Farooq Komal, Anwar Zahid, Khalid Waseem, Hasan Shoaib, Afzal Fareed, Zafar Muddassar, Ali Usman, Alghamdi Othman, Al-Farga Ammar, Al-Maaqar Saleh M
Department of Biochemistry and Biotechnology, University of Gujrat, Gujrat 54000, Pakistan.
University Institute of Food Science and Technology, The University of Lahore, Lahore 54000, Pakistan.
ACS Omega. 2024 Apr 4;9(15):17446-17457. doi: 10.1021/acsomega.4c00274. eCollection 2024 Apr 16.
The biotechnological process called solid-state fermentation (SSF) was applied for hyper production of protease by using a fungal strain called . From screening of 9 different local substrates (peanut shell, wheat bran, guava leaves, sugar cane bagasse, rice polish, wheat straw, corn straw, reed grass, and rice straw), peanut shells serve as the best substrates for protease production under optimized cultured conditions. The varying physiochemical parameters such as pH (2-9.5), temperature (30-52 °C), incubation time (1-10 days), inoculum size (1-8 mL), moisture level (20-125%), and substrate concentration (1-7 g) were optimized by response surface methodology (RSM). The highest activity of protease was recorded to be 1101.778 U/mL at 660 nm using peanut shell was optimum at pH 8, temperature 52 °C, incubation time 8 days, inoculum size 2 mL, moisture level 20%, and substrate concentration 2 g. The crude form of enzymes produced were further purified through ammonium sulfate precipitation, dialysis, and gel filtration chromatography. Then, purified enzymes were characterized at different pH, temperature, and incubation time. For characterization of purified protease, pH, temperature, and incubation time were 8, 52 °C, and 8 days for peanut shell and was done by one factor at a time method. Hence, isolated enzymes were alkaline in nature, i.e., alkaline proteases. Then, protease produced from peanut shells was applied to locally available detergents to increase their catalytic activity for strain removal. At last, the final results were interpreted in the form of 3D surface and contour plots using Microsoft Excel 2013 and Minitab 17 software. In conclusion, the utilization of and peanut shell as the substrate in the biotechnological process of SSF demonstrated successful hyper production of alkaline protease. The optimized conditions resulted in high enzyme activity and showcased the potential application of the isolated enzymes in improving the catalytic activity of locally available detergents.
名为固态发酵（SSF）的生物技术过程被用于通过一种名为 的真菌菌株超量生产蛋白酶。从对9种不同的本地底物（花生壳、麦麸、番石榴叶、甘蔗渣、米糠、小麦秸秆、玉米秸秆、芦苇草和稻草）的筛选中可知，在优化的培养条件下，花生壳是蛋白酶生产的最佳底物。通过响应面法（RSM）对不同的物理化学参数进行了优化，如pH值（2 - 9.5）、温度（30 - 52℃）、培养时间（1 - 10天）、接种量（1 - 8 mL）、水分含量（20 - 125%）和底物浓度（1 - 7 g）。使用花生壳时，在pH值为8、温度为52℃、培养时间为8天、接种量为2 mL、水分含量为20%、底物浓度为2 g的条件下，在660 nm处测得的蛋白酶最高活性为1101.778 U/mL。所产生的粗酶形式通过硫酸铵沉淀、透析和凝胶过滤色谱进一步纯化。然后，在不同的pH值、温度和培养时间下对纯化后的酶进行表征。对于纯化蛋白酶的表征，花生壳的pH值、温度和培养时间分别为8、52℃和8天，采用一次一个因素的方法进行。因此，分离出的酶本质上是碱性的，即碱性蛋白酶。然后，将从花生壳中产生的蛋白酶应用于当地可得的洗涤剂中，以提高它们去除污渍的催化活性。最后，使用Microsoft Excel 2013和Minitab 17软件以三维表面图和等高线图的形式解释最终结果。总之，在SSF生物技术过程中利用 和花生壳作为底物成功地超量生产了碱性蛋白酶。优化后的条件导致了高酶活性，并展示了分离出的酶在提高当地可得洗涤剂催化活性方面的潜在应用。
