Partial Purification Of Lipase Research Articles

Thermoseparating aqueous two-phase system for lipase recovery and partitioning from Nile tilapia viscera: Biochemical properties and effect of ultrasound

Lipase from Nile tilapia (Oreochromis niloticus) viscera were partitioned and recovered using a thermoseparating aqueous two-phase system (T-ATPS). Different phase partitioning parameters, including type and concentration of salts, concentration of EOPO, NaCl addition, the EOPO phase/distilled water ratio and temperature were optimized. In the primary ATPS, lipase was satisfactorily partitioned to the EOPO-rich top phase in the optimum system composed of 20% crude enzyme, 40% EOPO 3900, 10% (NH4)2SO4 and 4% NaCl. In the secondary ATPS, the optimum ratio between the EOPO-rich top phase and distilled water was 1:1 (w/w) and the optimum temperature for inducing phase separation was 60 °C. After thermoseparation, water solution containing lipase and EOPO were formed in the top and bottom phase of this step, respectively. Under the optimal partitioning condition, the yield (64.45%) and purification fold (PF: 6.30-fold) were obtained. Additionally, the lipases retained in the salt-rich bottom phase in the primary ATPS were successfully recovered from the recycling step no more than three times. The total yield of 93.59% was obtained from this separation system. The partitioned enzyme exhibited optimal activity at pH 8.5 and 40 °C and was stable at a temperature range of 0–40 °C and a pH range of 8–10. It showed high tolerance in the presence of ethanol and methanol. The effect of ultrasound on the partitioned lipase activity was also studied. The highest lipase activity was achieved when the sample was treated with ultrasound at 180 W and 24 kHz for 20 min at 40 °C, under which the activity was increased by 110.73% over the control. Therefore, the T-ATPS was found to be an attractive technique for the recovery and partial purification of lipase from Nile tilapia viscera. Also, ultrasound could obviously improve its activity.

Use of TPP and ATPS for partitioning and recovery of lipase from Pacific white shrimp (Litopenaeus vannamei) hepatopancreas.

Lipase recovery from Pacific white shrimp hepatopancreas using a three-phase partitioning (TPP) system in combination with an aqueous two-phase system (ATPS) was studied. TPP system was formed with a simultaneous addition of salt directly to crude extract (CE) followed by an organic solvent addition. The various process parameters required for efficient purification of lipase were optimized. The best lipase yield (87.41%) and purification fold (PF) (3.49-fold) were obtained in the interphase of TPP system, which consisted of the CE to t-butanol ratio of 1:1 (v/v) in the presence of 50% (w/v) (NH4)2SO4. Subsequently, TPP fraction was subjected to ATPS. Effects of phase compositions including PEG molecular weight and concentration, types and concentration of salts, NaCl addition and system pH on lipase partitioning were investigated. With the application of 25% (w/w) PEG1000 and 15% (w/w) MgSO4, at pH 5.0 was found most appropriate since high lipase PF (5.19-fold) and yield (78.46%) in top phase were obtained. The partitioned enzyme exhibited optimal activity at pH 8.0 and 55°C and was stable at a temperature range of 0-40°C and a pH range of 7-10. The partitioned lipase showed high tolerance in the presence of ethanol and methanol. Hence, the combined partitioning systems, TPP-ATPS, were found to be an attractive technique for the recovery and partial purification of lipase from Pacific white shrimp hepatopancreas.

Evaluation of partial purification and immobilization of lipase from Geotrichum candidum

Abstract Partial purification of lipase from Geotrichum candidum was obtained by precipitation using two methods: ethanol (70% v/v) and ammonium sulphate (80% w/v) followed by lyophylization. Recovery factors (RF) from 60% to 140%, concentration factors (CF) from 44 to 66 and lipase activities from 400 to 900 U g − 1 were obtained. Lyophylization is important to maintain the enzymatic activity. In relation to solvent affinity, lyophilized lipase had higher affinity for the polar phase when incubated in phosphate buffer:hexane (1:1) but it had lower stability when incubated in phosphate buffer:ethanol (1:1) at 37 °C. Immobilization onto different supports (alginate beads, niobium–graphite alloy, zeolite and Celite®) revealed the inability of niobium–graphite alloy as a support and good potential for the other supports. Under the circumstances applied, Celite® resulted in an immobilized lipase activity around 90 U g −1 and zeolite presented efficiency around 66%, which indicate to be good alternatives for immobilization of lipase from G.candidum .

Production, optimization, and partial purification of lipase fromPseudomonassp. strainBUP6, a novel rumen bacterium characterized fromMalabari goat

This study introduces a novel bacterium-Pseudomonas sp. strain BUP6-isolated from the rumen of the Malabari goat with efficiency for producing lipase. It showed significant production of lipase when grown in a newly designed basal medium, supplemented with vegetable oil. Suitability of five vegetable oils such as groundnut oil, coconut oil, olive oil, sunflower oil, and palm oil as inducer for the production of lipase was examined, and groundnut oil supported the highest production of lipase (96.15 U/mL). Various physical parameters required for the maximum production of lipase were statistically optimized. Plackett-Burmann design was employed to study the interactive effects of physical parameters and found that temperature, agitation, and pH effected the production of lipase significantly. The optimum conditions for lipase production (37°C, 200rpm, and pH 6.9) were detected by Box-Behnken design and response surface methodology, which resulted in the 0.3-fold increase (i.e., 126U/mL) of the lipase activity over the unoptimized condition. The apparent molecular mass of partially purified lipase was 35kDa, as judged by SDS-PAGE; the activity of lipase was also confirmed by native PAGE. Thus, this study focuses on the need for the exploitation of rumen microbes for the production of industrially significant and human-friendly biomolecules to meet the future needs.

Partial purification of lipase from cotyledons of oilseed rape (Brassica napus L.)

Partial purification of lipase from cotyledons of oilseed rape (<i>Brassica napus</i> L.)