Improvement Of Lovastatin Production By Fusarium Pseudocircinatum Ibrl B3-4 Via Solid Substrate Fermentation
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Date
2015-08
Authors
Ab Rashid, Syarifah
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Abstract
The ability of rice bran and brown rice in lowering cholesterol and supporting solid substrate fermentation (SSF) system are undeniable. Plus, the accessibility for these substrates is reachable at the mill factory with inexpensive price. For the last few years, researchers have investigated paddy and its other components like bran, rice, straw and husk, own the potentiality in producing secondary metabolite including statin. Malaysian local hospital has experienced a high demand on statin and lovastatin is the most outstanding drug with 51% prescription done for hypercholesterolemia patients. Thus, this recent investigation emphasized the improvement of lovastatin production by Fusarium pseudocircinatum IBRL B3-4 via SSF. Rice bran and brown rice denoted different composition values in moisture, protein, lipid, fibre, carbohydrate and ash. The highest component in rice bran was carbohydrate with 41.20 ± 2.10% while fibre was the superior composition in brown rice (48.53 ± 0.58%). Out of 78 filamentous fungi, only 55 isolates displayed positive dark spot on the thin layer chromatography plate (TLC) with retention factor (Rf) of 0.26 to 0.32. However, only 28 fungal isolates were detected to synthesise lovastatin through high performance liquid chromatography system (HPLC). IBRL B3-4 isolate depicted the highest lovastatin production with 281.67 ± 44.44 μg lovastatin/g dry solid. Based on the colony and structural morphologies on potato dextrose agar (PDA), the colony of IBRL B3-4 isolate was grown to 54 mm and 60 mm under dark and light conditions, respectively. For structural observation on carnation leaf agar (CLA) and Spezieller Nahrstoffarmer agar (SNA), the isolate owned oval shape microconidia (4.5-7.0 x 1.7-2.6 μm), slender and falcate shape macroconidia (32.0-46.4 x 1.2-2.9 μm), false head, coiled hyphae and also short chain structure. As a respond to the molecular approach, this isolate depicted 99.33% (Fusarium-ID database) and 99.00% (GenBank database) sequence similarity with F. pseudocircinatum. The best condition for F. pseudocircinatum IBRL B3-4 to produce lovastatin in the flask system was under the original substrate size, 70% (v/w) moisture content, incubation temperature of 30 ± 2°C, inoculum size of 1 x 105 spore/mL, pH 6.5, 5 g of substrate quantity (1:1 ratio) with static condition. It also needed external nutrients namely 1.5% (w/w) sucrose, 1% (w/w) yeast extract and 0.5% (w/w) calcium chloride. The tray system also required almost the same conditions with the flask system except for moisture content (60%; v/w) and substrate quantity (100 g or 0.5 cm thickness). The final productivity detected from flask system was 1770.00 ± 60.00 μg lovastatin/g dry solid while in tray system was 2436.67 ± 15.56 μg lovastatin/g dry solid, which represented 38% increment. Both systems required 12 days incubation period to synthesis the maximal productivity. During mycotoxin investigation, F. pseudocircinatum IBRL B3-4 has synthesized moniliformin (MON) and fumonisin B1 (FUM B1) at 4.20 ± 1.12 μg MON/g substrate and 1.73 ± 0.71 μg FUM/g substrate, respectively. However, the lethality concentration 50% values (LC50) for fractional lovastatin signified non-toxic activities as the value was higher than 1 mg/mL. Within 4 weeks treatment, fractional lovastatin concentration of 110 mg fraction sample/kg body weight (which represented 550 to 750 μg lovastatin/g dry solid of lovastatin) was slightly the best dose to reduce the low density lipoprotein (LDL) and increase the high density lipoprotein (HDL) in male and female Sprague dawley rats. The final LDL value detected via Thermo Scientific Multiskan® Spectrum microplate readers in male and female was 0.006 ± 0.001 μg/μl and 0.004 ± 0.001 μg/μl, respectively. While for the HDL, it amplified to 0.023 ± 0.001 μg/μl (in male) and 0.022 ± 0.003 μg/μl (in female). The results obtained from this work suggested the application of fractional lovastatin from F. pseudocircinatum IBRL B3-4 can greatly appointed this compound as an anti cholesterol lowering agent.
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