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Outline
Objective: To develop tannases resources with excellent enzymatic properties. Methods: Primary screening was performed using the clear zone plate method, followed by a secondary screening using the methanol rhodanine colorimetric assay. A fungal strain with the highest tannase activity was isolated from the Daqu samples. The strain was identified through morphological observation and ITS sequence analysis. The liquid fermentation conditions for tannase production were optimized. The enzyme (PbTanA) was purified through ammonium sulfate precipitation and DEAE weak anion-exchange chromatography, obtaining electrophoretic-grade pure enzyme. The enzymatic properties of PbTanA were further investigated, and its application in hydrolyzing tannic acid to produce gallic acid was evaluated. Results: The strain was identified as Penicillium bilaiae and named Penicillium bilaiae CAU46-1. After fermentation at 30 ℃ and pH5.5 for 4 days, a tannase activity of 28.7 U/mL was produced. This was achieved under conditions where 3% tannic acid served as inducer, 4% wheat bran as carbon source, and 0.2% ammonium chloride as nitrogen source, resulting in a tannase activity 3.3 times higher than the initial level. The tannase (PbTanA) was purified using ammonium sulfate precipitation followed by DEAE weak anion exchange chromatography. Its molecular weight was 79.0 kDa. The enzyme exhibited good acid tolerance and thermal stability, with an optimal pH of 5.0 and good stability within the pH range of 2.0~7.0. Its optimal temperature was 60 ℃, and it remained stable up to 50 ℃. PbTanA hydrolyzed 20 g/L tannic acid for 8 h, producing a gallic acid concentration of 18.4 g/L with a yield of 91.8%. Conclusion: Tannase PbTanA from Penicillium bilaiae CAU46-1 showed excellent enzymatic properties and efficiently hydrolyzed tannic acid to produce gallic acid, demonstrating its potential for application.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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