The esterase activity was determined in cheeses homogenate
using on a-naphthyl derivatives of fatty acids of 2–8 carbon atoms
as substrate (Sigma, St. Louis, MO) according to the method
describe by Medina et al. (2004). The assay mixture contained
160 mL of 100 mM sodium phosphate buffer, pH 7.0, 20 mL of anaphthyl
substrate (10 mM in ethanol), and 100 mL of cheese
homogenate. After incubation for 1 h at 37 8C, color was developed
by adding 0.6 mL of Fast Garnet GBC (Sigma) solution (5 mg/mL in
10% w/v SDS) and further incubation at room temperature for
15 min. The absorbance was measured at 560 nm in a spectrophotometer
(CECIL 2021, Cambridge, UK). Controls containing the
reaction mixture plus glacial acetic acid were also incubated to test
for the presence of background activity. A standard curve was
prepared using a-naphthol. A unit of esterase activity was defined
as the amount of enzyme that released 1 mmol of a-naphthol per
minute. Specific esterase activity was defined as units per
milligram of protein.
The esterase activity was determined in cheeses homogenateusing on a-naphthyl derivatives of fatty acids of 2–8 carbon atomsas substrate (Sigma, St. Louis, MO) according to the methoddescribe by Medina et al. (2004). The assay mixture contained160 mL of 100 mM sodium phosphate buffer, pH 7.0, 20 mL of anaphthylsubstrate (10 mM in ethanol), and 100 mL of cheesehomogenate. After incubation for 1 h at 37 8C, color was developedby adding 0.6 mL of Fast Garnet GBC (Sigma) solution (5 mg/mL in10% w/v SDS) and further incubation at room temperature for15 min. The absorbance was measured at 560 nm in a spectrophotometer(CECIL 2021, Cambridge, UK). Controls containing thereaction mixture plus glacial acetic acid were also incubated to testfor the presence of background activity. A standard curve wasprepared using a-naphthol. A unit of esterase activity was definedas the amount of enzyme that released 1 mmol of a-naphthol perminute. Specific esterase activity was defined as units permilligram of protein.
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