Improved Taxol production by combination of inducing factors in suspension cell culture of Taxus baccata

Abstract

To date enormous attempts have been devoted to improve Taxol production exploiting various methodologies from bioprocess engineering to biotechnological and synthetic approaches. We have developed a 2-stage suspension cell culture of Taxus baccata L. using modified B5 medium in order to improve cell growth as well as productivity. After callus induction and cell line selection, B5 medium was supplemented with vanadyl sulfate (0.1 mg/l), silver nitrate (0.3 mg/l) and cobalt chloride (0.25 mg/l) at the first day of stage I culture to maximize cell growth. This medium was further supplemented with sucrose (1%) and ammonium citrate (50 mg/l) on day 10 and sucrose (1%) and phenylalanine (0.1 mM) on day 20 (i.e., biomass growth medium). At stage II (day 25), two different concentrations of several elicitors such as methyl jasmonate (10 or 20 mg/l), salicylic acid (50 or 100 mg/l) and fungal elicitor (25 or 50 mg/l) were added to the biomass growth medium with the aim of improving cellular productivity. For morphological analysis, microscopic inspection was carried out during cultivation. Cell-associated and extracellular amount of Taxol were detected and measured using HPLC methodology. At stage I, overall Taxol amount of biomass growth medium was 13.75 mg/l (i.e., 5.6-fold higher than that of untreated B5 control). At stage II, treated cells with methyl jasmonate (10 mg/l), salicylic acid (100 mg/l) and fungal elicitor (25 mg/l) produced the highest amount of Taxol (39.5 mg/l), which is 16-fold higher than that of untreated B5 control (2.45 mg/l). Microscopic analyses of Taxus cells in suspension cultures showed various positional auto-fluorescence showing direct correlation with Taxol production. Our studies revealed that intervallic supplementation of B5 medium with combination of biomass growth factors at stage I and mixture of elicitors at stage II could significantly increase Taxol production. Thus, we suggest that the exploitation of this methodology may improve the production of Taxol since demands for Taxol pharmaceuticals are increasingly growing and resource paucities have limited its direct harvesting from Taxus trees.

Endophytes: exploiting biodiversity for the improvement of natural product-based drug discovery

“…Ketchum and Gibson 1996;Ketchum and Croteau 1998;Yukimune et al 2000;Wu and Lin 2003;Naill and Roberts 2005;Khosroushahi et al 2006;Vongpaseuth and Roberts 2007). This conclusion is further supported by the aforementioned work of Croteau and his associates who succeeded in the isolation of paclitaxel biosynthetic genes of plant origin.…”

New Approach To Improve Taxol Biosynthetic

“…So, Taxol and baccatin III levels increase 4-and 3.6-fold, over the controls in the presence MJ but Vanadyl sulphate only enhance 4-fold taxol production [53]. observed that Taxol remarkably is increased by use of these elicitors more than 16-fold as compared with untreated B5 media [45].…”

“…Cell suspensions suffer from low yeild of production of secondary metabolites and somaclonal variation. This pronlems can be reduced by careful selection of favorite calli, optimal growth conditions, medium and adding some elicitors [45]. Somaclonal variation can be positive or negative [46]; it is possible to choose desirable callus with high Taxol content.…”

“…Selection of high yielding cell lines, optimization of culture conditions, culture media components such as Sugar, phosphate, nitrate and plant growth regulators, carbohydrate source, explant, physical conditions, immobilization, phytohormones, elicitors, addition of precursors, adsorbants or additives, nutrient media and employment of a two-stage culture system are important factors can play a significant role to increase the growth and high accumulation of secondary metabolites [39,45].…”

“…These fast growing systems can be used for large scale culture of plant cells to obtain valuable products. Various strategies are being employed in continuing efforts to increase productivity [7,45]. Cell suspensions suffer from low yeild of production of secondary metabolites and somaclonal variation.…”