Abstract：
OBJECTIVE: Microbial transformation to modify saponins and enhance their biological activities has received increasing attention in recent years. This study aimed to screen the strain that can biotransform notoginsenoside R1, identify the product and study its biological activity.
RESULTS: A lactic acid bacteria strain S165 with glycosidase-producing activity was isolated from traditional Chinese fermented foods, which was identified and grouped according to API 50 CHL kit and 16S rDNA sequence analysis. Subsequently, notoginsenoside R1 underwent a 30-day fermentation period by the strain S165, and the resulting products were analyzed using High-performance liquid chromatography (HPLC), Ultra-performance liquid chromatography (UPLC)-mass spectrometry (MS)/MS, and 13C-Nuclear magnetic resonance (NMR) techniques. Employing a model of Lipopolysaccharide (LPS)-induced damage to Caco-2 cells, the damage of Caco-2 cells was detected by Hoechst 33 258 staining, and the activity of notoginsenoside R1 biotransformation product was investigated by CCK-8 and western blotting assay. The strain S165 was identified as Lactiplantibacillus plantarum and was used to biotransform notoginsenoside R1. Through a 30-day biotransformation, L. plantarum S165 predominantly converts notoginsenoside R1 into 3β,12β-dihydroxydammar-(E)-20(22),24-diene-6-O-β-D-xylopyranosyl-(1→2)-β-D-glucopyranoside, temporarily named notoginsenoside T6 (NGT6) according to HPLC, UPLC-MS/MS, and 13C-NMR analysis. Results from CCK-8 and Hoechst 33258 staining indicated that the ability notoginsenoside T6 to alleviate the intestinal injury induced by LPS in the Caco-2 cell was stronger than that of notoginsenoside R1. In addition, Western blotting result showed that notoginsenoside T6 could prevent intestinal injury by protecting tight junction proteins (Claudin-1, Occludin, and ZO-1).
CONCLUSIONS: Notoginsenoside R1 was biotransformed into the notoginsenoside T6 by L. plantarum S165, and the biotransformed product showed an enhanced intestinal protective effect in vitro.
RESULTS: A lactic acid bacteria strain S165 with glycosidase-producing activity was isolated from traditional Chinese fermented foods, which was identified and grouped according to API 50 CHL kit and 16S rDNA sequence analysis. Subsequently, notoginsenoside R1 underwent a 30-day fermentation period by the strain S165, and the resulting products were analyzed using High-performance liquid chromatography (HPLC), Ultra-performance liquid chromatography (UPLC)-mass spectrometry (MS)/MS, and 13C-Nuclear magnetic resonance (NMR) techniques. Employing a model of Lipopolysaccharide (LPS)-induced damage to Caco-2 cells, the damage of Caco-2 cells was detected by Hoechst 33 258 staining, and the activity of notoginsenoside R1 biotransformation product was investigated by CCK-8 and western blotting assay. The strain S165 was identified as Lactiplantibacillus plantarum and was used to biotransform notoginsenoside R1. Through a 30-day biotransformation, L. plantarum S165 predominantly converts notoginsenoside R1 into 3β,12β-dihydroxydammar-(E)-20(22),24-diene-6-O-β-D-xylopyranosyl-(1→2)-β-D-glucopyranoside, temporarily named notoginsenoside T6 (NGT6) according to HPLC, UPLC-MS/MS, and 13C-NMR analysis. Results from CCK-8 and Hoechst 33258 staining indicated that the ability notoginsenoside T6 to alleviate the intestinal injury induced by LPS in the Caco-2 cell was stronger than that of notoginsenoside R1. In addition, Western blotting result showed that notoginsenoside T6 could prevent intestinal injury by protecting tight junction proteins (Claudin-1, Occludin, and ZO-1).
CONCLUSIONS: Notoginsenoside R1 was biotransformed into the notoginsenoside T6 by L. plantarum S165, and the biotransformed product showed an enhanced intestinal protective effect in vitro.
摘要：
目的：微生物转化对皂苷进行修饰并增强其生物学活性近年来受到越来越多的关注。本研究旨在筛选能生物转化三七皂苷R1的菌株，鉴定产品并研究其生物学活性。
方法：从中国传统发酵食品中分离出一株具有产糖苷酶活性的乳酸菌S165，根据API50CHL试剂盒和16SrDNA序列分析进行鉴定和分组。随后，三七皂苷R1经过菌株S165的30天发酵期,UPLC-MS/MS,和13C-NMR技术。采用LPS诱导的Caco-2细胞损伤模型，通过Hoechst33.258染色检测到caco-2细胞的损伤，用CCK-8和免疫印迹法检测三七皂苷R1生物转化产物的活性。
结果：菌株S165被鉴定为植物乳杆菌，并用于生物转化三七皂苷R1。通过30天的生物转化,植物乳杆菌S165主要将三七皂苷R1转化为3β，12β-二羟基多巴胺-（E）-20（22），24-二烯-6-O-β-D-吡喃木糖基-(1→2)-β-D-吡喃葡萄糖苷,根据HPLC暂时命名为三七皂苷T6（NGT6），UPLC-MS/MS和13C-NMR分析。CCK-8和Hoechst33.258染色结果表明，三七皂苷T6减轻LPS诱导的Caco-2细胞肠损伤的能力强于三七皂苷R1。此外,Westernblotting结果显示三七皂苷可通过保护紧密连接蛋白(Claudin-1,Occludin,ZO-1).
结论：三七皂苷R1经植物乳杆菌S165生物转化为三七皂苷T6，其生物转化产物在体外具有增强的肠道保护作用。
方法：从中国传统发酵食品中分离出一株具有产糖苷酶活性的乳酸菌S165，根据API50CHL试剂盒和16SrDNA序列分析进行鉴定和分组。随后，三七皂苷R1经过菌株S165的30天发酵期,UPLC-MS/MS,和13C-NMR技术。采用LPS诱导的Caco-2细胞损伤模型，通过Hoechst33.258染色检测到caco-2细胞的损伤，用CCK-8和免疫印迹法检测三七皂苷R1生物转化产物的活性。
结果：菌株S165被鉴定为植物乳杆菌，并用于生物转化三七皂苷R1。通过30天的生物转化,植物乳杆菌S165主要将三七皂苷R1转化为3β，12β-二羟基多巴胺-（E）-20（22），24-二烯-6-O-β-D-吡喃木糖基-(1→2)-β-D-吡喃葡萄糖苷,根据HPLC暂时命名为三七皂苷T6（NGT6），UPLC-MS/MS和13C-NMR分析。CCK-8和Hoechst33.258染色结果表明，三七皂苷T6减轻LPS诱导的Caco-2细胞肠损伤的能力强于三七皂苷R1。此外,Westernblotting结果显示三七皂苷可通过保护紧密连接蛋白(Claudin-1,Occludin,ZO-1).
结论：三七皂苷R1经植物乳杆菌S165生物转化为三七皂苷T6，其生物转化产物在体外具有增强的肠道保护作用。
