour intensity, enhancing efficiency, and ensuring data reproducibility. In recent decade, Boronate affinity solid phase microextraction (BA-SPME) has emerged as a unique tool for the selective extraction of cis-diol-containing compounds. However, automatic hyphenation of SPME with high performance liquid chromatography (HPLC) has not yet been reported. Herein we report a new automated hyphenation of in-tube BA-SPME with HPLC for the determination of cis-diol compounds in tea beverages. The automated hyphenation was implemented through a flow-switch via the six-way valve of an autosampler. We prepared an in-tube SPME capillary, investigated the loading capacity of the coated column, characterized the morphology of the column, and investigated the factors influencing the separation. Finally, using the automatic hyphenated SPME-HPLC approach, we analyzed the cis-diol-containing compounds in three kinds of commercial tea beverages and investigated the effect of tea-making temperature on the content of the cis-diol-containing compounds in the prepared tea.

Understanding Biotic Stress and Hormone Signalling in Cassava (Manihot esculenta): Potential for Using Hyphenated Analytical Techniques

Biotic stresses often constitute major factors limiting global crop yields. A better understanding of plant responses to these stresses will facilitate efforts to improve stress tolerance and yields, especially in a climatically changing world. Numerous attempts have been made to confer tolerance/resistance to biotic stresses using both traditional and modern breeding methods. Mechanisms of biotic stress tolerance controlled by signalling networks and the analysis of genes controlling the yield and biotic stress tolerance are discussed. This review presents a report on the hormonal response of cassava to biotic stresses and the potential use of hyphenated analytical techniques to understand biotic stress hormonal responses. Hyphenated analytical techniques are reliable tools for understanding the response of cassava to biotic stresses, thereby accelerating the process of the development of biotic stress-tolerant/resistant genotypes for breeding purposes.

Omics to address the opportunities and challenges of nanotechnology in agriculture

Increasing global food demand, and risks associated with climate change and agrochemicals demand novel and sustainable agricultural practices to improve crop yield and quality. Various strategies using nanotechnology have been explored widely to provide solutions. However, their application in the food industry is constrained by limited understanding of nanomaterial safety. Growing interest in the potential of engineered nanomaterials (ENMs) in agricultural applications has also resulted in increased crop exposure, leading to unknown risks to plants, animals and humans. Recently, considerable research has been carried out on overt plant response to ENMs; however, conclusive mechanistic information is lacking. With advancements in hyphenated analytical techniques, research on ENM-biota interactions has witnessed a paradigm shift from low-throughput, single end-point bioassays to high-throughput, discovery-oriented omic tools, including transcriptomics, proteomics and metabolomics. This review summarizes the pioneering studies utilizing omics in plants exposed to ENMs to explain phenotypic expressions and elucidate associated biological pathways. Advantages and challenges of current analytical techniques employed in proteomics and metabolomics are also discussed. We acknowledge the need for integration and application of multiomics to identify sensitive biomarkers in plants in response to ENMs, and provide mechanistic insights, in order to design enhanced and safer nano-enabled products for agriculture.

Prediction of designer drugs: Synthesis and spectroscopic analysis of synthetic cathinone analogs that may appear on the Swedish drug market.

The use of hyphenated analytical techniques in forensic drug screening enables simultaneous identification of a wide range of different compounds. However, the appearance of drug seizures containing new substances, mainly new psychoactive substances (NPS), is steadily increasing. These new and other already known substances often possess structural similarities and consequently they exhibit spectral data with slight differences. This situation has made the criteria that ensure indubitable identification of compounds increasingly important. In this work, 6 new synthetic cathinones that have not yet appeared in any Swedish drug seizures were synthesized. Their chemical structures were similar to those of already known cathinone analogs of which 42 were also included in the study. Hence, a total of 48 synthetic cathinones making up sets of homologous and regioisomeric compounds were used to challenge the capabilities of various analytical techniques commonly applied in forensic drug screening, ie, gas chromatography-mass spectrometry (GC-MS), gas chromatography-Fourier transform infrared spectroscopy (GC-FTIR), nuclear magnetic resonance (NMR), and liquid chromatography quadrupole time-of-flight mass spectrometry (LC-QTOF-MS). Special attention was paid to the capabilities of GC-MS and GC-FTIR to distinguish between the synthetic cathinones and the results showed that neither GC-MS nor GC-FTIR alone can successfully differentiate between all synthetic cathinones. However, the 2 techniques proved to be complementary and their combined use is therefore beneficial. For example, the structural homologs were better differentiated by GC-MS, while GC-FTIR performed better for the regioisomers. Further, new spectroscopic data of the synthesized cathinone analogs is hereby presented for the forensic community. The synthetic work also showed that cathinone reference compounds can be produced in few reaction steps.

Identification of Impurities and Degradation Products in Pharmaceutical Products-Role of Hyphenated Techniques

Impurity profiling and identification of degradation product is an essential part of pharmaceutical development program. Impurity profiling is mandatory requirement by various regulatory agencies and is directly related with the quality, safety and efficacy of a drug product. The ultimate purpose of impurity profiling and stress testing is in the establishment of the degradation products pathways and to investigate the stability-indicating power of the analytical procedures which is ultimately helpful in the prediction of shelf life of drug product. Hyphenated technique is combination a combination of two different analytical techniques with the help of proper interface. Mainly chromatographic techniques are combined with the spectroscopic techniques. This write up provides a review on brief information about role of different hyphenated analytical techniques such as HPLC-UV, HPLC-MS, GC-MS and UPLC-MS/MS for identification of impurities and degradation products in pharmaceutical products.

Application of Bioassay-Guided Fractionation Coupled with a Molecular Approach for the Dereplication of Antimicrobial Metabolites

A systematically delineated dereplication approach was described based on genome mining and bioassay-guided fractionation using endophytic fungus Xylaria psidii FPL-52(S) isolated from leaves of Ficus pumila Linn., (Moraceae). A polyketide synthase gene-based molecular screening strategy by a degenerate oligonucleotide primer polymerase chain reaction technique coupled with a bioinformatic phylogenomic approach revealed the presence of an iterative polyketide synthase gene within the genome of Xylaria psidii FPL-52(S). Chemical dereplication of ethyl acetate extract derived from a submerged fermentation culture broth of Xylaria psidii FPL-52(S) by bioassay-guided chromatographic and hyphenated analytical spectroscopic techniques led to the identification of polyketide mycoalexin 3-O-methylmellein. Antimicrobial profiling and minimal inhibitory concentration values for 3-O-methylmellein were determined by disc diffusion and microbroth dilution techniques. Gram-positive bacteria, dermatophytic and phytopathogenic fungi were susceptible in terms of inhibition zone and minimum inhibitory concentration values when compared to co-assayed standards. Herein, we highlight and demonstrate an improved approach which facilitates efficient dereplication and effect-guided fractionation of antimicrobial metabolite(s). The present work flow serves as a promising dereplication tool to survey the biosynthetic potential of endophytic fungal diversity, thereby identifying the most promising strains and prioritizing them for novel polyketide-derived antimicrobial metabolite discovery.

Characterization of rhamnolipids by liquid chromatography/mass spectrometry after solid-phase extraction.

Rhamnolipids are surface-active agents with a broad application potential that are produced in complex mixtures by bacteria of the genus Pseudomonas. Analysis from fermentation broth is often characterized by laborious sample preparation and requires hyphenated analytical techniques like liquid chromatography coupled to mass spectrometry (LC-MS) to obtain detailed information about sample composition. In this study, an analytical procedure based on chromatographic method development and characterization of rhamnolipid sample material by LC-MS as well as a comparison of two sample preparation methods, i.e., liquid-liquid extraction and solid-phase extraction, is presented. Efficient separation was achieved under reversed-phase conditions using a mixed propylphenyl and octadecylsilyl-modified silica gel stationary phase. LC-MS/MS analysis of a supernatant from Pseudomonas putida strain KT2440 pVLT33_rhlABC grown on glucose as sole carbon source and purified by solid-phase extraction revealed a total of 20 congeners of di-rhamnolipids, mono-rhamnolipids, and their biosynthetic precursors 3-(3-hydroxyalkanoyloxy)alkanoic acids (HAAs) with different carbon chain lengths from C8 to C14, including three rhamnolipids with uncommon C9 and C11 fatty acid residues. LC-MS and the orcinol assay were used to evaluate the developed solid-phase extraction method in comparison with the established liquid-liquid extraction. Solid-phase extraction exhibited higher yields and reproducibility as well as lower experimental effort.

A selective and sensitive LC-MS/MS method for the simultaneous determination of twopotential genotoxic impurities in celecoxib

Abstract Background Impurity profiling is now receiving critical attention from regulatoryauthorities. For trace level quantification of potential genotoxic impurities(PGIs), conventional analytical techniques like high-performance liquidchromatography (HPLC) and gas chromatography (GC) are inadequate; consequently,there is a great need to apply hyphenated analytical techniques to developsensitive analytical methods for the analysis of pharmaceuticals. Methods A selective and sensitive liquid chromatography-tandem mass spectrometry(LC-MS/MS) method was developed for the simultaneous determination of(4-sulfamoylphenyl)hydrazine hydrochloride (SHH) and(4-methyl-acetophenone)para-sulfonamide phenylhydrazine hydrochloride(MAP) PGIs in celecoxib active pharmaceutical ingredient (API). The LC-MS/MSanalysis of SHH and MAP PGIs was done on Symmetry C18 (150 mm ×4.6 mm, 3.5 μm) analytical column, and the mobile phase used was5.0 mM ammonium acetate-acetonitrile in the ratio of 30:70(v/v). The flow rate used was 0.7 mL/min. Triplequadrupole mass detector coupled to positive electrospray ionization operated inmultiple reaction monitoring (MRM) mode was used for the quantification of SHH andMAP PGIs. The method was validated as per International Conference onHarmonization (ICH) guidelines and was able to quantitate both SHH and MAP PGIs at1.0 ppm with respect to 10 mg/mL of celecoxib. Results The proposed method was specific, linear, accurate, precise, and robust. Thecalibration curves show good linearity between the concentration range of 0.06 and7.5 ppm for both SHH and MAP PGIs. The correlation coefficient obtained was&gt;0.9998 in each case. The method has very low limit of detection (LOD) andlimit of quantification (LOQ). The obtained LOD and LOQ values were 0.02 and0.06 ppm, respectively, for both SHH and MAP PGIs. For both the PGIs,excellent recoveries of 95.0% to 104.0% were obtained at a concentration range of0.06 to 3.0 ppm. The developed method was also applied to determine the SHHand MAP PGIs in three formulation batches of celecoxib. Conclusions The proposed method is simple and accurate and is a good quality control tool forthe simultaneous quantitative determination of SHH and MAP PGIs at very low levelsin celecoxib during its manufacturing.

Chemometrics tools used in analytical chemistry: an overview.

This article presents various important tools of chemometrics utilized as data evaluation tools generated by various hyphenated analytical techniques including their application since its advent to today. The work has been divided into various sections, which include various multivariate regression methods and multivariate resolution methods. Finally the last section deals with the applicability of chemometric tools in analytical chemistry. The main objective of this article is to review the chemometric methods used in analytical chemistry (qualitative/quantitative), to determine the elution sequence, classify various data sets, assess peak purity and estimate the number of chemical components. These reviewed methods further can be used for treating n-way data obtained by hyphenation of LC with multi-channel detectors. We prefer to provide a detailed view of various important methods developed with their algorithm in favor of employing and understanding them by researchers not very familiar with chemometrics.

Automated Solid Phase Extraction (SPE) LC/NMR Applied to the Structural Analysis of Extractable Compounds from a Pharmaceutical Packaging Material of Construction

The structural analysis (i.e., identification) of organic chemical entities leached into drug product formulations has traditionally been accomplished with techniques involving the combination of chromatography with mass spectrometry. These include gas chromatography/mass spectrometry (GC/MS) for volatile and semi-volatile compounds, and various forms of liquid chromatography/mass spectrometry (LC/MS or HPLC/MS) for semi-volatile and relatively non-volatile compounds. GC/MS and LC/MS techniques are complementary for structural analysis of leachables and potentially leachable organic compounds produced via laboratory extraction of pharmaceutical container closure/delivery system components and corresponding materials of construction. Both hyphenated analytical techniques possess the separating capability, compound specific detection attributes, and sensitivity required to effectively analyze complex mixtures of trace level organic compounds. However, hyphenated techniques based on mass spectrometry are limited by the inability to determine complete bond connectivity, the inability to distinguish between many types of structural isomers, and the inability to unambiguously determine aromatic substitution patterns. Nuclear magnetic resonance spectroscopy (NMR) does not have these limitations; hence it can serve as a complement to mass spectrometry. However, NMR technology is inherently insensitive and its ability to interface with chromatography has been historically challenging. This article describes the application of NMR coupled with liquid chromatography and automated solid phase extraction (SPE-LC/NMR) to the structural analysis of extractable organic compounds from a pharmaceutical packaging material of construction. The SPE-LC/NMR technology combined with micro-cryoprobe technology afforded the sensitivity and sample mass required for full structure elucidation. Optimization of the SPE-LC/NMR analytical method was achieved using a series of model compounds representing the chemical diversity of extractables. This study demonstrates the complementary nature of SPE-LC/NMR with LC/MS for this particular pharmaceutical application. The identification of impurities leached into drugs from the components and materials associated with pharmaceutical containers, packaging components, and materials has historically been done using laboratory techniques based on the combination of chromatography with mass spectrometry. Such analytical techniques are widely recognized as having the selectivity and sensitivity required to separate the complex mixtures of impurities often encountered in such identification studies, including both the identification of leachable impurities as well as potential leachable impurities produced by laboratory extraction of packaging components and materials. However, while mass spectrometry-based analytical techniques have limitations for this application, newer analytical techniques based on the combination of chromatography with nuclear magnetic resonance spectroscopy provide an added dimension of structural definition. This article describes the development, optimization, and application of an analytical technique based on the combination of chromatography and nuclear magnetic resonance spectroscopy to the identification of potential leachable impurities from a pharmaceutical packaging material. The complementary nature of the analytical techniques for this particular pharmaceutical application is demonstrated.

Isoflavonoids of the Leguminosae

This account describes 275 new isoflavonoids published between 2008 and 2011 as constituents of the Leguminosae, commenting on their source, identification, biological activity, synthesis, and ecological or chemosystematic significance. Applications of hyphenated analytical techniques to the characterisation of Leguminosae isoflavonoids are also reviewed, together with advances in biosynthetic studies. A checklist of new compounds by species is given, and 226 references are cited.

Organometallic derivatizing agents in bioanalysis

Over the last few decades, the development of several innovative hyphenated analytical techniques and their routine use in laboratories has led to new possibilities for the quantitative analysis of biomolecules. Today, the identification and quantification of biomolecules such as peptides and proteins are essential to answer important medical, pharmaceutical, and biological questions. To allow efficient detection and structure elucidation of biomolecules, several approaches including derivatization strategies were investigated and applied during recent years. This article summarizes the current approaches for labeling and presents the different types of organometallic derivatizing agents used as labels. Furthermore, their analytical potential with respect to quantification and structure elucidation for different applications in the field of bioanalysis is discussed.

Selenomethionine Biotransformation and Incorporation into Proteins along a Simulated Terrestrial Food Chain

Selenium is an essential trace element in vertebrates, but there is a narrow concentration range between dietary requirement and toxicity threshold. Although a great deal is known about the biochemistry of Se from a nutritional perspective, considerably less attention has been focused on the specific biochemistry of Se as an environmental toxicant. Recent advances in hyphenated analytical techniques have provided the capability of quantifying specific chemical forms of Se in biological tissues as well as the distribution of Se among macromolecules. We applied liquid chromatography coupled to inductively coupled plasma mass spectrometryto investigate biotransformations of selenomethionine along a simulated terrestrial food chain consisting of selenomethionine exposed crickets (Acheta domesticus) fed to western fence lizards (Sceloporus occidentalis). Evidence was obtained for selenomethionine biotransformation as well as for sex-specific differences in the metabolism of Se compounds and their subsequent incorporation into proteins in the lizard. The results demonstrate the complexities involved in trophic transfer of Se due to the potential for extensive biotransformation and the species- and even sex-specific nature of these biotransformations.

Isoflavonoids of the Leguminosae

Isoflavonoids are found predominantly in subfamily Papilionoideae of the Leguminosae. This review describes more than 420 new examples of Leguminosae isoflavonoids, giving details of their source, identification, biological activity, synthesis, and ecological or chemosystematic significance. Other topics addressed include the application of hyphenated analytical techniques to the characterisation of legume-derived isoflavonoids, and advances made in biosynthetic studies. A checklist of new compounds by species is given, and 404 references are cited.