CHARACTERIZATION AND PRODUCTION OF MONO-RHAMNOLIPID AND DI-RHAMNOLIPID BIOSURFACTANTS IN RECOMBINANT ESCHERICHIA COLI

• Main Author: Hadi Kusuma, Subhan
• Format: Theses
• Language: Indonesia
• Subjects: Teknik (Rekayasa, enjinering dan kegiatan berkaitan)
• Online Access: https://digilib.itb.ac.id/gdl/view/47760
• Institution: Institut Teknologi Bandung

Surfactants are amphiphilic molecules and have an important role in various fields including agriculture, cosmetics, pharmaceuticals, bioremediation, and the petroleum industry. In the application of Enhanced Oil Recovery (EOR), surfactants are needed in large quantities, so, it is necessary to produce biodegradable surfactants such as rhamnolipid biosurfactants. Rhamnolipid is a glycolipid biosurfactant produced by Pseudomonas aeuruginosa. This species is a pathogen, so it is needed strategy to overcome this problem. Therefore, this research aims to do transforming the rhamnolipid gene into Escherichia coli for large-scale production. Rhamnolipid biosynthesis encoded by three main genes, rhlA, rhlB, and rhlC. The rhlAB produces mono-rhamnolipid and rhlABC produces di-rhamnolipid. The construction involved two plasmids, pPM RHLAB (mono-rhamnolipid) and pPM RHLABC (di-rhamnolipid) with T7lac promoter. Characterization of rhamnolipid biosurfactants by E24, IFT, and CMC analysis showed that di-rhamnolipid has better activity (70%, 0.8 mN/m, and 300 mg / L) than mono-rhamnolipid (58%, 2.1 mN/m and 900 mg / L) and SDS (46%, 4.7 mN/m, and 2000 mg / L) at pH 7, 25 oC, and 0% salinity. HRMS analysis shows that E. coli pPM RHLAB produces mono-rhamnolipid Rha-C14:2 (24%) and E. coli pPM RHLABC produces di-rhamno mono-lipid Rha-Rha-C10 (99.5%) with 8 novel rhamnolipid congeners. Production of rhamnolipids from E. coli pPM RHLABC using autoinduction media produced rhamnolipid concentrations of 1245.68 mg/L under optimal production conditions (production time of 20 hours, temperature 37 oC, and pH 7). Optimization of media variations obtained that the autoinduction media produced highest concentrations of rhamnolipid (1245.68 mg/L), LB+1% lactose (459.51 mg/L), M9+POME (Palm Oil Mill Effluent) 20% (318.42 mg/L), M9+POME 15% (305.59 mg/L), M9+POME 10% (211.58 mg/L), LB+POME 20% (193.48 mg/L), M9+POME 25% (180.40 mg/L), M9 (144.97 mg/L) and LB+0.5 mM IPTG (136.26 mg/L). The conclusion from this research indicate that di-rhamnolipid has better characteristics than mono-rhamnolipid for use as a surfactant in EOR technology, agriculture, and pharmacy.