大蒜化学成分不同测定方法比较及相关性探讨

目的:采用不同方法测定大蒜中化学成分含量并进行相关性探讨。方法:本实验参考美国药典大蒜中蒜氨酸含量测定方法,采用C₁₈色谱柱,以乙腈-1,4-二氧六环-四氢呋喃-0.045 mol·L^-1磷酸二氢钠溶液(25:2.9:2.2:69.9)为流动相,供试品经灭酶、衍生化处理后在337 nm下测定蒜氨酸含量;参考印度药典大蒜中蒜氨酸含量测定方法,采用C₁₈色谱柱,以0.1%磷酸为流动相,供试品经热水灭酶处理后,直接在210 nm下测定蒜氨酸含量;采用课题组建立的大蒜中蒜氨酸含量测定方法,采用C₁₈色谱柱,以0.04%三氟乙酸为流动相,供试品经微波灭酶处理后,直接在214 nm下测定蒜氨酸含量;参考欧洲药典和英国药典大蒜粉中大蒜辣素含量测定方法,采用C₁₈色谱柱,以甲醇-0.1%甲酸(60:40)为流动相,将供试品捣碎后,在254 nm下测定大蒜中大蒜辣素含量;参考中国药典大蒜中大蒜素含量测定方法,采用C₁₈色谱柱,以甲醇-0.1%甲酸(75:25)为流动相,将供试品捣碎、回流处理后在210 nm下测定大蒜中大蒜素含量。结果:课题组建立方法与美国药典方法测得蒜氨酸含量没有显著性差异,印度药典方法灭酶不完全,蒜氨酸测定结果偏低,欧洲药典方法合理可行,大蒜辣素测定值可反应真实情况,中国药典方法测定大蒜素的结果与多种因素相关。结论:5种方法测定的指标不同,欧盟药典和英国药典以大蒜辣素为测定指标,中国药典以大蒜素为测定指标,美国药典、印度药典和课题组自建方法都以蒜氨酸为测定指标;但方法之间仍具有一定差异,灭酶方法不同:美国药典采用蒜酶抑制剂灭酶,印度药典用热水灭酶,课题组方法采用微波灭酶;方法原理不同:美国药典采用柱前衍生化法,印度药典和课题组方法则直接测定蒜氨酸含量。

Objective: To determine the chemical constituents in garlic by different means and study the correlation between them.Methods: In this study, according to the United States Pharmacopoeia(USP), the content determination method of alliin in garlic, using C₁₈ chromatographic column, acetonitrile-1, 4-dioxane-tetrahydrofuran-0.045 mol L-1 sodium dihydrogen phosphate solution(25:2.9:2.2:69.9) as mobile phase, alliin of the sample was contented at 337 nm after the process of allinase destruction, derivatization. The Indian Pharmacopeia(IP), the content determination method of alliin in garlic, using C₁₈ chromatographic column, 0.1% phosphoric acid as mobile phase, alliin of the sample was contented at 210 nm after killing allinase by hot water; the method of our team, the content determination method of alliin in garlic, using C₁₈ chromatographic column, with 0.04% trifluoroacetic acid as mobile phase, alliin of the sample was contented at 210 nm after killing allinase by microwave;In the European Pharmacopoeia(EP) and the British Pharmacopoeia(BP), the content determination method of allicin in garlic powder, using C₁₈ chromatographic column, methanol-0.1% formic acid(60:40) as mobile phase, allicin of the sample was contented at 254 nm after the garlic is mashed; In the Chinese Pharmacopoeia(ChP), the content determination method of alltride in garlic, using C₁₈ chromatographic column, methanol-0.1% formic acid(75:25) as mobile phase, alltride in the garlic was contented at 210 nm after the garlic was mashed and reflux.Results: There were no significant difference between our team and USP in the determination of the content of alliin, in IP, allinase inactivation incomplete, the content of alliin was lower, EP method was reasonable and feasible, the determination of allicin could reflect the actual situation, in ChP, the determination of allitride content and the results correlate with many factors.Conclusion: The determining quota is different in these methods, in EP and BP, allicin as index, in ChP, alltride as index, in USP, IP and the method of our team, alliin is determining quota, but there are some differences among the three methods, the method of killing allinase: allinase is killed, in USP by allinase inhibitor, in IP by hot water, in the method of our team by microwave; the theory of these methods: contents of alliin in garlic, in USP with pre-column derivatization method, in IP and the method of our team with direct method.