| Summary: | 碩士 === 中國文化大學 === 生物科技研究所 === 87 === This study is aimed to optimized the culture conditions for the growth of cell derived from Ginkgo biloba and ginkgolide（GK）production.The callus was induced from the cotyledon, root,stem and leaf in the MS media supplemented with various growth regulators.The best conditions for callus induction and cultivation were at 25℃ in vessel container with light intensity of 2200-2300 lux and light period of 12 hr/day.The cell growth index（final wt./original wt.）was 2.5 after 14-day cultivation.
The optimal inoculum ratios for agar-to-liquid and liquid -to-liquid transfers were 4﹪（w/v）and 2.2﹪w/v）,respectively. The optimal medium for suspension culture was MS added with 1 ppm NAA and 0.1 ppm kinetin,with pH 6.0 and adequate ventilation. The light intensity did not affect the cell growth.Cell culture in rolling bottle with rotation rate of 1.5 rpm and adequate ventilation got the best results.The cell growth indexes（final PCV/original PCV）for shake flask and rolling bottle cultures were 4-5 and 5-7,respectively.The dry weight to fresh weight ratio of the cell from both cultures was 7-8﹪
Two types of bioreactor were utilized for cell mass production,including air-driver temporary immersion system （ADTIS）and rotary drum bioreactor （RDB）.In ADTIS,the optimal immersion to air-exposure time periods were 15/15,15/30 and 30/30 mins.They had the growth index of 3-4 after 14-day cultivation.In RDB,the growth indexes were 3.2 and 4.2 for 14-day and 28-day cultivation,respectively.The dry weight to fresh weight ratio of cell from both culture was 10-11﹪.
In RDB culture,the medium pH dropped from 6.0 to 4.9 at the day 2 and the remain 4.9 until the end of culture.Dissolved oxygen dropped gradually from 6.8 mg/L to 4.5 mg/L,after 28-day cultivation. In nutrient consumption of RDB culture,carbon sours was almost completely consumed,while 70-80﹪of the nitrogen and phosphate source were still left in the media at the end of culture.
Addition of banana meal at 10 or 50 g/l in solid media or 10 g/l in liquid media could promote the cell growth of ginkgolide. The coconut milk,however,did not improve the cell growth .
The GKA and GKB production rates were better in (1)stem cell in rolling bottle,(2)cotyledon cell in rolling bottle and(3) cotyledon cell in darks suspension culture as compared with other different type cultures.The highest ginkgolide content was 23×10-5﹪（of dry weight）.The cell growth rates,however,were better in(1)stem cell in rolling bottle,(2)cotyledon cell rolling bottle,and (3)leaf cell in liquid suspension culture.
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