Ramdas, Mausam Singh, Amrat Pal Singh, Neerja Tiwari, Karuna Shanker
{"title":"UHPLC-Orbitrap-MS/MS代谢物图谱结合HPLC-PDA靶向化学指纹图谱揭示了地理环境（G × E）对山核桃根化学成分的影响。","authors":"Ramdas, Mausam Singh, Amrat Pal Singh, Neerja Tiwari, Karuna Shanker","doi":"10.1016/j.ymeth.2025.09.010","DOIUrl":null,"url":null,"abstract":"<p><p>Aegle marmelos (AM) (Rutaceae family) holds significant economic value due to its uses in food, traditional medicines, and timber. Studies have confirmed that its extracts and phytochemicals possess various pharmacological actions, including anti-obesity, diuretic, anti-inflammatory, and chemopreventive. However, challenges remain due to the limited information on its secondary metabolites and the lack of validated methodologies. Natural variability in the raw material further complicates the assessment of therapeutic efficacy. Hence, we have conducted the untargeted metabolite profiling of A. marmelos root (AMR) using UHPLC-Orbitrap-MS/MS analysis, identifying 69 phytochemicals of diverse classes. Additionally, we have isolated and characterised 09 key phytochemicals to evaluate the chemical variability in AMR from 15 geographical locations. HPLC-PDA method, compliant with ICH Q2 (R2) guidelines, was developed and validated to quantitate 02 alkaloids (skimmianine, O-Methyl tembamide) and 07 coumarins [umbelliferone, xanthotoxol, marmin, and 7-(6-Hydroxy-7-methoxy-3,7-dimethyl-(2E)-2-octenyloxy) coumarin, 7-(3,7-Dimethyl-6-oxo-(2E)-2-octenyloxy) coumarin, marmelosin, and auraptene]. Chemometric analysis has distinguished 15 AMR ecotypes, and hierarchical cluster analysis resulted in AM-S4 as a distinctive ecotype, which was verified by principal component analysis with 96 % data variance. Partial least square-discriminate analysis predicted a relationship between targeted metabolites and ecotypes. The metabolites associated with a discriminatory pattern of AMR ecotypes were identified by variable importance for projection (VIP) score. Unlike previous reports, the present method fulfils the ISO: 17025-2017 requirement by evaluating the measurement of uncertainty (MU) to ensure the accuracy and traceability of the results. Overall, present study summarises comprehensive metabolite profiling, multi-components classification of AMR to define genetic-environment (G × E) effect, and quality evaluation of AMR derived medicinal product.</p>","PeriodicalId":390,"journal":{"name":"Methods","volume":" ","pages":""},"PeriodicalIF":4.3000,"publicationDate":"2025-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"UHPLC-Orbitrap-MS/MS metabolite profiling combined with HPLC-PDA targeted chemical fingerprint reveals the geographic-environment (G × E) effect on chemical constituents of Bilvamula (Aegle marmelos root).\",\"authors\":\"Ramdas, Mausam Singh, Amrat Pal Singh, Neerja Tiwari, Karuna Shanker\",\"doi\":\"10.1016/j.ymeth.2025.09.010\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>Aegle marmelos (AM) (Rutaceae family) holds significant economic value due to its uses in food, traditional medicines, and timber. Studies have confirmed that its extracts and phytochemicals possess various pharmacological actions, including anti-obesity, diuretic, anti-inflammatory, and chemopreventive. However, challenges remain due to the limited information on its secondary metabolites and the lack of validated methodologies. Natural variability in the raw material further complicates the assessment of therapeutic efficacy. Hence, we have conducted the untargeted metabolite profiling of A. marmelos root (AMR) using UHPLC-Orbitrap-MS/MS analysis, identifying 69 phytochemicals of diverse classes. Additionally, we have isolated and characterised 09 key phytochemicals to evaluate the chemical variability in AMR from 15 geographical locations. HPLC-PDA method, compliant with ICH Q2 (R2) guidelines, was developed and validated to quantitate 02 alkaloids (skimmianine, O-Methyl tembamide) and 07 coumarins [umbelliferone, xanthotoxol, marmin, and 7-(6-Hydroxy-7-methoxy-3,7-dimethyl-(2E)-2-octenyloxy) coumarin, 7-(3,7-Dimethyl-6-oxo-(2E)-2-octenyloxy) coumarin, marmelosin, and auraptene]. Chemometric analysis has distinguished 15 AMR ecotypes, and hierarchical cluster analysis resulted in AM-S4 as a distinctive ecotype, which was verified by principal component analysis with 96 % data variance. Partial least square-discriminate analysis predicted a relationship between targeted metabolites and ecotypes. The metabolites associated with a discriminatory pattern of AMR ecotypes were identified by variable importance for projection (VIP) score. Unlike previous reports, the present method fulfils the ISO: 17025-2017 requirement by evaluating the measurement of uncertainty (MU) to ensure the accuracy and traceability of the results. Overall, present study summarises comprehensive metabolite profiling, multi-components classification of AMR to define genetic-environment (G × E) effect, and quality evaluation of AMR derived medicinal product.</p>\",\"PeriodicalId\":390,\"journal\":{\"name\":\"Methods\",\"volume\":\" \",\"pages\":\"\"},\"PeriodicalIF\":4.3000,\"publicationDate\":\"2025-10-02\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Methods\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://doi.org/10.1016/j.ymeth.2025.09.010\",\"RegionNum\":3,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"BIOCHEMICAL RESEARCH METHODS\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Methods","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1016/j.ymeth.2025.09.010","RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"BIOCHEMICAL RESEARCH METHODS","Score":null,"Total":0}
UHPLC-Orbitrap-MS/MS metabolite profiling combined with HPLC-PDA targeted chemical fingerprint reveals the geographic-environment (G × E) effect on chemical constituents of Bilvamula (Aegle marmelos root).
Aegle marmelos (AM) (Rutaceae family) holds significant economic value due to its uses in food, traditional medicines, and timber. Studies have confirmed that its extracts and phytochemicals possess various pharmacological actions, including anti-obesity, diuretic, anti-inflammatory, and chemopreventive. However, challenges remain due to the limited information on its secondary metabolites and the lack of validated methodologies. Natural variability in the raw material further complicates the assessment of therapeutic efficacy. Hence, we have conducted the untargeted metabolite profiling of A. marmelos root (AMR) using UHPLC-Orbitrap-MS/MS analysis, identifying 69 phytochemicals of diverse classes. Additionally, we have isolated and characterised 09 key phytochemicals to evaluate the chemical variability in AMR from 15 geographical locations. HPLC-PDA method, compliant with ICH Q2 (R2) guidelines, was developed and validated to quantitate 02 alkaloids (skimmianine, O-Methyl tembamide) and 07 coumarins [umbelliferone, xanthotoxol, marmin, and 7-(6-Hydroxy-7-methoxy-3,7-dimethyl-(2E)-2-octenyloxy) coumarin, 7-(3,7-Dimethyl-6-oxo-(2E)-2-octenyloxy) coumarin, marmelosin, and auraptene]. Chemometric analysis has distinguished 15 AMR ecotypes, and hierarchical cluster analysis resulted in AM-S4 as a distinctive ecotype, which was verified by principal component analysis with 96 % data variance. Partial least square-discriminate analysis predicted a relationship between targeted metabolites and ecotypes. The metabolites associated with a discriminatory pattern of AMR ecotypes were identified by variable importance for projection (VIP) score. Unlike previous reports, the present method fulfils the ISO: 17025-2017 requirement by evaluating the measurement of uncertainty (MU) to ensure the accuracy and traceability of the results. Overall, present study summarises comprehensive metabolite profiling, multi-components classification of AMR to define genetic-environment (G × E) effect, and quality evaluation of AMR derived medicinal product.
期刊介绍：
Methods focuses on rapidly developing techniques in the experimental biological and medical sciences.
Each topical issue, organized by a guest editor who is an expert in the area covered, consists solely of invited quality articles by specialist authors, many of them reviews. Issues are devoted to specific technical approaches with emphasis on clear detailed descriptions of protocols that allow them to be reproduced easily. The background information provided enables researchers to understand the principles underlying the methods; other helpful sections include comparisons of alternative methods giving the advantages and disadvantages of particular methods, guidance on avoiding potential pitfalls, and suggestions for troubleshooting.
求助内容：
应助结果提醒方式：
京公网安备 11010802042870号
