Methods
The evaluate method to the pigment’s activity
of antiperoxidate of fat Taken 6 shares of 10 g
lard into conical flasks, and added 5 mL samples of
various concentration gradient (added 5 mL distilled
water into blank test and 5 mL 0.1 mg mL-1 Vc instead
of sample into control experiment) and chloroform-acetic
acid (3:2) 30 mL mixtured into each
flask. Then placed them in the constant incubator
in (60 ± 1)°C after well-mixed to test the peroxide
value (POV) every two days according to GB/T
5538-2005/ISO 3960:2001 (Standardization Administration
of the People’s Republic of China 2005).
Calculated the peroxide value by the following
equation:
X1
(meq kg-1) = 1 000(V-V0
) c/m (1)
Where X1
is the POV, in milligram equivalent per
kilogram (meq kg-1). V is the volume of sodium thiosulfate
solution (STS) used for the dete-rmination, in
millilitres (mL); V0
is the volume of STS used for the
blank determination, in millilitre (mL); c is the concentration
of the STS, in moles per litre (mol L-1); m is the
mass of the test portion, in grams (g).
The lard POV is stated less than 0.20 g 100 g-1
(2.00 g kg-1) according to GB10146-2005 (Standardization
Administration of the People’s Republic of
China 2005), so converted the peroxide value by the
following equation according to GB/T 5009.37-2003
(Standardization Administration of the People’s Republic
of China 2003)
X2= X1
/78.8 (2)
Where X2
is the peroxide value, in grams per
hektogram (g 100 g-1). 78.8 is conversion coeficient.
The evaluating methods to TAA, SARSA and
AOSA of the pigment These antioxidant assays
were taken according to the test manual supplied by
Nanjing Jiancheng Bio-Project Institute with a certain
mass of samples (Zong et al. 2006). The antioxidant
activities were measured by the following
equations:
TAA (U mL-1 pigment extraction) =100(A - Ac) 30×
[Total volume of reaction solution (mL)/Sample volume
(mL)]× The dilution multiple of the sample (3)
SARSA (U L-1)=[(Ac-A)/(Ac-As)]× 0.15 (mg mL-1)×
1 000 mL× The dilution multiple of the sample (4) AOSA (U mL-1) = [(Ac - A)/(As - Ab)] × 8.824
(mmol mL-1) × 1 (mL/sample volume) × The dilution
multiple of the sample (5)
Where A is absorption value of test tube. Ac is absorption
value of comparison tube. As is absorption
value of standard tube. Ab is absorption value of blank
tube. 0.15 mg mL-1 is concentration of Vc. 8.824
mmol mL-1 is concentration of standard tube.
As the antioxidant activity of every material varies
to each other under a fixed concentration reaction
system, and according to the Lambet-Beer’s
Law, the concentration of the samples should not
be too high or too low, which will affect on the
result. Therefore, before the experimentations of
SARSA and AOSA, a prearranged experimentation
should be done in order to determine the optimal
sample concentration by following the instruction
on the cases. There is a linearity relationship while
the sample depressing rate between 10-60% when
the activity is tested by the cases. The optimal concentration
tube should be selected of the depressing
rate between 40-50%, and if the rate is higher
than 60% or lower than 10%, the sample concentration
should be adjust to optimal one. Scavenging
rate is counted by the following equation.
Scavenging rate=(Ac - A)/Ac× 100% (6)
Where Ac is absorption value of comparison tube. A
is absorption value of test tube.
The evaluating method to DPPH.scavenging activity
of the pigment Added 0.10 mL various concentration
pigment samples (the comparison sample
added 0.10 mL 0.1 mg mL-1 Vc) into 3.00 mL DPPH
solution in 0.006 mol L-1, after homogenizing, placed
the samples into dark place for 30 min. Then tested
the absorption value of mixture solution at the 517
nm, and the DPPH.scavenging activity was calculated
by Eq. (6). In synergism test of the pigment
and Vc to scaveng DPPH., altered 0.10 mL pigment
to 0.05 mL pigment with 0.05 mL 0.1 mg mL-1 Vc,
other experimental operation was idem (Xie et al.
2006).