PROCESSES FOR INCREASING EXTRACTION OF ENZYMES FROM ANIMAL FEED AND MEASURING ACTIVITY OF THE SAME
Methods of increasing extraction of enzymes from animal feed and measuring enzyme activity are described herein.
1 . A method for measuring activity of phytase in an animal feed comprising:
mixing an amount of an animal feed with a carbonate-bicarbonate buffer to obtain a mixture, wherein the animal feed includes phytase, and the carbonate-bicarbonate buffer includes sodium carbonate at a concentration in a range from 10 mM to 500 mM and sodium bicarbonate at a concentration in a range from 10 mM to 500 mM;
extracting the phytase from the mixture; and
measuring the activity of an extracted phytase.
2 . The method of claim 1 further comprising grinding an animal feed to form flour prior to the step of mixing.
3 . The method of claim 2 , wherein the flour comprises particles having a size within a range of 250 μm to 6,000 μm.
4 . The method of claim 3 , wherein the size is at least 250 μm.
5 . The method of claim 1 , wherein a pH of the carbonate-bicarbonate buffer is 10.00, or greater.
6 . The method of claim 1 , wherein the carbonate-bicarbonate buffer further comprises a nonionic detergent.
7 . The method of claim 1 , wherein the amount of animal feed is within a range from 100 g to 500 g.
8 . The method of claim 1 , wherein a temperature of the mixture is in a range 20° C.-80° C.
9 . The method of claim 1 , wherein phytase is produced in a genetically engineered host.
10 . The method of claim 9 , wherein the host is selected from a plant cell, a bacterial cell, a mammalian cell, and a yeast cell.
11 . The method of claim 1 , wherein the phytase is an E. coli phytase.
12 . The method of claim 1 , wherein measuring of the phytase activity is performed at a temperature within a range of 20° C.-80° C.
13 . The method of claim 12 , wherein the temperature is 37° C.
14 . A method for extracting a feed enzyme comprising:
mixing an amount of animal feed with a carbonate-bicarbonate buffer to obtain a mixture, wherein the animal feed includes a feed enzyme, and the carbonate-bicarbonate buffer includes sodium carbonate at a concentration in a range from 10 mM to 500 mM and sodium bicarbonate at a concentration in a range from 10 mM to 500 mM; and
extracting the feed enzyme from the mixture.
15 . The method of claim 14 , wherein a pH of the carbonate-bicarbonate buffer is 10.00, or greater.
16 . The method of claim 14 , wherein the carbonate-bicarbonate buffer further comprises a nonionic detergent.
17 . The method of claim 16 , wherein the nonionic detergent is a polysorbate.
18 . The method of claim 17 , wherein the polysorbate is at a concentration in a range from 0.001% (v/v) to 1.0% (v/v).
19 . The method of claim 14 , wherein the mixture comprises the amount of animal feed and the carbonate-bicarbonate buffer at a ratio of less or equal to one selected from the group consisting of: 1:5 (w/v), 1:10 (w/v), 1:20 (w/v), 1:50 (w/v), 1:60 (w/v), 1:70 (w/v), 1:80 (w/v), 1:90 (w/v), 1:100 (w/v), 1:200 (w/v), 1:300 (w/v), 1:400 (w/v), 1:500 (w/v), 1:600 (w/v), 1:700 (w/v), 1:800 (w/v), 1:900 (w/v) and 1:1000 (w/v).
20 . The method of claim 14 , wherein the amount of animal feed is within a range from 100 g to 500 g.
21 . The method of claim 15 further comprising grinding the animal feed to form flour, wherein the flour comprises particles having a size within a range of 250 μm to 6,000 μm.
22 . The method of claim 21 , wherein the size is at least 250 μm.
23 . The method of claim 14 , wherein a temperature of the mixture is in a range 20° C.-80° C.
24 . The method of claim 14 , wherein the temperature is 50° C.
25 . The method of claim 14 , further comprising measuring the activity of the feed enzyme.
26 . The method of claim 14 , wherein the feed enzyme is produced in a genetically engineered host.
27 . The method of claim 26 , wherein the host is selected from a plant cell, a bacterial cell, a mammalian cell, and a yeast cell.
28 . The method of claim 14 , wherein the feed enzyme is selected from the group comprising phytase, xylanase, glucanase, endoglucanase, cellobiohydrolase, amylase, protease, mannanase, arabinofuranosidase, xylosidase, glucoamylase, pectinase, lignin peroxidase, esterase, or cellulase.
29 . The method of claim 28 , wherein the phytase is an E. coli phytase.
30 . The method of claim 25 , wherein the step measuring is performed at a temperature within a range of 30° C.-80° C.
31 . The method of claim 30 , wherein the temperature is 37° C.