Current Research Journal of Biological Sciences 5(6): 273-284, 2013 ISSN: 2041-076X, e-ISSN: 2041-0778 © Maxwell Scientific Organization, 2013 Submitted: July 9, 2013 Accepted: July 24, 2013 Published: November 20, 2013 273 Production of Poly-β-Hydroxybutyric Acid (PHB) by Rhizobium elti and Pseudomonas stutzeri Elsayed B. Belal Department of Agricultural Botany, (Agricultural Microbiology), Faculty of Agriculture, Kafrelsheikh University, 33516, Kafr El-Sheikh, Egypt, Tel./Fax: 0020479102930 Abstract: The amount of chemosynthetic plastic waste increases every year and exact time for its degradation is unknown. Two poly-β-hydroxybutyrate producing bacteria from different microbial sources were isolated and characterized for their morphological, biochemical properties. Based on their 16S rDNA, they were identified as Rhizobium elti E1 and Pseudomonas stutzeri E114. The bacterial strains were screened for PHB production and compared for the intensity of fluorescence using Nile red staining methods. PHB production conditions were optimized with different carbon and nitrogen sources, the highest PHB production was observed with mannitol, sucrose and ammonium sulphate by R. elti and P. stutzeri, respectively. Regarding incubation time as well as temperature and pH, optimum PHB production conditions were 48 h, 30°C and 7, respectively. R. elti and P. stutzeri are capable of accumulating appreciable levels of PHB from glucose, xylose, lactose, whey, molasses, sugar cane bagasse, rice straw hydroysate when 2% from all substrates were used an alternative carbon for the PHB production. Ammonium sulphate was the best nitrogen source. C/N ratio was also one of the factors that affected the production of PHB. The ratio of C/N that reaches 20:1 was considered the best ratio to produce the highest production of PHB. The highest yield of PHB was done by P. stutzeri more than R. elti. The present study provide the useful data about the optimized conditions for PHB production by R. elti and P. stutzeri that can be utilized for industrial production of PHB, a fast emerging alternative of non biodegradable plastics. Keywords: PHB, Rhizobium elti, polyhydroxybutyrate, Pseudomonas stutzeri INTRODUCTION Synthetic polymers-designated as plastics-are applied in a wide range of packing-, household, agricultural, marine and architectural applications. Plastics were developed as light-weight and durable materials and they have replaced natural resources, such as metals and stones. However, its properties of durability have caused serious problems since plastic waste accumulates in the environment. The accumulation of abandoned plastics has caused a global environmental problem. Nature usually cannot handle plastic waste, since the majority of plastics are not degraded by microorganisms. At present, about one hundred million tons per year of plastics are produced in the world. With the increase in production, the amount of plastics wastes has raised enormously (Mukai and Doi, 1995) and increased the costs of solid waste disposal (owing to the reductions in available landfill space) dramatically. Alternatives to waste disposal such as plastic recycling are quite limited from an economical view point and partially include potential hazards (such as dioxine emission from PVC incineration) (Belal, 2003). Biodegradable plastics opened the way for new waste management strategies since these materials are designed to degrade under environmental conditions or in municipal and industrial biological waste treatment facilities. Most of the plastics on the market, claimed to be biodegradable, are based on synthetic and microbial polyesters (Augusta et al., 1993; Witt et al., 2001). Polyesters are potentially biodegradable due to the hydrozable ester bonds. In addition, they combine several properties that make them attractive candidates for various industrial applications. Among various biodegradable polymer materials are Polyhydroxyalkanoates (PHAs) (Page, 1992b; Zhang et al., 2003). This polymer family is made of two major groups-aliphatic and aromatic. Polyhydroxyalkanoate (PHAs) are alilphatic polymer naturally produced via a microbial process on sugar- based medium, where they act as carbon and energy storage material in bacteria. The main member of the PHA family is Polyhydroxybutyrate (PHB). PHB is accumulated inside in numerous bacteria under nutrient-limiting conditions with excess carbon. Many references show that number of microorganisms like Alcaligene eutrophus, Azotobacter beijierinckia, Pseudomonas oleovorans, Rhizobium sp., etc., produce PHAs as reserve food material. They can be used in different applications such as packaging film and containers, surgical pins and sutures and bone