Extraction, purification and characterisation of a milk-clotting protease from ‘kesinai’ (Streblus asper Lour.) leaves
Extracts from ‘kesinai’ (Streblus asper) leaves were investigated as a potential source of enzymes that can serve as an alternative to calf rennet in cheese making. Different types of extraction buffers were investigated namely sodium acetate buffer (pH 4.2-5.0), phosphate buffer (pH 6.0-7.0) and...
| Main Authors: | , , |
|---|---|
| Format: | Article |
| Published: |
Faculty of Food Science and Technology, Universiti Putra Malaysia
2019
|
| Online Access: | http://psasir.upm.edu.my/id/eprint/79840/ |
| _version_ | 1848858770543738880 |
|---|---|
| author | Pagthinathan, M. Ghazali, H. M. A. M., Yazid |
| author_facet | Pagthinathan, M. Ghazali, H. M. A. M., Yazid |
| author_sort | Pagthinathan, M. |
| building | UPM Institutional Repository |
| collection | Online Access |
| description | Extracts from ‘kesinai’ (Streblus asper) leaves were investigated as a potential source of enzymes
that can serve as an alternative to calf rennet in cheese making. Different types of extraction
buffers were investigated namely sodium acetate buffer (pH 4.2-5.0), phosphate buffer (pH
6.0-7.0) and Tris-HCl buffer (pH 7.0-9.0). Finally, the milk-clotting enzyme was extracted
using 100 mM Tris-HCl buffer (pH 7.4) with and without 5.0 mg/mL polyvinylpyrrolidone,
0.015 mL/mL Triton X-100 and 2 mM sodium metabisulphite. Purification was carried out
using acetone precipitation, and ion-exchange and size-exclusion chromatographic techniques.
Results showed that 100 mM Tris-HCl buffer (pH 7.4) was the most efficient extraction buffer
among the buffers used in the extraction study. After the final purification step of size-exclusion
chromatography, the enzyme was purified 3.3-fold with 42.3% of recovery. The enzyme
showed an optimum temperature and pH at 60°C and pH 7.4, respectively. The enzyme was
stable up to 70°C for one hour and the partially purified enzyme retained 83% and 96% of its
original activity at pH 6.0 and 8.0, respectively. The molecular weight of the partially enzyme
was estimated to be 75.8 kDa on SDS-PAGE. The milk-clotting activity of ‘kesinai’ enzyme
was found to be lower than that of commercial Mucor rennet. |
| first_indexed | 2025-11-15T12:18:44Z |
| format | Article |
| id | upm-79840 |
| institution | Universiti Putra Malaysia |
| institution_category | Local University |
| last_indexed | 2025-11-15T12:18:44Z |
| publishDate | 2019 |
| publisher | Faculty of Food Science and Technology, Universiti Putra Malaysia |
| recordtype | eprints |
| repository_type | Digital Repository |
| spelling | upm-798402022-11-14T04:35:58Z http://psasir.upm.edu.my/id/eprint/79840/ Extraction, purification and characterisation of a milk-clotting protease from ‘kesinai’ (Streblus asper Lour.) leaves Pagthinathan, M. Ghazali, H. M. A. M., Yazid Extracts from ‘kesinai’ (Streblus asper) leaves were investigated as a potential source of enzymes that can serve as an alternative to calf rennet in cheese making. Different types of extraction buffers were investigated namely sodium acetate buffer (pH 4.2-5.0), phosphate buffer (pH 6.0-7.0) and Tris-HCl buffer (pH 7.0-9.0). Finally, the milk-clotting enzyme was extracted using 100 mM Tris-HCl buffer (pH 7.4) with and without 5.0 mg/mL polyvinylpyrrolidone, 0.015 mL/mL Triton X-100 and 2 mM sodium metabisulphite. Purification was carried out using acetone precipitation, and ion-exchange and size-exclusion chromatographic techniques. Results showed that 100 mM Tris-HCl buffer (pH 7.4) was the most efficient extraction buffer among the buffers used in the extraction study. After the final purification step of size-exclusion chromatography, the enzyme was purified 3.3-fold with 42.3% of recovery. The enzyme showed an optimum temperature and pH at 60°C and pH 7.4, respectively. The enzyme was stable up to 70°C for one hour and the partially purified enzyme retained 83% and 96% of its original activity at pH 6.0 and 8.0, respectively. The molecular weight of the partially enzyme was estimated to be 75.8 kDa on SDS-PAGE. The milk-clotting activity of ‘kesinai’ enzyme was found to be lower than that of commercial Mucor rennet. Faculty of Food Science and Technology, Universiti Putra Malaysia 2019 Article PeerReviewed Pagthinathan, M. and Ghazali, H. M. and A. M., Yazid (2019) Extraction, purification and characterisation of a milk-clotting protease from ‘kesinai’ (Streblus asper Lour.) leaves. International Food Research Journal, 26 (3). pp. 913-922. ISSN 1985-4668; ESNN: 1505-5337 http://www.ifrj.upm.edu.my/26%20(03)%202019/21%20-%20IFRJ161395.R2-Final.pdf |
| spellingShingle | Pagthinathan, M. Ghazali, H. M. A. M., Yazid Extraction, purification and characterisation of a milk-clotting protease from ‘kesinai’ (Streblus asper Lour.) leaves |
| title | Extraction, purification and characterisation of a milk-clotting protease from ‘kesinai’ (Streblus asper Lour.) leaves |
| title_full | Extraction, purification and characterisation of a milk-clotting protease from ‘kesinai’ (Streblus asper Lour.) leaves |
| title_fullStr | Extraction, purification and characterisation of a milk-clotting protease from ‘kesinai’ (Streblus asper Lour.) leaves |
| title_full_unstemmed | Extraction, purification and characterisation of a milk-clotting protease from ‘kesinai’ (Streblus asper Lour.) leaves |
| title_short | Extraction, purification and characterisation of a milk-clotting protease from ‘kesinai’ (Streblus asper Lour.) leaves |
| title_sort | extraction, purification and characterisation of a milk-clotting protease from ‘kesinai’ (streblus asper lour.) leaves |
| url | http://psasir.upm.edu.my/id/eprint/79840/ http://psasir.upm.edu.my/id/eprint/79840/ |