Studies On The Culture System Of Scutellaria Baicalensis Georgi Hairy Roots Ⅱ

Scutellaria baicalensis Georgi is one kind of traditional Chinese medicines, a herb of the Labiatae genus, widely distributes in Hebei, Liaoning, Shaanxi, Shandong and Inner Mongolia, and the medical functions of it are cleaning the heat and humidity, detoxifying, stanching and soothing fetus. The modern pharmacological studies indicated that baicalin, the major functional component of S.baicalensis, has abilities of anti-tumor and anti-human immunodeficiency virus (HIV). It also has potential application value to develop baicalin to be new drugs, but for the reason of slow growth of S.baicalensis, a perennial herb, which limites development of the products made from S.baicalensis. One effective way to solve the problem of sustainable development is to establish a culture system for culturing S.baicalensis hairy roots based on some modern biotechnologies.In this research, the culture conditions, baicalin accumulation pattern, amplification culture and determination of baicalin by high performance liquid chromatography (HPLC) were mainly studied. The results are as follows:1. The determination of baicalin in hairy roots by HPLC was performed on SciC18 column with a mobile phase of methanol water (50:50), contains 0.2% phosphate. The flow rate is 1 mL/min, the temperature is room temperature and the detective wave length is 274 nm. The retention time of baicalin is 9 min and the good linearity in the range of 12~72μg/mL (r=0.9992) is got under those conditions, and the average recovery is 99.32% with RSD of 1.36%.2. Effects of different carbohydrates on hairy roots growth and baicalin synthesis were investigated in this study. The results showed that the growth curve all turn on an"s"appearance using sucrose, glucose and fructose as carbon sourse respectively. Sucrose is benefit to the growth of hairy roots and in the case the highest dry weight reaches to 13.1g/L and baicalin content is 8.16%. Fructose makes the accumulation of baicalin to 10.66% with the dry weight of 11.4 g/L.3. Temperature is an important factor affecting the hairy root culture. At 28℃, the highest dry weight is 12.80g/L, while at 20℃the maximum baicalin content is 8.80%. The culture temperature giving the maximum baicalin content is not consistent with that giving the maximum biomass, so the method of two-step culture with different temperature was taken, and the dry weight and baicalin content are 13.3g/L and 12.58% and the yield of baicalin achieves 1.62g/L which increases by 66.34% compared to the conference (28℃).4. The effects of elicitors, hormones, precursor and replenish medium on the production of baicalin were studied. The results show that the yield of baicalin increases by 9.18%, 10.20% and 13.27% respectively, while concentrations of elicitors of Asp.niger, Asp.oryzae and Co2+ are 40mg/L, 20mg/L and 15.3×10-5 mmol/L respectively. The yield of baicalin increases by 25.74% and 24.75%, while concentrations of NAA and 6-BA are 0.4mg/L and 0.2 mg/L. The baicalin yield increases by 36.63% is achieved, when 0.4mmol/L phenylanine is added. The baicalin yield increases by 56.44% on 70 days after the medium was replenished.5. The research of amplification culture shows that the hairy roots of S.baicalensis grew rapidly and accumulated baicalin efficiently in air-lift reactor. The baicalin production decreased 22.41% compared to rotational culture implying the culture style influences the accumulation of baicalin in some extent.The culture system of hairy root of S.baicalensis was established by studying different factors affecting on biomass and baicalin accumulation, which builds the theoretical foundation for production of baicalin by hairy roots of S.baicalensis in a large-scale.