Department of Pharmacy, China Pharmaceutical University Nanjing Drum Tower Hospital, Nanjing 210008, China; School of Basic Medicine and Clinical Pharmacy, China Pharmaceutical University, Nanjing 211112, China.
Department of Pharmacy, Nanjing Drum Tower Hospital, The Affiliated Hospital of Nanjing University Medical School, Nanjing 210008, China.
J Pharm Biomed Anal. 2024 Jan 5;237:115784. doi: 10.1016/j.jpba.2023.115784. Epub 2023 Oct 10.
B vitamins in the human distal gut are primarily derived from the gut microbiota because daily dietary vitamins are fully absorbed in the small intestine under normal dietary and physiological conditions. Quantitative determination of B vitamins in the distal gut and faecal samples is crucial for understanding the intricate relationship between gut B vitamins, gut microbiota, and host health. In this study, we developed a rapid, robust, and reliable method with a simple extraction procedure for the simultaneous analysis of seven B vitamins in human faeces using high-performance liquid chromatography-electrospray ionisation-tandem mass spectrometry (HPLC-ESI-MS/MS) with stable isotope-labelled internal standards. A protein precipitation approach using methanol as the precipitant was employed to extract vitamin B from human faecal samples. Seven B vitamins were adequately separated and quantified within 9 min by HPLC-ESI-MS/MS with a Pursuit PFP column (2.0 ×150 mm, 3.0 µm), including vitamins B1, B2, B3, B5, pyridoxic acid, pyridoxine, and B7. The lower limits of quantification were within the range of 0.1-25 ng mL. The intra-day and inter-day precision and accuracy were both within 15 %. The validated method was successfully applied to 55 faecal samples collected from healthy individuals, patients with type 2 diabetes, and obese patients. Compared with healthy controls, obese patients had lower faecal concentrations of vitamins B1 and B3 and pyridoxic acid, whereas patients with type 2 diabetes had lower faecal concentrations of vitamins B1 and B5.
人体远端肠道中的 B 族维生素主要来源于肠道微生物群，因为在正常饮食和生理条件下，每日膳食中的维生素会在小肠中被完全吸收。定量测定远端肠道和粪便样本中的 B 族维生素对于了解肠道 B 族维生素、肠道微生物群和宿主健康之间的复杂关系至关重要。在本研究中，我们开发了一种快速、稳健且可靠的方法，该方法采用高效液相色谱-电喷雾离子化-串联质谱（HPLC-ESI-MS/MS），使用稳定同位素标记的内标物，结合简单的提取程序，可同时分析人体粪便中的七种 B 族维生素。采用甲醇沉淀法从人粪便样品中提取维生素 B。采用 Pursuit PFP 柱（2.0×150mm，3.0µm），通过 HPLC-ESI-MS/MS 可在 9 min 内充分分离和定量七种 B 族维生素，包括维生素 B1、B2、B3、B5、吡哆醛、吡哆醇和 B7。定量下限在 0.1-25ng·mL 范围内。日内和日间精密度和准确度均在 15%以内。该验证方法成功应用于 55 例来自健康个体、2 型糖尿病患者和肥胖患者的粪便样本。与健康对照组相比，肥胖患者粪便中维生素 B1 和 B3 以及吡哆醛的浓度较低，而 2 型糖尿病患者粪便中维生素 B1 和 B5 的浓度较低。
