Liquid Chromatography Research Articles

Bioactivity-Based Analysis and Chemical Characterization of Hypoglycemic Components from Helicteres angustifolia L.

Helicteres angustifolia L. (H. angustifolia), a well-known traditional Chinese medicine, has been demonstrated to have hypoglycemic activity. We found that the EtOAc extract of H. angustifolia (HAEF) showed stronger α-glucosidase inhibitory activity than that of positive control. Furthermore, the hypoglycemic activity of HAEF was evaluated in streptozotocin (STZ)-induced type 2 diabetes mellitus (T2DM) rats. The results demonstrated that HAEF reduced the drinking quantity, feeding quantity, and controlled weight loss in diabetic rats. Besides, the fasting blood glucose (FBG), viscera index, and the area under time-blood glucose curve (AUC) were significantly decreased, and the oral glucose tolerance was also improved after 5 weeks. Then, the high-performance liquid chromatography with quadrupole time of flight tandem mass spectrometry (HPLC-Q-TOF-MS/MS) method was performed for qualitative analysis of the chemical constituents in HAEF. Twenty-one compounds were identified from in HAEF. Four compounds were further isolated from HAEF and subjected to α-glucosidase inhibition experiments. At the end, molecular docking was empolyed simulate the interaction of three compounds with α-glucosidase. This is the first report on major hypoglycaemic components has been identified in the roots of H. angustifolia. These findings provide a material basis for the use of H. angustifolia in the treatment of diabetes.

Identification of Estrogen-Responsive Proteins in Mouse Seminal Vesicles Through Mass Spectrometry-Based Proteomics

Background: Although estrogenic compounds promise therapeutic potential in treating various conditions, concerns regarding their endocrine-disrupting effects have been raised. Current methodologies for screening estrogenicity in rodent models are limited to the female-specific uterotrophic bioassay. Studies have reported enlargement of the seminal vesicles in orchiectomized males treated with estrogens. However, identifying estrogenicity strictly through changes in wet weights is uninformative regarding the molecular mechanisms of these agents. Therefore, protein-based biomarkers can complement and improve the sensitivity of weight-based assessments. To this end, we present a discovery-driven proteomic analysis of 17β-estradiol’s effects on the seminal vesicles. Methods: We treated orchidectomized mice with the hormone for five days and used the vehicle-treated group as a control. Seminal vesicles were analyzed by shotgun approach using data-dependent nanoflow liquid chromatography–tandem mass spectrometry and label-free quantification. Proteins found to be differentially expressed between the two groups were processed through a bioinformatics pipeline focusing on pathway analyses and assembly of protein interaction networks. Results: Out of 668 identified proteins that passed rigorous validation criteria, 133 were regulated significantly by 17β-estradiol. Ingenuity Pathway Analysis® linked them to several hormone-affected pathways, including those associated with immune function such as neutrophil degranulation. The altered protein interaction networks were also related to functions including endocrine disruption, abnormal metabolism, and therapeutic effects. Conclusions: We identified several potential biomarkers for estrogenicity in mouse seminal vesicles, many of them not previously linked with exogenous 17β-estradiol exposure.

Application of QuEChERS extraction and LC-MS/MS for determination of pharmaceuticals in sewage sludges sampled across the Czech Republic.

The use of pharmaceuticals entails a significant risk of environmental contamination. Wastewater treatment plants (WWTPs) are considered to be the main contributors to contamination as they ineffectively eliminate these compounds from wastewater. Simultaneously, they produce solid waste, sludge, which often contains a variety of retained pollutants, including pharmaceuticals. Since sewage sludge is frequently applied to agricultural soil due to its rich nutrient content, pollutants are introduced into the environment in this way. Only a few studies have been carried out on the topic of the analysis of pharmaceuticals in sludge. Therefore, information on the occurrence of pharmaceuticals in sludge is limited. The present study employed quick, easy, cheap, effective, rugged and safe (QuEChERS) extraction and liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) analysis to establish a simple and reliable procedure for determining 16 pharmaceuticals (antibiotics, anticonvulsants, antidepressants and β-blockers) in sewage sludge. The method has been thoroughly validated, and parameters such as linear range, accuracy, precision, matrix effects and detection and quantification limits were assessed. Our method achieved low limits of quantification (0.5-9.0µgkg-1) and satisfactory recoveries (51-101%). Forty sludge samples from different WWTPs across the Czech Republic were analysed. Fourteen compounds were detected and quantified in most samples, with antidepressants having the highest detection frequency and overall content. Sertraline, with a mean concentration of 521.0µgkg-1, was notably prevalent alongside its metabolite norsertraline (mean concentration 204.9µgkg-1). The antibiotic azithromycin was also found at higher levels (mean concentration 185.1µgkg-1).

Glucose influence cold tolerance in the fall armyworm, Spodoptera frugiperda via trehalase gene expression

The fall armyworm (FAW), Spodoptera frugiperda is a cold-sensitive species that overwinters in temperate climates without diapause. Overwintering in insects involves rapid cold hardening (RCH), supported by trehalose (TRE), which serves as an intermediate between glycogen (GLY) and glucose (GLU). However, both GLU and TRE help maintain homeostasis under stress. TRE is hydrolyzed by the enzyme trehalase (Treh) into GLU. This study retrieved Sf-Treh1a, Sf-Treh1b, and Sf-Treh2 from the FAW transcriptome analysis. RNA interference (RNAi) targeting these three Treh genes resulted in significant downregulation of mRNA levels and altered survival rates in RNAi-treated FAW larvae following RCH treatment. High-pressure liquid chromatography (HPLC) quantification of TRE and GLU in treated groups suggests that GLU is an essential component of the hemolymph for survival adaptation to cold conditions in S. frugiperda. This study reveals limited cold adaptability of FAW, as evidenced by lower glucose concentration levels. We found that FAW requires alternative molecules, in conjunction with glucose and trehalose for freeze tolerance and survivability. Our study aims to discover the molecular mechanisms that contribute to freeze tolerance in FAW by exploring the roles of trehalose, glucose, and glycogen.

Potential Production of Mycotoxins by the Fungal Flora Isolated from "Garba" (Côte D'ivoire)

Garba, an Ivorian dish composed of attiéké (a fermented cassava couscous), fried tuna, and condiments, is increasingly consumed by the population, particularly in the streets of Abidjan. However, garba could pose a health risk to consumers. This study, aimed at preserving public health, sought to identify the toxigenic fungal flora present in garba sold on the streets of Abidjan. To this end, 300 garba samples were collected from four districts of Abidjan. Fungal strains contaminating the garba were enumerated and identified using classical mycological methods. Mycotoxins (aflatoxins and ochratoxins) were quantified using High-Performance Liquid Chromatography (HPLC). Results showed that the average mold counts ranged from 0 to 1.8 x 10^3 CFU/g. The isolated mold strains belonged to ten species grouped into four genera: Aspergillus, Mucor, Penicillium and Paecilomyces, with Mucor spp. (36.89%) and Aspergillus niger (30.09%) being the most predominant. The mycotoxins detected in garba were aflatoxins (B1, B2, G1, and G2) and ochratoxin A, with average levels ranging from 0.42 to 8.07 µg/kg. Approximately 37% and 23% of garba samples had total aflatoxin and aflatoxin B1 levels exceeding regulatory limits. The presence of potentially mycotoxigenic fungal strains in garba could pose a health risk to consumers. Compliance with good hygiene practices during the production and sale of garba will therefore be necessary to considerably reduce contamination and protect consumer health.

Tea Quality: An Overview of the Analytical Methods and Sensory Analyses Used in the Most Recent Studies

Tea, one of the world’s most consumed beverages, has a rich variety of sensory qualities such as appearance, aroma, mouthfeel and flavor. This review paper summarizes the chemical and volatile compositions and sensory qualities of different tea infusions including black, green, oolong, dark, yellow, and white teas based on published data over the past 4 years (between 2021 and 2024), largely focusing on the methodologies. This review highlights the relationships among the different processing methods of tea and their resulting chemical and sensory profiles. Environmental and handling factors during processing, such as fermentation, roasting, and drying are known to play pivotal roles in shaping the unique flavors and aromas of different types of tea, each containing a wide variety of compounds enhancing specific sensory characteristics like umami, astringency, sweetness, and fruity or floral notes, which may correlate with certain groups of chemical compositions. The integration of advanced analytical methods, such as high-performance liquid chromatography (HPLC) and gas chromatography–mass spectrometry (GC–MS), with traditional sensory analysis techniques was found to be essential in the evaluation of the chemical composition and sensory attributes of teas. Additionally, emerging approaches like near-infrared spectroscopy (NIRS) and electronic sensory methods show potential in modern tea evaluation. The complexity of tea sensory characteristics necessitates the development of combined approaches using both analytical methods and human sensory analysis for a comprehensive and better understanding of tea quality.

Zic-HILIC MS/MS Method for NADomics Provides Novel Insights into Redox Homeostasis in Escherichia coli BL21 Under Microaerobic and Anaerobic Conditions

Background: Nicotinamide adenine dinucleotide (NAD+), its precursors, and its derivatives (collectively NADome) play a crucial role in cellular processes and maintain redox homeostasis. Understanding the dynamics of these metabolic pools and redox reactions can provide valuable insights into metabolic functions, especially cellular regulation and stress response mechanisms. The accurate quantification of these metabolites is challenging due to the interconversion between the redox forms. Methods: Our laboratory previously developed a zwitterionic hydrophilic interaction liquid chromatography (zic-HILIC)–tandem mass spectrometry method for the quantification of five essential pyridine nucleotides, including NAD+ derivatives and it’s reduced forms, with 13C isotope dilution and matrix-matched calibration. In this study, we have improved the performance of the chromatographic method and expanded its scope to twelve analytes for a comprehensive view of NAD+ biosynthesis and utilization. The analytical method was validated and applied to investigate Escherichia coli BL21 under varying oxygen supplies including aerobic, microaerobic, and anaerobic conditions. Conclusions: The intracellular absolute metabolite concentrations ranged over four orders of magnitude with NAD+ as the highest abundant, while its precursors were much less abundant. The composition of the NADome at oxygen-limited conditions aligned more with that in the anaerobic conditions rather than in the aerobic phase. Overall, the NADome was quite homeostatic and E. coli rapidly, but in a minor way, adapted the metabolic activity to the challenging shift in the growth conditions and achieved redox balance. Our findings demonstrate that the zic-HILIC-MS/MS method is sensitive, accurate, robust, and high-throughput, providing valuable insights into NAD+ metabolism and the potential significance of these metabolites in various biological contexts.

A Comprehensive Quality Evaluation for Gentiana Rigescens Franch. by Fingerprinting Combined with Chemometrics and Network Pharmacology.

Gentiana rigescens Franch. (G. rigescens) is a unique traditional medicinal herb from southwestern China, and its clinical mechanism for the treatment of hepatitis and the quality differences between different origins are not clear. The research aims to analyze the mechanisms for the treatment of hepatitis and differences in inter-origin differences using analytical techniques, chemometrics, and network pharmacology. Through infrared spectroscopy, spectral images, and high-performance liquid chromatography (HPLC) analysis, it was found that there were differences in absorbance intensity and significant differences in compound content among the samples' origin. G. rigescens iridoids and flavonoids exert therapeutic effects on hepatitis through multiple targets (GAPDH, EGFR, and MMP9, etc.) and multiple pathways (non-small cell lung cancer, hepatitis C, etc.). The above HPLC, chemometrics, and network pharmacology results revealed that gentiopicroside, and swertiamarine was the best quality marker among origins. The accuracy of the ResNet model train, test, and external validation sets for synchronous spectral images were 100 %, which could be utilized as an effective tool for tracing G. rigescens's origins. The R2 of the calibration and validation sets of the PLSR model was higher than 0.70. This model had excellent predictive performance in determining the content of gentiopicroside and swertiamarine, and could quickly, accurately, and effectively predict these two compounds. The research investigates the differences in G. rigescens origins from multiple perspectives, establishes image recognition models and prediction models, and provides new methods and theoretical basis for quality control of G. rigescens.

Lactobacillus johnsonii CCFM1376 improves hypercholesterolemia in mice by regulating the composition of bile acids

Aim: Strains with high bile salt hydrolase (BSH) activity have the potential to regulate cholesterol metabolism. This study aimed to assess the alleviating effect of Lactobacillus johnsonii (L. johnsonii ) CCFM1376, a strain with high BSH activity, on mice with hypercholesterolemia and explore the mechanism of its effect through the modulation of bile acid metabolism. Methods: The BSH activity was measured using the ninhydrin method. C57BL/6J mice were given a high-cholesterol diet to induce hypercholesterolemia with simultaneous gavage of L. johnsonii CCFM1376 for 8 weeks. The biochemical parameters in the serum and liver of hypercholesterolemic mice were measured to assess the alleviating effect of L. johnsonii CCFM1376 on hypercholesterolemia. Bile acid content in the mouse liver, serum, distal ileum contents, and feces was determined using liquid chromatograph mass spectrometer (LC-MS). RNA was extracted from mouse ileum and liver, and the expression levels of relative genes implicated in bile acid metabolism were measured by quantitative real-time PCR (qPCR). Results: Compared to the model group, the group treated with L. johnsonii CCFM1376 exhibited significantly reduced levels of serum total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C), along with a significant increase in high density lipoproteins cholesterol (HDL-C) level. Moreover, hepatic levels of TC and LDL-C in the CCFM1376 group also decreased significantly. Furthermore, the content and amount of unconjugated bile acids in the hepatic-enteric circulation of the L. johnsonii CCFM1376 group significantly increased, and the total bile acid content in the feces also significantly increased. In the L. johnsonii CCFM1376 group, the relative expression levels of ileal farnesoid X receptor (FXR) and fibroblast growth factor 15 (FGF15) were downregulated, while the relative expression level of CYP7A1 was upregulated. Conclusion: These results indicated L. johnsonii CCFM1376 improves hypercholesterolemia in mice by regulating the composition of bile acids. This provides a reference for probiotic strategy to regulate cholesterol metabolism.

The Hydrophobic Amino Acid-Rich Fish Collagen Peptide Ameliorates Dextran Sulfate Sodium-Induced Ulcerative Colitis in Mice via Repairing the Intestinal Barrier, Regulating Intestinal Flora and AA Metabolism.

The incidence of ulcerative colitis (UC) is increasing annually, but treatment option is limited. Fish collagen peptide (FCP) is a food source collagen peptide that has shown promise in alleviating UC symptoms. However, its impact on the intestinal barrier and intestinal metabolic homeostasis in UC remains unclear. This study aimed to analyze the peptide sequences and absolute amino acid (AA) content of FCP, assessing its effects on UC in mice induced by dextran sulfate sodium (DSS). FCP was examined by liquid chromatography and tandem mass spectrometry (LC-MS/MS) analysis. The 3% DSS was utilized to induce UC in murine models, followed by the assessment of the therapeutic efficacy of FCP. Clinical manifestations of UC mice were meticulously evaluated and scored. Subsequently, samples were procured for histological examination and intestinal epithelial barrier integrity analysis as well as macrogenomic and metabolomic profiling. Here, it shows that abundant peptide sequences and AAs were in FCP, particularly enriched in hydrophobic AAs (HAAs). Furthermore, it was observed that FCP effectively reversed colon shortening and reduced the extent of histological damage. Additionally, FCP suppressed the abnormal expression of inflammatory factors and intestinal barrier proteins and modulated the dysbiosis of gut microbiota toward a balanced state. These alterations led to the activation of intestinal alkaline AA and various AA metabolisms, ultimately contributing to the mitigation of UC symptoms. In summary, the diverse peptide sequences and high AAs in FCP, particularly rich in HAAs, can alleviate DSS-induced UC via preserving intestinal barrier integrity, regulating gut microbiota, and modulating AA metabolism.

Herbo-vitamin medicine Livogrit Vital ameliorates isoniazid induced liver injury (IILI) in human liver (HepG2) cells by decreasing isoniazid accumulation and oxidative stress driven hepatotoxicity

BackgroundTuberculosis (TB) is a leading cause of infection related mortality. Isoniazid is one of the frontline drugs for anti-TB therapy. Hepatotoxicity induced by isoniazid is a major cause of drug-discontinuation which may lead to development of resistant TB or increased mortality.PurposeTo characterize pharmacological properties of plant-based prescription medicine, Livogrit Vital (LVV) against isoniazid-induced liver injury (IILI) using HepG2 cells.MethodPhytometabolite characterization of LVV was performed by High-performance liquid chromatography (HPLC). The effects of LVV on cytosafety, IC50 shift, oxidative stress, ER stress, apoptosis, liver injury markers, and accumulation of isoniazid and hydrazine was performed on HepG2 cells induced with isoniazid. Silymarin was used as the positive control.ResultsHPLC based phytometabolite characterization of LVV revealed the presence of several anti-oxidant, anti-apoptotic, and hepatoprotective compounds. In isoniazid-induced HepG2 cells, LVV reduced cytotoxicity of isoniazid and shifted its IC50 value. Treatment with LVV reduced ROS generation and lipid peroxidation; enhanced GSH enzyme levels in isoniazid-induced HepG2 cells. As per the mechanistic evaluation, LVV modulated gene expression level of Caspase-3, FGF21, and IRE-1α. LVV treatment also normalized isoniazid-induced elevated Caspase-3 activity and cPARP1 protein levels, indicating its potentials to regulate liver cell apoptosis. Concomitantly, biomarkers of hepatotoxicity, ALT and GGT, also decreased by LVV treatment. Interestingly, LVV treatment reduced intracellular accumulation of isoniazid and its toxic metabolite hydrazine, in isoniazid-stimulated HepG2 cells.ConclusionTreatment of hepatic cells with the herbo-vitamin medicine, Livogrit Vital, regulates IILI by modulation of oxidative and ER stress, apoptosis, and bioaccumulation of isoniazid and hydrazine. Collectively, Livogrit Vital could well be explored as an adjuvant hepatoprotective agent alongwith anti-TB medicines.

Prevalence and screening of hemoglobinopathies and glucose-6-phosphate dehydrogenase deficiency in Yemeni blood donors

ABSTRACT Hemoglobinopathies and Glucose-6-Phosphate Dehydrogenase (G6PD) deficiency pose significant health threats in Yemen, which can be alleviated through robust preventive strategies, highlighting the importance of implementing a cost-effective and efficient blood donor screening method to facilitate early diagnosis and management of erythrocyte disorders, ultimately improving both donor safety and patient health outcomes. Objectives This study aimed to determine the prevalence of total blood cell abnormalities, hemoglobinopathies and G6PD deficiency and evaluate the efficacy of red blood cell (RBC) indices, mentzer index (MI) and naked-eye single tube red cell osmotic fragility (NESTROF) test as screening tools for diagnosis of β thalassemia trait among Yemeni blood donors. Methods A cross-sectional study was conducted with 106 volunteer blood donors who met the national standard criterion of blood donation. Various tests were performed, including complete blood count (CBC), serum ferritin, sickling test, G6PD assay, NESTROF test and high-performance liquid chromatography (HPLC). Results The prevalence of hematological abnormalities among blood donors reached 68.9%, with functional RBC abnormalities at 51.9%, leukopenia at 10.4%, thrombocytosis at 1.9%, and thrombocytopenia at 4.7%. Additionally, hemoglobinopathies were found in 21.7% of donors, with β-thalassemia trait at 3.8%, sickle cell trait at 1.9%, and suspected α-thalassemia trait at 16%, while G6PD deficiency and iron deficiency were present in 12.3% and 17.9% of donors, respectively. The NESTROF test, MCV and MCH demonstrated a sensitivity rate of 100%. MI and MCH exhibited the highest specificity followed by NESTROF test in the screening of β-thalassemia trait. Conclusions The prevalence of hemoglobinopathies and G6PD deficiency appear to be common among Yemeni blood donors. These results emphasize the necessity of comprehensive blood donation screening programs to safeguard the blood supply and promote early detection and management of hemoglobinopathies and G6PD deficiency in Yemen.

Pharmaceutical Residues in Fishes and Macro Invertebrates around the Wastewater Treatment Plant Site at the University Teaching Hospital of Yaoundé, Cameroon

Introduction: Hospital effluents are one of the main sources of dissemination of pharmaceutical residues in hydro system. Worldwide, analyses of these effluents have revealed the presence of various pharmaceuticals. Very few studies assessing the contamination of solid matrices by pharmaceutical residues have been carried out in Africa, specifically in Cameroon. Objectives: The study aimed to detect pharmaceutical residues in solid environmental matrices at sites around the discharge of the effluent from the Yaoundé University Teaching Hospital. Methods: Four hundred and fifty-nine (459) macro invertebrates and twenty (20) Nile tilapia were collected in three sampling sites before and beyond the wastewater treatment plant of the Yaoundé University Teaching Hospital. The specimens were conserved in a cooler prior to freeze drying and grinding. The powders were sent for analysis at the Sefako Makgatho Health Sciences University in South Africa. The analysis was carried out using an ultra-high-performance liquid chromatography coupled with a triple quadrupole tandem mass spectrometer. Results: A total of ninety-nine pharmaceutical residues related to seventeen pharmacological classes were detected in all samples of macro invertebrates (72) and Nile tilapia (55 for fish viscera and 58 for fish flesh) collected. Antibiotics were the most represented (23.2 %), followed by anti-inflammatory (18.2 %), and anti-parasitics (9.1 %). More than half (53.6 %) of the drug residues detected in the fish samples were of the anti-inflammatory (21,71 %), antihypertensive (17,67 %) and antibiotic (13,13 %) classes. The number of drug residues detected in the macroinvertebrates before the treatment plant (41) was smaller than those detected after the treatment plant (67). Conclusion: Five therapeutic classes (antibiotics, anti-inflammatories, antihypertensive, antiparasitics and hormones) were mostly represented.These results are in line with the importance of considering diet of fishes and/or humans as an exposure pathway based on the great similarity in pharmaceutical composition between macroinvertebrates and Nile tilapia.

A Fully Validated LC-MS Quantitation Method for Psychoactive Compounds Found in Native South American Plant Species

Psychoactive drugs are compounds that alter the function of the central nervous system, resulting in changes in perception, mood, cognition, and behavior. A subclass of psychoactive drugs, psychedelics, are hallucinogenic drugs that can trigger psychedelic experiences and possible changes in mental perception. The potential use of psychedelics as a therapeutic has led to an increase in clinical research focusing on the treatment of mental disorders including anxiety and depression. There are numerous species belonging to Psychotria and Banisteriopsis which have been reported to contain psychedelic and psychoactive compounds; however, there is a lack of validated analytical methods for raw plant material, which is crucial if these plants are to be commercially cultivated for medicines. This study provides a fully validated method using ultra-high performance liquid chromatography (UHPLC) coupled to mass spectrometry (MS) for the following six compounds: tryptamine, N,N-dimethyltryptamine (DMT), 5-methoxy-N,N-dimethyltryptamine (5-MeO-DMT), tetrahydroharmine (THH), harmaline, and harmine. The validated method was used to determine the psychoactive concentrations in Psychotria viridis, Psychotria carthagenensis, Banisteriopsis caapi, and Alicia anisopetala. Validation parameters were established; linearity (R2 = 0.988–0.999), limit of detection (LOD) (0.06–0.11 ng/mL), limit of quantitation (LOQ) (0.18–0.34 ng/mL), accuracy, precision, extraction efficiency (>98%), recovery (74.1–111.6%), and matrix effect (70.6–109%) were all evaluated. All six compounds eluted within nine minutes, with a total analysis time of 20 min including column equilibration. This method establishes a high-throughput method for the robust analysis of psychedelics which may see future use in agricultural research and industry.

The Impact of Feed Management Technologies on Mineral Oil Hydrocarbons (MOH) Contamination: A Comparative Farm Level Approach

Legislative frameworks about contamination are often limited to foods and underestimate the role of animal nutrition for safe production. This study aims to assess mineral oil hydrocarbon (MOH) contamination in feed and identify the technological factors that are contributing to this issue, particularly focused on mechanised harvesting and processing. Three dairy farms, classified by contamination risk (low, medium, and high), were selected, and fifteen feed samples were analysed using the coupled liquid chromatography–gas chromatography–flame ionisation detection (LC–GC–FID) method, with a microwave-assisted saponification (MAS) step to determine mineral oil saturated (MOSH) and aromatic (MOAH) hydrocarbon levels. Important contamination levels were observed depending on the technological development of each farm. MOSH levels ranged from 11.4 mg/kg to 81.40 mg/kg, while MOAH levels ranged from 0.5 mg/kg to 4.6 mg/kg. MOAH accounted for 4.74% of the total MOH content. The results showed a connection between feed production technologies and MOH contamination levels. Factors such as the mechanisation, the machinery used, and the storage conditions were potentially contributors to contamination, while chemical treatments had no direct impact but some potential risks. The contamination levels varied across farms, indicating certain contamination sources beyond technological factors. Advanced technological measures and proper equipment maintenance are suggested to mitigate MOH contamination risks in feed.

Rapid Raman spectroscopy analysis assisted with machine learning: a case study on Radix Bupleuri.

Radix Bupleuri has been widely used for its plentiful pharmacological effects. But it is hard to evaluate their safety and efficacy because the concentrations of components are tightly affected by the surrounding environment. Thus, Radix Bupleuri samples from different regions and varieties were collected. Based on the experimental and computational Raman spectrum, machine learning is emphasized for certain obscured characteristics; for example, linear discriminant analysis (LDA), support vector machine (SVM), eXtreme gradient boosting (XGBoost) and light gradient boosting machine (LightGBM). After dimension reduction by LDA, models of SVM, XGBoost and LightGBM were trained for classification and regression prediction of Bupleurum production regions. Support vector classifiers achieved the best accuracy of 98% and an F1 score above 0.96 on the test set. Support vector regression has a good fitting performance with an R2 score above 0.90 and a relatively low mean square error. However, complex models were prone to overfitting, resulting in poor generalization ability. Among these machine learning models, the typical LDA-SVM models, consistent with the high-performance liquid chromatography results, demonstrate great performance and stability. We envision that this rapid classification and regression technique can be extended to predictions for other herbs. © 2024 Society of Chemical Industry.

Ultra-High-Performance Liquid Chromatography–Tandem Mass Spectrometry Analysis of Δ9-Tetrahydrocannabinol and Cannabidiol in Commercial Suk-Saiyasna Herbal Remedy: Applying Hansen Solubility Parameters for Sample Extraction to Ensure Regulatory Compliance

Background: Suk-Saiyasna is a traditional Thai herbal remedy that comprises 12 distinct herbs. Among these, cannabis leaves constitute 12 of the total 78 components in this formulation. This study specifically examines the portion of the cannabis plant, which accounts for approximately 15.8% of the overall herbal composition used in the entire remedy. According to the Thailand Narcotics Act of 2022, the Δ9-tetrahydrocannabinol (Δ9-THC) concentration in herbal extracts must not exceed 0.2% by weight. This study aims to quantify the levels of Δ9-THC and cannabidiol (CBD) in commercial Suk-Saiyasna products. Methodology: This research utilizes Hansen Solubility Parameters (HSPs) to identify the optimal solvent for ultrasonic extraction, thereby maximizing cannabinoid yield. An advanced method was developed employing ultra-high-performance liquid chromatography–tandem mass spectrometry (UHPLC-MS/MS), compliant with AOAC standards to meet regulatory guidelines. The method validation emphasized specificity, linearity, sensitivity, accuracy, and precision. Results: Dichloromethane was chosen due to its favorable HSP values, enabling highly efficient extraction of Δ9-THC and CBD, achieving recovery rates of over 99.9% after the second extraction. This investigation benefits from the accuracy of the UHPLC-MS/MS technique in quantifying cannabinoids in commercial products, with Δ9-THC concentrations observed between 0.00231% and 0.14218%, and CBD concentrations ranging from 0.00002% to 0.01541%, all remaining below the legal limit. Conclusions: The variability in cannabinoid concentrations among various commercial products highlights the need for standardization in the herbal industry. This finding underscores the critical role of rigorous quality control measures in ensuring the safety and efficacy of cannabis-derived products.

Therapeutic Peptides Encapsulated into Lipid-Based Nanovesicles: Scaling from Lab to Plant

Lipid-based peptide-encapsulated nanovesicles represent a cutting-edge approach in drug delivery systems, offering enhanced stability, targeted delivery, and controlled release of therapeutic peptides. This review explores the comprehensive science and techniques behind the robust formulation and analytical methods for these nanovesicles, spanning from lab-scale research to large- scale manufacturing. Critical aspects include lipid selection, detailing how the choice of lipids impacts the stability and efficacy of the nanovesicles, and various peptide encapsulation techniques, emphasizing methods to achieve high encapsulation efficiency and controlled release. Key processes in vesicle formation, such as thin film hydration, sonication, and microfluidics, are discussed to highlight their roles in producing nanovesicles with consistent size and morphology. The importance of stability studies, including physical, chemical, and thermal assessments, is underscored. Analytical techniques such as Dynamic Light Scattering (DLS), Zeta Potential measurement, High-Performance Liquid Chromatography (HPLC), and Mass Spectrometry (MS) are detailed, showcasing their crucial roles in characterizing nanovesicles and ensuring their quality, safety, and efficacy for pharmaceutical use. The review addresses the challenges and solutions associated with scaling up production, including maintaining consistent product quality, optimizing manufacturing processes, and controlling production costs. It also navigates the regulatory landscape, discussing the stringent guidelines set by agencies such as the FDA and EMA and the necessity of strategic regulatory planning to facilitate market approval. This comprehensive review aims to serve as an in-depth resource for experts in the field, aiding in the translation of lipid-based peptide-encapsulated nanovesicle formulations from bench to bedside, ultimately enhancing therapeutic outcomes and expanding treatment options for various disease. Key Words: Peptide, Amino acid, Degradation, Pharmacokinetics, Peptide encapsulation, drug delivery systems, Peptide Characterization

Synthesis, Chiroptical Properties, and Absolute Configuration Determination of Phenyl‐4‐pyridyl‐2,5‐dipyrimidinylmethane

Phenyl‐4‐pyridyl‐2,5‐dipyrimidinylmethane (1) has been synthesized in five steps from 5‐methylpyrimidine. The structural feature of 1 is the symmetry elements with respect to the 180º rotation of the aryl group along the central C‐C bonds. Due to this structural feature, the compound 1 is expected the good crystallinity despite of the asymmetric molecular structure. The optical resolution of 1 was achieved by high performance liquid chromatography using chiral stationary phase (chiral HPLC). The circular dichroism (CD) spectra of the two fractions showed opposite signs. As expected, the X‐ray crystallographic analysis was successfully performed for both the racemic and enantiomerically pure crystals of compound 1, with the nitrogen atoms being unambiguously assigned in both cases. It was concluded the first‐eluted fraction in chiral HPLC was determined to be S configuration both by the CD calculation for reproducing the experimental CD spectrum and by the anomalous diffraction of X‐ray crystallographic analysis of enantiomerically pure 1.

Application of Quantitative Proton Nuclear Magnetic Resonance Spectroscopy for Determination of the Content of Geniposide in Gardeniae fructus

Background: Gardeniae fructus (Zhi-Zi) is the dry ripe fruit of the plant Gardenia jasminoides Ellis (Rubiaceae), which can be used as both food and medicine. Geniposide, a key constituent of Gardeniae fructus, serves as an indicator component for evaluating the quality of Gardeniae fructus. Traditionally, the quantification of geniposide in Gardeniae fructus is achieved through High-performance Liquid Chromatography (HPLC)-based methods. Objective: The present study aimed to introduce a rapid approach to quantifying geniposide content in Gardeniae fructus along with validating its effectiveness. Methods: The experiments involved finding a suitable deuterium solvent, Internal Standard (IS), specific peak, and Nuclear Magnetic Resonance (NMR) parameters for quantitation, and validating specificity, accuracy, precision, and stability. Results: The results have indicated that methanol-d4 as a solvent has facilitated excellent signal separation in the proton Nuclear Magnetic Resonance (1H NMR) spectroscopy, with trimethyl 1,3,5- benzenetricarboxylate emerging as the ideal IS. The specific signal at δ 7.45, corresponding to H-3 in geniposide, has been identified as the optimal peak for integration. The application of distinctive signals from the 1H NMR spectroscopy has allowed for the precise quantification of geniposide in Gardeniae fructus. Conclusion: This study has suggested using 1H NMR to quantify geniposide in Gardeniae fructus to be a viable alternative to HPLC-based methods, providing a suitable approach for quality control of Gardeniae fructus.