2.2. Extraction of proteins and oil bodies
Peanut protein and oil bodies were extracted using the graded extraction
method (Sukhotu, 2014) and aqueous enzymatic extraction of maize germ
(Li, Peng, & Li, 2015), respectively, with some modifications. The
skinless peanut seeds were ground in a high-speed universal grinder
(FW-100; Beijing Ever Bright Medical Treatment Inc, Beijing, China).
Twenty grams of peanut powder was dispersed in deionized water (the
solid-to-liquid ratio of 1:7, 1:6, 1:5, 1:4, 1:3, and 1:2 w/v).
Viscozyme® L (0.25, 0.50, 0.75, 1.00, 1.25, 1.50,
1.75, and 2.0%, calculated by the weight of total substrate) was added
to the mixture and enzymolysis was conducted in a digital water-bathing
constant temperature vibrator (THZ-82; Jinhua Huafeng Instrument Inc,
Jinhua, China) for different times (20, 40, 60, 80, 100, 120, and 140
min) and at different temperatures (35, 40, 45, 50, 55, 60, 65, and 70
°C). Thereafter, the enzyme was deactivated by placing the samples in a
boiling water bath for 5 min. The cooled solution was centrifuged at
5000 × g for 20 min (DZ267-32C6; Anting Scientific Instrument
Factory, Shanghai, China). The supernatant (oil bodies) was decanted and
dried at 50 ℃ for 10 h in a vacuum drying oven (DZF-2B; Beijing Ever
Bright Medical Treatment Inc, Beijing, China), whereas the precipitation
was dried in a freezer dryer (LGJ-25; Beijing Sihuan Scientific
Instrument Inc, Beijing, China) for 24 h and weighed. The amount of
protein was measured by an Automatic Kjeldahl Apparatus (K1100; Jinan
Haineng Instrument Inc, Shandong, China), and the yields of protein and
oil bodies were calculated using the formula1 and 2, respectively:
\begin{equation}
\mathrm{Yield\ of\ peanut\ protein=}\frac{\mathrm{\text{protein\ content\ of\ precipitate}}}{\mathrm{\text{protein\ content\ of\ peanut}}}\mathrm{\times 100\ }\left(\mathrm{\%}\right)\mathrm{\text{\ \ \ \ \ \ \ \ \ \ \ \ \ \ \ }}\mathrm{\ (1}\mathrm{)}\nonumber \\
\end{equation}\begin{equation}
\mathrm{Yield\ of\ oil\ bodies\ =}\frac{\mathrm{\text{dry\ weight\ of\ oil\ bodies}}}{\mathrm{\text{weight\ of\ peanut\ sample}}}\mathrm{\times 100\ }\left(\mathrm{\%}\right)\mathrm{\text{\ \ \ \ \ \ \ \ \ \ \ \ \ \ \ \ \ }}\mathrm{\text{\ \ \ \ \ \ \ \ \ \ \ \ }}\mathrm{\text{\ \ }}\mathrm{(2)\ }\nonumber \\
\end{equation}