237 / 311
to 4.5 with 1 M NaOH solution. Dilute to 1 L with water.
(
2
)
100 mM, pH 5.0
.—Same procedure as for (
a
)(
1
), except
pH is adjusted to 5.0.
(
b
)
Heat-stable
-amylase solution.
—Heat-stable
-amylase, ca 20 000 liquefon units/g, specific gravity 1.25
(Product Multifect AA 21L, Genencor International,
Rochester, NY; origin:
Bacillus licheniformis
; pH optima,
5.5–5.8).
(
c
)
Amyloglucosidase solution.
—(
1
)
7 U/8 mL.—
Pipet
0.067 mL concentrated amyloglucosidase and dilute to
250 mL with 100 mM sodium acetate buffer, (
a
)(
1
)
.
(
2
)
100 U/mL
.—Dilute 0.77 mL concentrated
amyloglucosidase with 100 mM sodium acetate buffer, (
a
)(
2
)
,
to 25 mL (concentrated amyloglucosidase, 3260 U/mL,
Product E-AMGDF, Megazyme International Ireland, Ltd,
Bray Co., Wicklow, Ireland; origin:
Aspergillus niger
;
optimum pH, 4.0; stable pH, 4.0–5.5).
(
d
)
Glucose oxidase–peroxidase–aminoantipyrine buffer
mixture.
—Mixture of glucose oxidase, 7000 U/L; peroxidase,
7000 U/L; and 4-aminoantipyrine, 0.74 mM. Prepare by
dissolving 9.1 g Na
2
HPO
4
and 5.0 g KH
2
PO
4
in ca 300 mL
distilled water in a volumetric flask. Use distilled water to
rinse chemicals into bulb of flask. Swirl flask to dissolve
completely. Add 1.0 g phenol (ACS grade) and 0.15 g
4-aminoantipyrine. Use distilled water to rinse chemicals into
bulb of flask. Swirl flask to dissolve completely. Add glucose
oxidase (7000 U) and peroxidase (7000 U), rinse enzymes
into flask with distilled water, and gently swirl flask to
dissolve contents without causing excessive foaming. Dilute
to 1 L with distilled water. Seal flask and invert repeatedly to
mix. Filter solution through a glass fiber filter with 1.6 m
retention. Store in a sealed amber bottle at ca 4 C. Determine
standard curve for the reagent, using a 4-point standard curve
with distilled water and (
e
), according to
Preparation of
Reagent Blanks and Standard Curves
, (
b
)(
1
).
(
e
)
Glucose standard solutions.
—40, 60, and 80 g/mL.
Determine the dry matter of powdered crystalline glucose
(purity 99.5%). Weigh approximately 20, 30, and 40 mg
glucose, and record weight to 0.0001 g. Dissolve each portion
separately in distilled water. Dilute each solution to 500 mL to
obtain 3 independent glucose standard solutions. Multiply the
weight of glucose by the percentage of dry matter, and divide
H
ALL
: J
OURNAL OF
AOAC I
NTERNATIONAL
V
OL
. 92, N
O
. 1, 2009
45
Table 2. Results for determinations of free glucose and starch + maltooligosaccharides corrected for free glucose in
food and feed samples (dry matter basis)
Hot water
Acetate buffer
Extension of
AOAC
996.11
Sample
DM, %
a
Mean
s
r
b
Mean
s
r
Mean
s
r
Free glucose
Alfalfa silage
90.0
0.12
0.00
0.14
0.00
0.12
0.01
Corn grain, ensiled
93.5
0.44
0.00
0.38
0.00
0.18
0.00
Corn silage
91.1
0.05
0.00
0.07
0.00
0.07
0.00
Rice
90.4
0.18
0.01
0.20
0.01
0.02
0.02
Soybean meal
92.2
0.08
0.00
0.04
0.00
0.05
0.00
Split peas
91.4
0.07
0.00
0.04
0.00
0.02
0.02
Wheat flour
90.9
0.17
0.02
0.13
0.00
0.05
0.02
Starch + maltooligosaccharides corrected for free glucose
Alfalfa silage
90.0
1.5
0.1
1.6
0.0
1.4
0.0
Corn grain, ensiled
93.5
69.0
0.1
75.6
0.8
70.4
0.2
Corn silage
91.1
7.3
0.8
37.1
0.4
34.2
0.9
Rice
90.4
82.0
0.4
83.3
1.3
82.4
0.1
Soybean meal
92.2
1.4
0.0
0.8
0.0
0.9
0.0
Split peas
91.4
48.4
0.3
50.1
0.7
48.1
0.1
Wheat flour
90.9
74.3
0.5
79.6
0.2
74.8
0.1
Average of feed/food samples
Mean
40.6
46.9
44.6
s
r
0.31
0.48
0.20
CV, %
c
2.57
1.55
0.78
a
DM = Dry matter.
b
s
r
= Standard deviation of replicates.
c
CV = Coefficient of variation (s
r
/mean).