Metabolite Composition Research Articles

Differentiation of Ayahuasca Samples According to Preparation Mode and Botanical Varieties Using Metabolomics

ABSTRACT Ayahuasca is a brew traditionally prepared with a mixture of Psychotria viridis leaves and Banisteriopsis caapi vine and has demonstrated therapeutic properties for depression. Knowledge of the brew composition is important to improve the therapeutic potential and decrease side effects if ayahuasca becomes an option for refractory depression treatment. Ultra-high performance liquid chromatography coupled to high-resolution mass spectrometry (UHPLC-HRMS) was applied to analyze 126 ayahuasca samples collected from different ayahuasqueiro groups and geographic origins. We were able to observe a differentiation in the metabolite composition of ayahuasca samples prepared by diverse ayahuasqueiro groups. These samples presented different antioxidant effects based on FRAP and ORAC assays. Exploratory statistical analysis demonstrated a trend of separating the samples according to the religious group. The most important identified compounds for differentiation of the brew prepared by distinct religious groups are glycosylated and/or phenolic compounds. The comparison based on the mode of ayahuasca preparation presented more variability than the comparison based on the botanical variety of B. caapi used. We conclude that ayahuasca samples prepared with “caupuri” or “tucunacá” separately exhibited differences in the analysis of L-glutamate and the metabolism of arginine and proline. This suggests that a possible variation in this pathway could explain the occurrence of swollen stem nodes in “caupuri,” one of the B. caapi varieties.

HRMAS-NMR-Based Metabolomics Approach to Discover Key Differences in Cow and Goat Milk Yoghurt Metabolomes

This study highlights the differences in the metabolomes of cow milk yoghurt (CY) and goat milk yoghurt (GY) using a nuclear magnetic resonance (NMR)-based metabolomic approach. The 1H HRMAS-NMR spectrum displayed 21 metabolites comprising organic acids, sugars, amino acids, amino acid derivatives and phospholipids. The orthogonal partial least squares discriminant analysis model clearly separated CY and GY groups, implying differences in metabolite composition. The corresponding Variable Importance in Projection (VIP) plot revealed that choline, sn-glycero-3-phosphocholine, O-phosphocholine, fucose, citrate, sucrose, glucose and lactose mainly contributed to the group separation (VIP > 1). Hierarchical cluster analysis further confirmed the metabolome similarities and differences between CY and GY. Additionally, 12 significantly differential metabolites (with a fold change > 1.5 and p-value < 0.05) were identified, with 1 downregulated and 11 upregulated. Pathway impact analysis revealed the correlation of significant metabolites with starch and sucrose metabolism, galactose metabolism, and the citrate cycle. Furthermore, receiver operating characteristic curve analysis identified eight metabolites (choline, sn-glycero-3-phosphocholine, fucose, O-phosphocholine, glucose, citrate, 2-oxoglutarate, lactose and sucrose) as candidate biomarkers. This study represents the first utilization of HRMAS-NMR to analyze the metabolomic profiles of yoghurt made from cow and goat milk. In conclusion, these findings provide preliminary information on how NMR-based metabolomics can discriminate the metabolomes of yoghurt prepared from the milk of two different animals, which may be valuable for authenticity and adulteration assessments.

Oral supplementation of heat-killed Enterococcus faecalis strain EC-12 relieves gastrointestinal discomfort and alters the gut microecology in academically stressed students.

Stress significantly affects gastrointestinal and mental health, and the gut microbiota plays a pivotal role in this process. Enterococcus faecalis strain EC-12 (EC-12) is a lactic acid bacterium that has several health benefits. To investigate the impact of oral supplementation with heat-killed EC-12 on the discomfort caused by stress, a randomised, double-blind, placebo-controlled trial was conducted with students under academic stress taking EC-12 (n= 14) or a placebo (n= 13) daily for one week. Improvement in the students' symptoms was assessed using the visual analogue scale. Faecal microbiota was characterised by next-generation sequencing of 16S rRNA genes, and faecal metabolites and short-chain fatty acids were analysed using a GC-MS metabolomics approach. Significant improvements in abdominal pain and rumbling of the stomach were found in the EC-12 group compared to the placebo group, but no changes were observed in mental symptoms or salivary cortisol levels. The relative abundance of E. faecalis significantly increased in the EC-12 group after the trial; however, the composition and diversity of the gut microbiota did not change significantly. Functional analysis of the gut microbiota suggested that EC-12 intake alters specific metabolic pathways. Although the levels of faecal short-chain fatty acids did not change between the groups before and after the trial, EC-12 intake altered the composition of faecal metabolites, with a significant increase in tryptamine levels. The ratio of students with improved symptoms to those with increased tryptamine levels was calculated based on the number of students with elevated faecal tryptamine levels who showed symptomatic improvements. The ratio of improved rumbling stomach was higher than that of other types of digestive discomfort. These results suggest that oral supplementation with EC-12 has a potentially beneficial effect on stress-induced gastrointestinal discomfort, which may occur through alterations in gut microbiota composition and metabolism. This study was registered at the University Hospital Medical Information Network Center (UMIN) under the UMIN ID: UMIN000048184.

Metabolomic Fingerprints: Seasonal and Farm-Specific Differences in Heterotrigona itama Bee Bread

Stingless bees’ products such as honey and bee bread are beneficial to human health. However, the metabolite composition within the products may differ according to spatial and temporal factors. This study aims to investigate the impact of spatial and temporal factors on the significant metabolite composition present in Heterotrigona itama bee bread collected from different seasons and types of bee farms. Thus, tandem mass spectrometry (LCMS/MS) based metabolomics analysis was used to achieve the said goal, focusing on H. itama bee bread samples, followed by multivariate analysis using the MetaboAnalyst platform. Our findings revealed significant metabolites that set bee bread samples apart. In terms of temporal factors, the analysis highlights specific metabolites exclusive to the wet season, such as flavones and flavonoids. However, in contrast, during the wet season, there are no significant metabolites in herbal farm samples. The results also underscore the significance of phenolic compounds in wet season samples and as vital metabolites distinguishing the groups. This study highlights a total of 24 annotated significant metabolites identified in different bee bread samples, as well as their capacity to differentiate between seasons and bee farms. Notably, these results suggest a wide variety of potential plant families as a source of bee bread. These findings shed light on the impact of seasons and diverse plant families contributing to bee bread composition, which may impact the growing meliponiculture industry greatly.

Widely Targeted Metabolomics Analysis Revealed the Component Differences of Hemerocallis citrina Borani in Different Production Areas of Datong

Hemerocallis citrina Borani (H. citrina) has garnered significant attention due to its abundant nutritional quality. Datong, located in Shanxi Province, is recognized as one of the four major production regions for high-quality H. citrina. While Datong boasts multiple production areas, the nutritional composition of daylilies varies across regions due to environmental factors and planting patterns, which remain unclear. This study focuses on the total polyphenol and flavonoid contents (TPCs and TFCs) and protein content of H. citrina from three areas in Datong: Sanshilipu (DTSSLP), Dangliuzhuang (DTDLZ), and Jijiazhuang (DTJJZ). Additionally, a widely targeted metabolomics analysis was used to analyze the metabolite composition of H. citrina from these three areas. The results showed that H. citrina in DTSSLP had the highest contents of protein and amino acids, as well as TPCs and TFCs. A total of 798 differential metabolites were identified in H. citrina across the areas, with DTSSLP showing the highest levels of different classifications of metabolites, indicating its enhanced health benefits and physiological activities. Nine metabolic pathways were related with the different characteristics among DTSSLP, DTDLZ, and DTJJZ. This study provides theoretical support for distinguish H. citrina from different producing regions and elucidates the mechanisms underlying its metabolic pathways.

The Impact of Fermented Milk Products Containing Bifidobacterium longum BB536 on the Gut Environment: A Randomized Double-Blind Placebo-Controlled Trial

Probiotics, particularly those native to the gut microbiota, have a profound influence on the gut environment. In this study, we conducted a randomized placebo-controlled, double-blind, parallel-group comparison trial to investigate the effects of Bifidobacterium longum BB536 (B. longum BB536) on the fecal microbiota and metabolite compositions in healthy individuals. We compared the effects of fermented milk produced solely with Streptococcus thermophiles and Lactobacillus bulgaricus (placebo group) and fermented milk supplemented with B. longum BB536 (BY group). Our findings revealed a significantly greater relative abundance of Faecalibacterium in the BY group than in the placebo group by the 3rd day, a trend that persisted until the end of the trial on the 17th day. Additionally, the BY group presented significantly increased concentrations of tryptophan (Trp), Indole-3-lactic acid, and Indole-3-aldehyde on the 17th day. A significant positive correlation was observed between the relative abundance of Faecalibacterium and the number of viable B. longum BB536 bacteria in the feces. The concentrations of Trp and Indole-3-acetic acid were also significantly correlated with the number of viable B. longum BB536 bacteria in the feces. Our results suggest that B. longum BB536 intake can modulate the gut microbiota and metabolite profiles, which are general indicators for monitoring the gut environment, potentially conferring health benefits to the host.

Unveiling the Metabolic Trajectory of Pig Feces Across Different Ages and Senescence.

Porcine models are increasingly recognized for their similarities to humans and have been utilized in disease modeling and organ grafting research. While extensive metabolomics studies have been conducted in swine, primarily focusing on conventional cohorts or specific animal models, the composition and functions of fecal metabolites in pigs across different age groups-particularly in the elderly-remain inadequately understood. In this study, an untargeted metabolomics approach was employed to analyze the fecal metabolomes of pigs at three distinct age stages: young (one year), middle-aged (four years), and elderly (eight years). The objective was to elucidate age-associated changes in metabolite composition and functionality under standardized rearing conditions. The untargeted metabolomic analysis revealed a diverse array of age-related metabolites. Notably, L-methionine sulfoxide levels were found to increase with age, whereas cytidine-5-monophosphate levels exhibited a gradual decline throughout the aging process. These metabolites demonstrated alterations across various biological pathways, including energy metabolism, pyrimidine metabolism, lipid metabolism, and amino acid metabolism. Collectively, the identified key metabolites, such as L-methionine sulfoxide and Cholecalciferol, may serve as potential biomarkers of senescence, providing valuable insights into the mechanistic understanding of aging in pigs.

Enhanced Production of Microalgal Metabolites Through Aeration Coupled with Stirring

Adequate mixing is a key factor for microalgal cultivation to achieve high biomass production, so it is essential to clarify the comparative effects of different mixing methods on microalgal productivity, which has rarely been studied previously. This work therefore aimed to investigate the effects of different mixing methods (stirring, aeration, and aeration coupled with stirring) on the growth and metabolite composition of Chlorella sorokiniana SDEC-18, a strain with potential for large-scale application. The results showed that mixing was beneficial for carbohydrate accumulation, while dual mixing (aeration coupled with stirring) promoted growth and achieved the highest dry mass and metabolite productivities (including carbohydrates, proteins, and lipids) through enhancement of light energy capture in the entire system. The stirring speed in the dual mixing approach of aeration coupled with stirring was also considered: the optimal condition was found to be 800 rpm. The maximum biomass was 3.56 g L−1, and the carbohydrate productivity was as high as 119.45 mg L−1 d−1, which was the highest metabolite productivity (higher than proteins or lipids), obtained from aeration coupled with stirring at 800 rpm. Our study suggests that aeration coupled with stirring provides a feasible strategy for microalgal production, due to the optimal availability of CO2 and light achieved through effective mixing.

Secondary Metabolites Screening Using GC-MS and In-Vitro Biological Assessment Against Human Pathogenic Bacterial Strains of Viola Indica

Viola indica, a medicinal herb with a long history of traditional use in treating various diseases. In this study we analyzed methanolic extract of Viola indica for metabolite composition using GC-MS, and antibacterial activity through disc diffusion method. In GC-MS analysis, we found total of 25 metabolites in which the major metabolites were 9,12-Octadecadienoic acid (Z,Z)- (20%; 20.34 min), followed by Stigmasterol, 22,23-dihydro- (16%; 26 min) and α-Amyrin (12.38%; 26.65 min) . In antibacterial activity viola indica methanolic extract showed significant antibacterial effects against all bacterial strains. The highest antibacterial activity was recorded against Bacillus subtilis (8.50±0.20 mm), followed by Klebsiella pneumoniae (8.10±0.25 mm) zone of inhibition, while the lowest was recorded against Staphylococcus aureus (2.10±0.30 mm). The result of this study suggests that viola indica has potent antibacterial metabolites with good potential as a natural alternative to synthetic antibacterial agents. However, this study also suggest that more instrumental and biological research is required to explore it’s in-depth phytochemistry, pharmacological potential and toxicological properties.

Multi-omics analysis of soil microbiota and metabolites in dryland wheat fields under different tillage methods

No-tillage and subsoiling can improve soil aggregate structure and realize a synergistic effect of soil carbon and nitrogen retention compared with deep tillage. This study aimed to investigate the effects of different tillage methods on the microbiome and metabolites in wheat rhizosphere. Results indicated that no significant differences in the diversity of soil bacterial and fungal communities were observed among the tillage methods. Analysis revealed that no-tillage enriched specific genera such as Cryptosporangium, Crossiella, Rhodothermaceae, Leptothrix, Stilbella, Diutina, and Pyrenochaetopsis, while subsoiling was associated with Rubrobacter, Latescibacteraceae, Nitrospira, Rokubacteriales, and Ctenomyces. Deep tillage, on the other hand, showed significant associations with Nocardia, Aeromicrobium, Sphingopyxis, Cordyceps, and Subulicystidium. Metabolomic analysis identified differential metabolites involved in various pathways, including the biosynthesis of plant secondary metabolites, ABC transporters, and starch and sucrose metabolism. Correlation analysis revealed a significant interaction between microorganisms and metabolites in wheat rhizosphere. Bacteria at the genus level exhibited greater associations with differential metabolites. In conclusion, different tillage practices can alter the composition of microbial communities and metabolites in wheat rhizosphere, and their interactions may affect soil fertility and wheat growth.

Endophytic Fungus UJ3-2 from Urtica fissa: Antibacterial Activity and Mechanism of Action against Staphylococcus aureus

Taking the endophytic fungus UJ3-2, isolated from Urtica fissa, as the experimental material, this study aimed to explore the composition of its metabolites and the underlying mechanisms by which it inhibits Staphylococcus aureus. Initially, the MIC, MBC, inhibitory curves, biofilm growth, and extracellular nucleic acids and proteins of S. aureus in response to the metabolites were measured. Secondly, PI staining and SEM were used to evaluate the impact of the metabolites on the integrity of the cell wall and overall morphology of S. aureus. Additionally, UPLC-MS was employed to analyze the composition of the secondary metabolites. The UJ3-2 strain was identified as Xylaria grammica based on ITS sequencing and designated as Xylaria grammica UJ3-2. Our results revealed that the metabolites of UJ3-2 exhibited excellent in vitro antibacterial activity against S. aureus, with both MIC and MBC values of 3.125 mg/mL. The inhibitory curve confirmed that 1 MIC of UJ3-2 metabolites could completely inhibit the growth of S. aureus within 24 h. With increasing concentrations of UJ3-2 metabolites, the growth of S. aureus biofilms was significantly suppressed, and obvious leakage of nucleic acids and proteins was observed. PI fluorescence staining indicated that various concentrations of UJ3-2 metabolites disrupted the integrity of the S. aureus cell membrane. SEM observation revealed that the treated S. aureus surfaces became rough, and the bacteria shrank and adhered to each other, showing a dose-dependent effect. UPLC-MS analysis suggested that the main components of the fermented metabolites were 6-oxocineole (17.92%), (S)-2-acetolactate (9.91%), 3-methyl-cis,cis-muconate (4.36%), and 8-oxogeranial (3.17%). This study demonstrates that the endophytic fungus UJ3-2 exhibits remarkable in vitro antibacterial effects against S. aureus, primarily by enhancing the permeability of the S. aureus cell membrane, causing the leakage of its intracellular contents, and altering the bacterial surface morphology to inhibit the pathogen. The endophytic fungus UJ3-2 has a good antibacterial effect on S. aureus, which gives it certain application prospects in the screening and industrial production of new and efficient natural antibacterial active substances.

Electron-Transferring Flavoprotein and Its Dehydrogenase Required for Fungal Pathogenicity in Arthrobotrys oligospora.

Electron transfer flavoprotein (ETF) plays an important function in fatty acid beta oxidation and the amino acid metabolic pathway. It can provide pathogenicity to some opportunistic fungi via modulating cellular metabolite composition. Arthrobotrys oligospora is a typical invasion fungus to nematodes. Its ETF characterization is still unknown. Here, we showed that the mutations of A. oligospora ETF (Aoetfα and Aoetfβ) and its dehydrogenase (Aoetfdh) led to severe defects in mitochondrial integrity and blocked fatty acid metabolism. The pathogenicity-associated trap structures were completely suppressed when exposed to nematode-derived ascarosides and nutrition signals, including ammonia and urea. Compared to the wild-type strain, the nematode predatory activity was significantly reduced and delayed. But surprisingly, the rich nutrition could restore the massive trap and robust predatory activity in the mutant Aoetfβ beyond all induction cues. Moreover, the deletion of Aoetfβ has led to the accumulation of butyrate-like smell, which has a strong attraction to Caenorhabditis elegans nematodes. Ultimately, ETF and its dehydrogenase play a crucial role in nematode-trapping fungi, highlighting mitochondrial metabolite fluctuations that are connected to pathogenesis and further regulating the interactions between fungi and nematodes.

Lactiplantibacillus plantarum P101 Alleviated Alcohol-Induced Hepatic Lipid Accumulation in Mice via AMPK Signaling Pathway: Gut Microbiota and Metabolomics Analysis.

Mitigating steatosis is essential for delaying the progression of alcoholic liver disease. The effect and mechanism of Lactiplantibacillus plantarum P101 (LP.P101) on alleviating alcohol-induced hepatic lipid accumulation were investigated in our study. The mouse model was constructed by a short-term (10-day)-plus-binge ethanol feeding and gavaged with 108 CFU/mL of LP.P101 daily. Lipid droplet in the liver was significantly reduced by LP.101 intervention on AMPK activation. However, when AMPK was inhibited by dorsomorphin, the levels of related indicators (ALT, TG, etc.) and the expression levels of AMPK and relevant genes in the liver converged to that of the alcohol-fed group. Compared with the alcohol-fed group, LP.P101 reduced the relative abundance of Firmicutes and increased that of Bacteroidetes. Parabacteroides merdae was negatively correlated with lipid accumulation, and unclassified Negativibacillus was negatively associated with AMPK activation. Importantly, LP.P101 modified the compositions of the serum metabolites. The potential biomarker stercobilinogen was positively correlated with AMPK activation and negatively associated with lipid accumulation. This work confirmed that LP.P101 attenuated alcohol-induced hepatic lipid accumulation in mice through AMPK activation, and the alterations in gut microbiota and metabolites may play a significant role on AMPK activation.

Goat Milk Exosomes Ameliorate Ulcerative Colitis in Mice through Modulation of the Intestinal Barrier, Gut Microbiota, and Metabolites.

Goat milk is rich in a variety of nutrients that are important for intestinal health and disease prevention. However, the role of exosomes in goat milk remains to be elucidated. This study investigated for the first time the therapeutic efficacy and molecular underlying mechanisms of mature milk exosomes (M-exo) and goat colostrum exosomes (C-exo) on dextran sodium sulfate-induced ulcerative colitis (UC) in mice. The findings demonstrate that M-exo and C-exo significantly improved physiological indices, suppressed the secretion of proinflammatory cytokines, and diminished oxidative stress and apoptosis in UC mice. Moreover, C-exo and M-exo restored the intestinal barrier function, remodeled the gut microbiota, and improved metabolite composition in the feces of colitis mice. In conclusion, goat milk exosomes ameliorate UC in mice, which provides a basis for the development of functional food applications for the prevention and treatment of inflammatory bowel disease.

Effect of host breeds on gut microbiome and fecal metabolome in commercial pigs

BackgroundGut microbial composition and its metabolites are crucial for livestock production performance. Metabolite profiles from autopsied biospecimens provide vital information on the basic mechanisms that affect the overall health and production traits in livestock animals. However, the role of the host breed in the gut microbiome and fecal metabolome of commercial pigs remains unclear. In this work, differences in microbiota composition among three commercial pig breeds Duroc, Yorkshire, and Landrace were measured by 16S rRNA gene sequencing. Fecal metabolite compositions of the three pig breeds were detected using untargeted metabolomics.ResultsThere were significant differences in the gut microbiomes of the three species, indicating that host breed affects the diversity and structure of gut microbiota. Several breed-associated microorganisms were identified at different taxonomic levels. Notely, most microbial taxa were annotated as Lactobacillacea, Muribaculaceae, and Oscillospiraceae. Several bacteria, including Lactobacillus, Subdoligranulum, Faecalibacterium, Oscillospira, Oscillospiraceae_UCG-002, and Christensenellaceae_R-7_group, could be considered as biomarkers for improving the backfat thickness (BF) for commercial pigs. Additionally, KEGG analysis of gut microbiota further revealed that arginine biosynthesis, pyruvate metabolism, and fatty acid biosynthesis varied greatly among pig breeds. Multiple gut bacterial metabolites (e.g., spermidine, estradiol, and palmitic acid) were identified as breed-associated. Mediation analysis ultimately revealed the cross-talk among gut microbiota, metabolites, and BF thickness, proclaiming that the microbial and metabolic biomarkers identified in this study could be used as biomarkers for improving BF phenotype.ConclusionsThis work provides vital insights into breed effects on gut microbiota and metabolite compositions of commercial pigs and uncovers potential biomarkers that are significant for pig breed improvement.

A shift in substrate requirement might cause speciation of the lichenized fungi, Varicellaria hemisphaerica and V. lactea (Pertusariales, Ascomycota)

Proper species recognition is required to correctly estimate species preferences and their vulnerability or for eco-evolutionary inference. Varicellaria hemisphaerica and Varicellaria lactea are almost completely morphologically homogeneous species with unclear identification features. To evaluate the importance of morphological, chemical, and ecological characteristics used in recognition of these species, we tested 670 specimens, of which 42 were analyzed phylogenetically using nucITS rDNA, SSU rDNA, and LSU rDNA markers. This integrated taxonomical approach showed that V. hemisphaerica is distinct from V. lactea, and that substrate requirements, together with phylogenetic differences and the size of soredia, differentiate these species. The chemical composition of secondary lichen metabolites in both analyzed species showed similar variation and, therefore, this feature is not diagnostic in species recognition, although suggested by previous studies. The potential speciation of the two species seems to be caused by the shift in the substrate requirements.

Dietary modulation of human milk bioactives is associated with maternal FUT2 secretor phenotype: an exploratory analysis of carotenoids and polyphenol metabolites.

Maternal diet modifies profiles of human milk oligosaccharides (HMOs), carotenoids, and polyphenols in human milk (HM). However, substantial variability in profiles exists between women, highlighting the complexity of non-dietary factors modulating these profiles. The objective of this study was to carry out a secondary analysis exploring the effect of maternal diet on HM carotenoids and polyphenols and relationships between dietary modulation of HM bioactives (carotenoids, polyphenols, and oligosaccharides) and maternal α1,2-fucosyltransferase 2 (FUT2) secretor phenotype. In this pilot study, 16 exclusively breastfeeding women with obesity were enrolled between 4 and 5 months postpartum. The women were provided a 4-week meal plan consistent with the 2020 Dietary Guidelines for Americans (DGA). HM was collected for 24 h at baseline and post-intervention. Maternal FUT2 secretor phenotype was determined by 2'-fucosyllactose concentration in HM (non-secretor: < 100 nmol/ml; secretor: ≥100 nmol/ml). Concentrations of carotenoids and HMOs were determined by LC and polyphenol metabolites by UPLC-MS/MS. Thirteen women completed the study (6 secretors, 7 non-secretors). The change in HM concentrations of the HMOs lacto-N-tetraose (LNT, p = 0.007), lacto-N-fucopentaose II (LNFP II, p = 0.02), difucosyllacto-N-tetraose (DFLNT, p = 0.003), and disialyllacto-N-tetraose (DSLNT, p = 0.003) and polyphenol metabolites 4-hydroxybenzoic acid (4-HBA, p = 0.08) and ferulic acid (p = 0.02) over the intervention time frame was differentially associated with maternal secretor status. 4-HBA and ferulic acid positively correlated with HMOs LNT and DSLNT (rrm = 0.82-0.90, p = 0.03-0.06) for secretors but not for non-secretors. Only secretors demonstrated a negative correlation between 4-HBA and DFLNT (rrm = -0.94, p = 0.001). The influence of maternal diet on composition of HMOs and polyphenol metabolites in HM differs based on maternal secretor status. Consideration of non-dietary factors is needed to evaluate differences in response of HM bioactives to dietary modulation.

Integrated Metabolomics and Transcriptomics Analyses Reveal the Regulatory Mechanisms of Anthocyanin and Carotenoid Accumulation in the Peel of Coffea arabica.

The color of coffee fruits is influenced by several factors, including cultivar, ripening stage, and metabolite composition. However, the metabolic accumulation of pigments and the molecular mechanisms underlying peel coloration during the ripening process of Coffea arabica L. remain relatively understudied. In this study, UPLC-MS/MS-based metabolomics and RNA sequencing (RNA-seq)-based transcriptomics were integrated to investigate the accumulation of anthocyanins and carotenoids in the peel of Coffea arabica at different ripening stages: green peel (GP), green-yellow peel (GYRP), red peel (RP), and red-purple peel (RPP). This integration aimed at elucidating the molecular mechanisms associated with these changes. A total of ten anthocyanins, six carotenoids, and thirty-five xanthophylls were identified throughout the ripening process. The results demonstrated a gradual decrease in the total carotenoid content in the peel with fruit maturation, while anthocyanin content increased significantly. Notably, the accumulation of specific anthocyanins was closely associated with the transition of peel colors from green to red. Integrated metabolomics and transcriptomics analyses identified the GYRP stage as critical for this color transition. A weighted gene co-expression network analysis (WGCNA) revealed that enzyme-coding genes such as 3AT, BZ1, and lcyE, along with transcription factors including MYB, NAC, and bHLH, which interact with PHD and SET TR, may regulate the biosynthesis of anthocyanins and carotenoids, thereby influencing peel pigmentation. These findings provide valuable insights into the molecular mechanisms underlying the accumulation of anthocyanins and carotenoids in Coffea arabica peel during fruit maturation.

Increased artemisinin production in Artemisia annua L. by co-overexpression of six key biosynthetic enzymes

Malaria remains a global health issue, especially in resource-limited regions. Artemisinin, a key antimalarial compound from Artemisia annua, is crucial for treatment, but low natural yields hinder large-scale production. In this study, we employed advanced transgenic technology to co-overexpress six key biosynthetic enzymes—Isopentenyl Diphosphate Isomerase (IDI), Farnesyl Pyrophosphate Synthase (FPS), Amorpha 4,11-diene Synthase (ADS), cytochrome P450 monooxygenase (CYP71AV1), cytochrome P450 oxidoreductase (AACPR) and artemisinic aldehyde D11 reductase (DBR2)—in A. annua to significantly enhance artemisinin production. Our innovative approach utilized a co-expression strategy to optimize the artemisinin biosynthetic pathway, leading to a remarkable up to 200 % increase in artemisinin content in T1 transgenic plants compared to non-transgenic controls. The stability and efficacy of this transformation were confirmed in subsequent generations (T2), achieving a potential 232 % increase in artemisinin levels. Additionally, we optimized transgene expression to maintain plant growth and development, and performed untargeted metabolite analysis using GC–MS, which revealed significant changes in metabolite composition among T2 lines, indicating effective diversion of farnesyl diphosphate into the artemisinin pathway. This metabolic engineering breakthrough offers a promising and scalable solution for enhancing artemisinin production, representing a major advancement in the field of plant biotechnology and a potential strategy for more cost-effective malaria treatment.

Metabolomic profiling of in vitro and in situ grown Nilgiris tea reveals unique signatures for breeding decaffeinated varieties

This study was begun by establishing an in vitro culture in UPASI 9, a Nilgiris tea clone (Camellia sinensis) by optimising various factors. Anatomical studies demonstrated that use of lower carbendazim concentration for sterilisation (0.2%) produced viable and healthy explants for callus initiation. To confirm the genetic consistency of the regenerated plants, gene-specific SSR markers were developed and utilised. GC-MS was employed to analyse volatile metabolites extracted from callus, stem, micro shoots, and leaves of the UPASI 9 tea genotype. The results revealed distinct compositions of metabolites in each sample. More interestingly, caffeine was exclusively detected in leaf samples but absent in all other investigated tissues, despite the presence of Tea Caffeine Synthase (TCS) gene-specific SSRs. Thus, this study provided unique information on the absence of caffeine in in vitro grown Nilgiris tea clone, UPASI 9, as decaffeinated tea has a unique niche in the global market.