凤丹愈伤组织培养体系建立及丹皮酚含量测定

Callus culture system and paeonol determination of Paeonia ostii T.Hong et J.X.Zhang var. lishizhenii B.A.Shen

本文旨在建立凤丹(Paeonia ostii var.lishizhenii)愈伤组织培养及制取丹皮酚的技术体系,为细胞工程制药提供技术基础。凤丹去皮种子以70%乙醇(V/V)浸泡2.0 min和有效氯1.0%的二氯异氰尿酸钠溶液浸泡20 min,去污染率高达97.78%;剥取种胚接种在含6-苄氨基嘌呤1.5 mg/L、2,4-二氯苯氧乙酸2.0 mg/L的MS培养基上,愈伤组织诱导率达90.00%;愈伤组织在含6-苄氨基嘌呤1.8 mg/L、萘乙酸0.6 mg/L、蔗糖30 g/L、水解酪蛋白0.4 g/L、琼脂5 g/L、pH 7.3的改良WPM培养基上暗培养25天,增殖倍数达3.53,褐化率为11.78%;基于天然产物丹皮酚含量和愈伤组织生长曲线,培养25天时丹皮酚含量达到最高,为最佳收获期。

The paper aims to establish a technical system for the cultivation of paeonol in P.ostii var.lishizhenii callus,and to provide a technical basis for cell engineering pharmacy.The mature and plump seeds of P.ostii var.lishizhenii were peeled and soaked in 70% ethanol(V/V) for 2.0 min and then soaked in 1.0% sodium dichloroisocyanurate solution for 20 minutes.The seed decontamination rate was 97.78%.Place sterilized on MS medium and then add 6-BA 1.5 mg/L and 2,4-D 2.0 mg/L.The callus induced was 90.00%.Optimized medium for cell culture was the modified WPM with 6-BA 1.8 mg/L,NAA 0.6 mg/L,sucrose 30 g/L,hydrolyzed casein 0.4 g/L,agar 5 g/L and pH 7.3.In the conditions,cell proliferation ratio was 3.53 and the browning rate was 11.78% on the 25th day.Based on the natural product paeonol content and callus growth curve,paeonol content reached the highest when cultured for 25 days,which was the best harvest time.