GC-MS/-HTC-IRMS同时测定瘤胃液中乙酸含量及其氢稳定同位素比值  

Simultaneous Determination of Content and Stable Hydrogen Isotope Ratio of Acetic Acid in Rumen Fluid by GC-MS/-HTC-IRMS

作　　者：贺珍 袁红朝[1,2] 张秀敏[1] 张丽萍[1] 耿梅梅[1] 许丽卫[1] 李春勇[1] 彭灿 陈闻 唐思宇 钟欢 王久荣[1] HE Zhen;YUAN Hongzhao;ZHANG Xiumin;ZHANG Liping;GENG Meimei;XU Liwei;LI Chunyong;PENG Can;CHEN Wen;TANG Siyu;ZHONG Huan;WANG Jiurong(Institute of Subtropical Agriculture,Chinese Academy of Sciences,Changsha,Hunan 410125,China;Key Laboratory of Agro-ecological Processes in Subtropical Region,Institute of Subtropical Agriculture,Chinese Academy of Sciences,Changsha,Hunan 410125,China)

机构地区：[1]中国科学院亚热带农业生态过程重点实验室,长沙410125 [2]中国科学院亚热带农业生态研究所,农业生态工程湖南省重点实验室,长沙410125

出　　处：《中国无机分析化学》2025年第5期713-722,共10页Chinese Journal of Inorganic Analytical Chemistry

基　　金：国家自然科学基金资助项目(42177330,U21A2007);中国科学院技术支撑人才项目;中国科学院仪器功能开发项目(2025g040,2023g132);中国科学院开放课题项目。

摘　　要：日粮纤维在反刍动物瘤胃内降解生成挥发性脂肪酸(VFA)和甲烷,其中乙酸是VFA的主要成分之一。氢是瘤胃内连接日粮纤维降解生成VFA和甲烷代谢生成的重要中间体。因此,准确量化瘤胃液中乙酸含量及其氢稳定同位素比值对于探明反刍动物瘤胃的氢代谢规律及其对VFA合成路径和甲烷生成的内在影响机理具有重要意义。在前期工作的基础上,以瘤胃液中的乙酸为研究对象,通过液-液萃取的前处理方法,排除了样品基底水中氢的干扰,结合气相色谱-单四极杆质谱仪-热裂解-稳定同位素比例质谱仪(GC-MS/-HTC-IRMS),分析了不同浓度下自然丰度、2H富集乙酸标准溶液的氢稳定同位素比值,开发了瘤胃液中乙酸含量及其氢稳定同位素丰度同步分析的方法。结果表明,乙酸氢量(上柱量)的定量范围为44.4~222.1 nmol,定量相关系数为0.998,氢稳定同位素比值结果也能获得较理想的分析精度,其中自然丰度标准偏差小于2.6‰,在保证理想的测试准确度和精密度条件下,推荐氢上柱量为111 nmol(与参考值差值小于1.2‰,标准偏差小于0.6‰)。2H富集乙酸atom%测试结果在一定范围和参考值(0.014 atom%~0.100 atom%)具有良好线性关系,相关系数达到0.997。将方法应用于氘气供应的湘东黑山羊瘤胃发酵体系的瘤胃液中乙酸含量和氢稳定同位素比值的测定,均能获得较好的精度(RSD小于1.4%)。方法前处理操作简单,检出限低,耗时短,重复性好,实现了瘤胃液中低含量乙酸(氢量在nmol级)含量及其氢稳定同位素比值的同步批量检测。Dietary fiber in ruminants is degraded to yield volatile fatty acids(VFA) and methane.Acetic acid,a major component of VFA,plays a crucial role in regulating the composition and content of VFA and impacts protein metabolism in the rumen.Hydrogen serves as an important intermediate connecting dietary fiber degradation to VFA production and methane metabolism in the rumen.Thus,accurately quantifying the content of acetic acid and its hydrogen stable isotope ratio is highly significant for elucidating:1) the metabolic rules of hydrogen sources;2) their inherent effects on VFA synthetic pathways;and 3) methane generation.Building on previous work,our objective was to establish a suitable pretreatment method for analyzing the acetic acid content and its hydrogen stable isotope ratio in ruminal fluid,aiming to eliminate sample hydrogen interference.Subsequently,by combining gas-chromatography-mass spectrometry/high temperature conversion-specific isotope ratio mass spectrometry(GC-MS/-HTC-IRMS),the hydrogen stable isotope ratios of natural abundance and D-enriched acetic acid standard solutions at different concentrations were analyzed.As a result,a synchronous analysis method for the acetic acid content in ruminal fluid and its hydrogen stable isotope abundance was developed.The results demonstrated that the quantitative range of this method for hydrogen measurement(on-column amount) was 44.4-222.1 nmol of acetic acid,with a correlation coefficient exceeding 0.998.Meanwhile,the hydrogen isotope ratio test could also achieve excellent analysis accuracy,with the standard deviation of natural abundance being less than 2.6‰,which was basically consistent with the given values of standard references.Additionally,the 2H atom% of the enriched acetic acid standard sample(0.014 atom%-0.100 atom%) also exhibited a good linear relationship,with a correlation coefficient reaching 0.997.Finally,the newly established method was applied to Xiangdong black goat ruminants and achieved good reproducibility(RSD<1.4%).Therefore,our new

关 键 词：瘤胃液 乙酸 氢同位素 GC-MS/-HTC-IRMS 

分 类 号：O657.63[理学—分析化学]