Small talk about the determination of lipase activity - Database & Sql Blog Articles

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method:

1. Preparation of crude enzyme solution

The crude lipases were weighed 0.010 g, 0.020 g and 0.030 g, respectively, and dissolved in distilled water to a volume of 100 mL to prepare crude enzyme solutions at concentrations of 0.01%, 0.02% and 0.03%, respectively.

2, experimental design

In this experiment, 10 mL of salad oil was used as the substrate, and the optimal conditions for hydrolysis were selected by the determination of acid value based on the enzyme dosage, hydrolysis temperature and reaction time.

3. Determination of acid value

The acid value is the number of milligrams of NaOH required to neutralize 1 mole of free fatty acid, which is used to measure the degree of hydrolysis of the oil. In the experiment, the acid value of the hydrolyzate was determined by acid-base titration. Referring to the method used in the literature [3, 4], 1 mL of 95% ethanol solution was added to the obtained hydrolyzed droplets, shaken, the reaction was terminated, and the reaction was added. 2 drops of phenolphthalein indicator, quickly titrated with 0.05 mol/L NaOH solution until the solution is reddish, not disappearing within 30 s as the end point, record the number of milliliters (V) of the consumed NaOH solution. The blank value was measured in the same manner, and each test was repeated twice, and the average value was used as the measurement result.

The acid value is calculated as follows:

X = C (V - V0 ) × 40 /M

Where: X — fat acid value (mgNaOH / g oil)

C-NaOH standard solution concentration (mol/L)

M — mass of the sample ( g)

40-NaOH mmol mass (mg/mmol)

4 Determination of crude lipase activity

Under the optimal hydrolysis conditions, the acid values ​​X1 and X2 of the crude lipase and the standard lipase hydrolysate were measured, and the activity of the crude lipase was determined according to the formula U1 /U= X1 / X2, wherein U1 is the crude lipase activity. , U is the standard lipase activity.