Production and stabilization of microbial rennet from Rhizopus chinensis

Microbial rennet was initially produced from Rizophus chinensis by submerged fermentation using 30L stirred tank batch bioreactor with 4% wheat bran as substrate. The substrate was substituted with the less expensive coconut paring meal which was found to be a better substrate for fermentation. The optimized condition was able to shorten the fermentation condition to less than 24 hour and resulted to higher product yield coefficient of 1.7 liters per kilogram substrate. Aflatoxin assay of coconut paring meal and the resulting concentrated rennet gave negative results therefore safe to be used as food ingredient. Solid substrate production of microbial rennet was done due to malfunction of all the fermenter at BIOTECH pilot plant. Production using 12'X8'X 2.5 enamel tray loaded with 200 grams hydrated coconut paring meal was able to produce more rennet. The process had a product yield coefficient of 2.5 liters per kilogram substrate which is higher compared to submerged fermentation. Comparative study of the cost of production of microbial rennet using submerged and solid substrate fermentation showed that the later was scale up the production using trays compared to the use of the stirred tank reactor and the enzyme yield is higher. Granulation of liquid rennet was done by addition of binders and fillers to further stabilize the microbial rennet. The study on the production of high milk clotting enzyme (MCE) activity granule through the wet granulation of liquid rennet using the excipients cornstarch as filler and maltodextrin as binder was done. Large scale experiment showed consistent among trial runs which simply highly reproducible agglomeration mechanism during the granulation step. The granulated rennet with the highest activity was tried in preparation of white cheese at the rate of 2%, however, the corn starch from the granule settled at the bottom of the container. Rennet granules composed of gelatin, lactose and skim milk was developed. Result showed that the only concentration of the rennet and the drying temperature significantly affect the milk- clotting activity of the granules produced. The amount of gelatin was found not to affect the activity of the granules. Result also showed consistent Sauter means and average particle sizes between trial runs and a direct relationship occurred between particle size and the activity of the enzyme. The liquid rennet was pack in high density polyethylene bottle as per advise of the DOST Packaging Center while the granulated rennet was packed in foil packs. Evaluation of the microbial rennet which was conducted at the Department of Paraclinical Sciences, College of Veterinary Medicine, UPLB [University of the Philippines Los Baños] showed that the microbial rennet is safe to be used as food ingredient based on its toxicological and pathological studies using Swiss Webster Mice. The consumer preferences survey conducted showed that the microbial rennet has a better comparative advantage over animal rennet in terms of quality and is highly competitive with the imported rennet, in terms of the product attributes such as reliability, economy, ease of use and availability. Based on the analysis, production and distribution of BIOTECH rennet will generate an average net profit after tax of PhP 1.9 M every year with 42.7% average return of investment, while the net present value of income streams when discounted at 15% will be 1.92M pesos. The cost and benefit of using microbial rennet in white cheese making compared to other milk coagulants was done. Result showed that the use of the microbial rennet can increase the yield of cheese from cow's and carabao's milk by 28% and 22%, respectively compared to use of the animal rennet and ChyMax. Analysis of the by-product or biomass of microbial rennet fermentation showed an increase on protein and fiber content which suggested enrichment of the coconut paring meal substrate which can possibly be used as feed supplement for dairy animals. The washed biomass was acceptable to the young water buffalo when added to the fluid supplement which also contains honey. The study was suspended until enough material is produced because feeding trials required 40 to 50 kilograms per animal.