Application of Response Surface Methodology for Production of Capsaicin from Capsicum Annum L. L. A. Sagwan*, S. A. Survase and U. S. Annapure** Food Engineering and Technology Department, Institute of Chemical Technology, University of Mumbai, Matunga, Mumbai 400 019, India ABSTRACT Plant cell cultures are an attractive alternative source to whole plant for the production of high-value secondary metabolites. Taking advantage of this an optimization of different concentration of plant growth regulator was screened to obtain maximum callus growth and capsaicin production. A Statistical Design (Response Surface Methodology) was used in media optimization of chilli callus suspension culture. An increase in capsaicin yield from 29.89 μg/g dry weight cell mass to 36.32 μg/g dry weight cell mass was observed. Specific growth rate, Doubling time and Capsaicin production were higher for 25 g FW/L Inoculum size. Thus RSM was successfully used for increasing secondary metabolite production in suspension cell cultures. Keywords: Capsaicin, Response Surface Methodology, Plant Tissue Culture, Capsicum Annum, Secondary Metabolite, Suspension Cell Culture, Chilli Callus Capsaicinoids. *Corresponding author: *laxmi.sagwan@gmail.com; **usa@udct. org; INTERNATIONAL JOURNAL OF BIOTECHNOLOGY RESEARCH • January-June 2011 • Volume 4 • No. 1, pp. 11-18 IJBR  International Science Press Many higher plants are major sources of natural products used as pharmaceutical, agrochemicals, flavor and fragrance ingredients, food additives, and pesticides (Balandrin and Klocke, 1988). These belong to a group collectively known as secondary products or secondary metabolites. Research in the area of plant tissue culture technology has resulted in the production of many pharmaceutical substances for new therapeutics such as alkaloids, terpenoids, steroids, saponins, phenolics, flavanoids, and amino acids. The Phenolic compounds characteristic to fruits of the genus Capsicum, are commonly called capsaicinoids. Capsaicin and dihydrocapsaicin, are the most abundant pungent principles in hot peppers, and are responsible for 90% of the total pungency (Kosuge and Furata, 1970). Capsaicinoids are derivatives of phenylopropanoid compounds (Sudhakar Johnson et al., 1992). They are characterized by a high biological activity and their pharmacological, neurological and dietic effectiveness is well known. For effective capsaicin production, it is highly essential to optimize all the culture conditions and composition of medium. However, practically to optimize all the parameters and to establish the best possible conditions by interrelating all the parameters, numerous experiments have to be carried out which is not economical and practical. The traditional one-factor at a time approach to optimization is time-consuming and incapable of reaching the true optimum especially due to interaction among factors. Moreover, it assumes that the various growth parameters do not interact and that the process response is a direct function of the single varied parameter. Unlike conventional optimization, statistical optimization methods such as response surface methodology can take into account the interaction of variables in generating the process response (Haaland, 1989). Response surface methodology is a collection of mathematical and statistical techniques useful for analyzing the effects of several independent variables (Box and Hunter, 1978). RSM can be used to identify the effect of individual variables, evaluate the relative significance and seek the optimum constituents (Adinarayana et al. 2003; Elibol, 2004). These methods are often employed after a ‘‘vital few’’ controllable factors are identified and you want to find the factor settings that optimize the response. 1. MATERIALS AND METHODS Materials Kinetin, 2, 4-Dichlorophenoxy acetic acid, agar, di-sodium EDTA were procured from HIMEDIA Laboratories Pvt, Ltd. Mumbai. CaCl 2 .2H 2 O, MgSO 4 .7H 2 O, (NH 4 ) 2 SO 4 , Ca(NO 3 ) 2 .4H 2 O, NaH 2 PO 4 .H 2 O, MnSO 4 .H 2 O, CuSO 4 .5H 2 O, Fe 2 SO 4 .7H 2 O, ZnSO4.7H 2 O, KI, KH 2 PO 4 , MgSO 4 .7H 2 O, FeCl 3 .6H 2 0, inositol, sucrose, glucose, nicotinic acid, pyridoxine, glycine thiamine, ethanol, dry acetone, phosphomolybdic acid, Mercuric acid were procured from S. D. Fine chemicals Ltd. Mumbai. KNO 3 , HCl, CoCl 2 .6H 2 O, NH 4 NO 3 ,H 3 BO 3 , Na 2 MoO 4 .2H 2 O, Na 2 SO 4 , sodium potassium tartarate, ammonium molybdate, ascorbic acid, sodium hypochlorite,