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Use of Potato Peel Waste- a Cost Effective Alternative Substrate for the Production of Industrially Useful α-amylase Enzyme

F. Islam
Centre for Advanced Research in Sciences, University of Dhaka, Dhaka 1000, Bangladesh & Department of Microbiology, Jagannath University, Dhaka, Bangladesh

M. A. A. Mamun
Centre for Advanced Research in Sciences, University of Dhaka, Dhaka 1000, Bangladesh

A. K. Azad
Department of Microbiology, Jagannath University, Dhaka, Bangladesh

S. Yamashoji
Microbial Technology Laboratory, Kobe, Japan

M. L. Bari
Centre for Advanced Research in Sciences, University of Dhaka, Dhaka 1000, Bangladesh

Abstract/ Executive Summary:

Potato (Solanum tuberosum L.) is the fourth main crop worldwide for human consumption and widely used in food-processing industries, thus generating extremely large amounts of potato peel waste (PPW) as zero value by-product. This study was designed to utilize this PPW as biomaterials for the production of alpha-amylase by submerged fermentation using Aspergillus oryzae, isolated from commercial white koji. The study results demonstrated that the maximum amylase production (30 U/ml) was achieved in PPW medium supplemented with yeast extract and salt after 72 hrs of incubation at ambient temperature (28 ± 2°C), pH 7.0 and 120 rpm agitation. Extraction of the whole amylase from culture filtrate was found suitable with 80% ammonium sulfate saturation and maximum amylase activity of 42.07 U/ml was achieved after desalting. The purified α-amylase was found optimally active and stable up to 50°C.

Keywords: α-amylase, Aspergillus oryzae, Potato peel waste, Submerged fermentation. Enhanced nutritional composition.

Location: Local market in Dhaka city

Start Date:

End Date:

Program Area: Socio-economic and Policy

Non-commodity: Profitability

Objectives:

The main objective of this study was to produce industrially important amylase enzyme by submerged fermentation using Aspergillus oryzae and potato peel waste.

Methodology:

The fungal strain Aspergillus oryzae was isolated from commercial white koji preparation and stored on potato dextrose agar (PDA) plates at 4°C and used for this study. Substrate preparation- The potatoes were bought from the local market in Dhaka city, washed with tap water and boiled in an aluminum pot. Then peels were taken from boiled potatoes, washed and allowed to dry in laminar airflow for 1 h, and used as a substrate for amylase production. In addition, various parboiled rice (2%) including SonaGuti, GutiSharna, Najirshal, IRRI 28, and IRRI 29 produced in Bangladesh were also used as a substrate for amylase production. Submerged fermentation (SmF)- Fermentation medium (pH 7.0) was composed of 100 ml water, 2.0 g potato peel, 0.5 g yeast extract, 0.1 g K₂HPO₄, 0.05 g MgSO₄, and 0.001 g FeSO₄/7H₂O, and taken in 500 ml Erlenmeyer flask, and autoclaved at 121°C for 15 min at 15 lbs pressure. After autoclaving, the medium was cooled to room temperature and 1.0 cm×1.0 cm of PDA medium containing Aspergillus oryzae culture was added to the fermentation medium. The flask was then placed in a shaking incubator (120 rpm) at ambient temperature (28 ± 2°C) for 72 hrs. Optimization of process parameters- Various process parameters were optimized for maximal enzyme production as follows: incubation period (1-7 days), initial pH (6.5-8.5), substrate concentration (1%, 2% and 3%). Recovery of enzyme- After the specified incubation period (in each case), the medium was centrifuged at 6,000 rpm for 10 min, the supernatant was taken and used as a crude enzyme to measure α-amylase activity. Amylase assay-α-amylase was assayed by estimating liberated reducing sugars employing the 3,5-dinitrosalicylic acid (DNS) method (Miller, 1959). The reaction mixture containing 0.5 ml enzyme extract and 1.0 ml soluble starch solution was incubated for 20 min at 40°C. The reaction was stopped by adding a 3.0 ml DNS reagent. Then the mixture was heated in a boiling water bath (90°C) for 15 minutes. After boiling, test tubes containing the reaction mixture were cooled under running tap water. Absorbance was taken at 545 nm against the blank in a spectrophotometer. The liberated reducing sugars were estimated by dinitrosalicylic acid (DNS) method and taking glucose as standard (Miller, 1959). The blank contained 0.5 ml distilled water instead of 0.5 ml enzyme extract. Purification of α-amylase- Precipitation of the α-amylase was performed by adding ammonium sulfate in the enzyme solution following the method described by Wingfield (Wingfield, 2016). Solid (NH₄)₂SO₄ was added gradually into every 50 ml enzyme contained in conical flask in order to make it 50, 70, 80, 90 percent saturation and left overnight at 4°C. Then the precipitates were collected by centrifugation at 10,000 rpm for 10 min. The precipitates were dissolved in 5 ml of 0.2M phosphate buffer solution (pH 7). Desalting of dissolved precipitates was done by gel filtration using Sephadex G-25. The column was prepared by adding 20 ml of Tris-HCl buffer (pH 7.5) and 1.0 g of Sephadex G-25. One ml of sample was added slowly into the column. Then 20 ml elution buffer was added into the column and fractions (1.0 ml each) were collected. The enzyme activity and protein concentration (Bradford, 1976) were then assayed with appropriate dilution. Statistical analysis- All trials were replicated three times and the data presented in the table and figures were the average value ± standard deviation. The significant differences were subjected to analysis of variances using the Microsoft Excel program (Redmond, Washington DC, USA.).

Source of Information: Journal of Agriculture, Food and Environment, Vol 1 No 4 December 2020 Pages 133-137

Article Link (Online Journal): http://doi.org/10.47440/JAFE.2020.1420

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

In this study, 2% potato peel at pH 7.0 after 72 hrs of submerged fermentation at ambient temperature using A. oryzae, showed higher amylase production than that of different types of rice. Thus, this study results concluded that potato peel waste could be a cost-effective alternative substrate for the production of the industrially useful α-amylase enzyme. Further studies on the analysis of the nutritional composition of the fermented potato peel will be done to evaluate whether the fermented potato peel possesses enhanced nutritional composition and could be useful as nutrient-rich animal feed.

Knowledge Management: Journal