沙棘粕醇提物的定性定量分析及其对衰老小鼠肝脏抗氧化指标的影响

Qualitative and Quantitative Analysis of Sea Buckthorn Seed Extract and Its Antioxidant Effect in Liver of Aging Mice

利用沙棘粕为研究对象,经乙醇提取、大孔吸附树脂纯化获得沙棘粕醇提物(sea buckthorn seed extract,SBSE)。利用超高效液相色谱-串联四极杆飞行时间质谱(ultra-performance liquid chromatography coupled with quadruple time-of-flight tandem mass spectrometry,UPLC-QTOF-MS/MS)对SBSE进行定性研究,通过超高效液相色谱-串联三重四极杆质谱(ultra performance liquid chromatography coupled with triple quadruple mass spectrometry,UPLC-QQQ-MS)定量分析SBSE活性成分。最后研究分析SBSE对以D-半乳糖诱导的衰老小鼠的抗氧化功效,不同浓度SBSE灌胃衰老小鼠模型42 d后,比较小鼠肝脏组织中总抗氧化能力(total antioxidant capacity,T-AOC)、谷胱甘肽过氧化物酶(glutathione peroxidase,GSH-Px)、超氧化物歧化酶(superoxide dismutase,SOD)和过氧化氢酶(catalase,CAT)活性以及丙二醛(malondialdehyde,MDA)和脂褐素(lipofuscin,LP)水平。结果表明:通过UPLC-QTOF-MS/MS分析方法共鉴定出24种化合物,其中与标准品比对共确认14种物质;利用UPLC-QQQ-MS法,采用负离子多反应监测方式,建立SBSE的14种化合物的定量分析方法,通过检测分析,SBSE中山柰酚及其衍生物所占比例最大,质量浓度为(667.94±5.61)μg/mL;其次为槲皮素及其衍生物((204.01±0.04)μg/mL),原花青素类物质为(33.97±0.49)μg/mL。经过42 d的衰老小鼠灌胃实验,与模型组相比,灌胃一定剂量SBSE可以提高小鼠肝脏组织中GSH-Px、SOD和CAT的活力;低剂量组小鼠肝脏组织中的T-AOC水平极显著高于模型组(P<0.01);3种灌胃剂量小鼠的肝脏组织LP和MDA含量均极显著低于模型组(P<0.01)。

This research studied the qualitative analysis of sea buckthorn seed extract (SBSE), prepared from sea buckthorn seed residue by ethanol extraction and macroporous resin purification, by ultra-performance liquid chromatography coupled with quadruple time-of-flight tandem mass spectrometry (UPLC-QTOF-MS/MS). The bioactive components of SBSE were quantitatively determined by ultra performance liquid chromatography coupled with triple quadruple mass spectrometry (UPLC-QQQ-MS). Meanwhile, the antioxidant effect of SBSE in D-galactose-induced aging mice was studied. The mice were administrated by gavage with SBSE at different dosages for 42 days. Total antioxidant capacity (T-AOC), the activities of glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and catalase (CAT) and the contents of malondialdehyde (MDA) and lipofuscin (LP) in liver tissue were measured. The results obtained were as follows: A total of 24 compounds were identified by UPLC-QTOF-MS/MS, 14 compounds of which were characterized by comparing with reference substances and quantitatively determined by UPLC-QQQ-MS in the negative ion multiple reaction monitoring (MRM) mode. Kaempferol and its derivatives were the most abundant constituents in SBSE ((667.94 ± 5.61) μg/mL), quercetin and its derivatives were in the middle ((204.01 ± 0.04) μg/m L), and procyanidins were the least abundant ((33.97 ± 0.49) μg/m L). The administration of SBSE to aging mice could increase the activities of GSH-Px, SOD and CAT in liver tissue. Compared with the D-galactose-induced model group, hepatic T-AOC activity in the low-dosage group was significantly increased (P < 0.01), and the levels of MDA and LP were decreased significantly by SBSE at three dosages (P < 0.01).