| 徐婧,王丹,王俊,赵明磊,李建国.超高效液相色谱-三重四级杆质谱联用法同时定量荔枝果实组织中IAA和ABA[J].中国南方果树,2022,51(1): |
| 超高效液相色谱-三重四级杆质谱联用法同时定量荔枝果实组织中IAA和ABA |
| Simultaneous quantification of indole-3-aceticacid and abscisic acid in litchi fruit tissues by Ultra-high Performance Liquid Chromatography-Tandem Mass Spectrometry |
| 投稿时间：2020-12-19 修订日期：2021-03-04 |
| DOI： |
| 中文关键词: 荔枝 超高效液质联用仪 生长素 脱落酸 定量测定 |
| 英文关键词:Litchi UPLC-MS/MS IAA ABA Quantification |
| 基金项目:国家重点研发计划课题（2020YFD1000103）；广东省重点领域研发计划项目（2018B020202011） |
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| 中文摘要: |
| 摘 要：本研究旨在荔枝上建立应用超高效液相色谱-三重四级杆质谱联用仪（UPLC-MS/MS）同时定量测定两种内源激素-IAA和ABA的新方法。采用UPLC-MS/MS仪器，首先设定IAA和ABA标准品测定所需色谱和质谱条件参数，通过考察不同抗氧化剂 、提取剂、除色剂、萃取剂、提取方式、pH调节对荔枝果实样品中IAA和ABA保留行为及富集效果的影响，确认适合荔枝果实样品前处理流程，最后考察和验证方法准确度及精密度。荔枝样品经预冷的80%丙酮常规振荡浸提，石油醚纯化（纯化前调pH至8~9），乙酸乙酯萃取（加萃取剂前调pH至2~3），氮吹仪浓缩，80%甲醇复溶，内标法定量， BEH C18柱分离，0.1%甲酸乙腈和0.1%甲酸水为流动相，两种激素及其内标物峰型较好，保留时间稳定。IAA检出限为0.283pg/μL，定量限为0.339pg/μL；ABA检出限为0.102pg/μL，定量限为0.943pg/μL。对实际样品中IAA和ABA进行测定，结果显示其相对标准偏差分别为3.75%和1.38%。本研究建立的新方法能够较精确地同时定量检测出荔枝样品中IAA和ABA含量，并为荔枝样品中其他内源激素的测定提供新思路。 |
| 英文摘要: |
| ? Abstract: A new method was developed to simultaneously determine the contents of two endogenous phytohormones, 3-indoacetic acid (IAA) and abscisic acid (ABA), in litchi by ultra- high performance liquid chromatography-tandem mass spectrometry (UPLC- MS/MS). Proper parameters of chromatography and mass spectrometry were firstly assessed for the determination of the contents of IAA and ABA standards. Then pre-processing with different antioxidants, isolation solutions, purifiers, extractants, different extraction approaches, and PH was evaluated upon their effects on the rate of phytohormone retention and enrichment from litchi fruit tissues. At last, the most suitable protocol was developed to test for its accuracy and precision. Stable-labeled internal standards were added after a series of processing steps, including oscillation extraction with pre-chilled 80% acetone, petroleum ether purification (adjusted to pH 8~9 before purification), ethyl acetate extraction (adjusted to pH 2~3 before the addition of extractant), concentration using nitrogen blowing instrument, and re-dissolution in 80% methanol. The separation was performed on a BEH C18 column by using acetonitrile (0.1% formic acid) and water (0.1% formic acid) as mobile phases with gradient elution. The results showed the peak shape of the endogenous phytohormones and the internal standards were sharp and smooth and had stable retention time. The detection limit of IAA and ABA were 0.283 pg/μL and 0.102 pg/μL, and the limit of quantification were 0.339 pg/μL and 0.943 pg/μL, respectively. The relative standard deviations of the IAA and ABA were 3.75% and 1.38% respectively. In conclusion, the method we optimized can accurately determine the contents of IAA and ABA in litchi fruits and sheds new lights on the hormone quantification in litchi. |
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