Dextranase Liquid

Dextranase liquid activity assay method

(1) Preparation of Citric acid-disodium hydrogen phosphate buffer solution（PH5.5）:A solution: take out 20.369gNa2HPO4·12H2O dissolving in about 200ml water; B solution: take out 4.530g Citric Acid Monohydrate dissolving in about 200ml water then mix and dilute to 500 ml. 121 ℃, 20min sterilizing to standby.
(2) Weigh 2.0g dextran(T-2000) dissolving in buffer solution and up to  100ml as the substrate.
(3) Preparation of DNS solution
 take out 3.15g 3,5-Dinitrosalicylic acid dissolving in about 500 ml water, it can be heated to dissolved, but the temperature should not exceed 45℃.
weigh 20gNaOH dissolving in about 200 ml water.

Add  to  and then adding 91g KNaC4H4O6·4H2O, 2.5g phenol, 2.5g Anhydrous Sodium Sulfite, complete dissolution and dilute to 1000ml,then put it into a brown bottle, placed in a cool dark place 10 days later after filter paper filter to standby.

(4)Formulation of the enzyme activity standard curve :

take out 1g anhydrous glucose, roast 30 minutes under 60 degrees and raise the temperature to 80℃, and bake for 30 minutes, and then raise the temperature to 104℃, bake 1 hour, cooling in reserve.
The preparation of glucose standard solution: accurate weighing 0.1000 g of anhydrous glucose, dissolving and dilute to 100mL.

Take 6 colorimetric tube with the 25mL scale, according to the following table test.

 

                        
The value of OD540 (Y, ordinate) and glucose concentration (X, abscissa) on a computer or manual fitting curve, obtained the values of a and b in the  binary linear equation Y=aX+b

Requirements of the drawing curve correlation coefficient  r2 ≥ 0.999.

(5) detection of sample: take out 0.9mL substrate respectively added to the three 25mL colorimetric tube, placed the colorimetric tube in the constant temperature of 55℃ water bath and preheat at least 5 min, then respectively adding 0.1ml appropriate dilute enzyme solution to the two tubes, the third tube add 0.1mL distilled water as blank, exact reaction of 10min, DNS were immediately respectively joined 2mL and placed in a boiling water bath for accurate reaction of 5min, and then rapidly cooling, respectively with distilled water dilute to 25mL. Using ultraviolet spectrophotometer to measure，the blank reaction liquid as zero point, measuring the remaining two of the OD540 value and average.
The enzyme activity(U/mL) =1000 n (A-b) / (aMt)
Type A: is the average of the sample of OD540
b and a as the coefficients of the standard curve
n: Dilution ratio of the enzyme solution
t: enzymatic reaction time =10

M:The molar mass of glucose =180

 

CERTIFICATE OF ANALYSIS

 

Packaging and storage

Solid food packaging bag, 25 kg / barrel.

 

Dextranase liquid should be stored in dry, cool and well ventilated places away from the sun, heat.

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