74
Z. Xiao et al.
that means add Nutrase into seed mixture first, the mixture was incubated at 55 °C,
pH 7.0 for 2 h, and then add Cellulase into the same mixture, temperature and pH
were adjusted to 45 °C, and 4.5, respectively, and the mixture was incubated for 2 h
more).
Followed by centrifugation for 30 min at 10,000 rpm (Sigma, 3 K30, Osterode
am Harz, Germany). After centrifugation, the upper oily phase was withdrawn using
a pipette and weighted (w 1 , g). Emulsion was demulsified by vacuum dewatering at
90 kPa and at 50 °C for 4 h, followed by centrifugation for 30 min at 10,000 rpm,
upper oily phase was obtained and weighted (w 2 , g). The control samples were
prepared and treated identically without addition of enzymes. The free oil content
(%) and total oil recovering rate (ORR) of AEE compared to SE were calculated by
Eqs. (2.3) and (2.4):
Free oil content(%) =
w 1 + w 2
w 0
× 100%
(2.3)
ORR(%) =
Free oil content (%)
Total oil content(%)
× 100%
(2.4)
As shown as Table 2.8, a higher free oil recovery from castor seeds, in the range of
37.31–63.23% for the aqueous extraction process involving enzymes was observed
compared to only 9.16% for the control (oil obtained without enzyme treatment).
Among the enzymes studied, nutrase was found to be the best enzyme offering the
highest oil yield (63.23%), which was significantly (p < 0.05) higher than other
enzymes, whereas the oil recovering rate was minimum (37.31%) for the enzyme
of amylase treated seed samples. Overall, the tested enzyme mixtures exhibited
efficacy in the order: nutrase > pectinase > hemicellase > cellulose > amylase. The
high efficacy of proteolytic enzymes for free oil extraction is in agreement with the
findings of soybean and rapeseed. The enzyme action of nutrase during aqueous
extraction can be attributed to the degradation of cytomembrane, breakdown of the
protein networks of cotyledon cells and oleosin-based membranes that surround lipid
bodies, which results in liberating more oils than other enzymes.
The treatments of enzymatic combinations might improve the oil recovering rate,
nutrase, cellulose, and pectinase were chosen to combine with other enzymes to evaluate their cooperative effects on oil yield in this study. As shown in Table 2.9, the
enzymatic combinations obtained were in the range of 48.02–60.62% oil recovering
rate. Among the enzymatic combinations, nutrase coupled with cellulase had the
highest oil recovering rate of 60.62%, whereas the oil recovering rate was minimum
(48.02%) for cellulase combined with hemicellase. Overall, all enzymatic combinations did not increase the oil recovering rate compared to 63.23% for using the enzyme
of nutrase only. Similar results were also observed on peanut seeds oil enzymatic
extraction, which reported that the compounding alcalase with other enzymes did
not improve the peanut seeds oil yield significantly. Therefore, nutrase was chosen
for subsequent enzymatic hydrolysis process optimization in the present study.
Z. Xiao et al.
that means add Nutrase into seed mixture first, the mixture was incubated at 55 °C,
pH 7.0 for 2 h, and then add Cellulase into the same mixture, temperature and pH
were adjusted to 45 °C, and 4.5, respectively, and the mixture was incubated for 2 h
more).
Followed by centrifugation for 30 min at 10,000 rpm (Sigma, 3 K30, Osterode
am Harz, Germany). After centrifugation, the upper oily phase was withdrawn using
a pipette and weighted (w 1 , g). Emulsion was demulsified by vacuum dewatering at
90 kPa and at 50 °C for 4 h, followed by centrifugation for 30 min at 10,000 rpm,
upper oily phase was obtained and weighted (w 2 , g). The control samples were
prepared and treated identically without addition of enzymes. The free oil content
(%) and total oil recovering rate (ORR) of AEE compared to SE were calculated by
Eqs. (2.3) and (2.4):
Free oil content(%) =
w 1 + w 2
w 0
× 100%
(2.3)
ORR(%) =
Free oil content (%)
Total oil content(%)
× 100%
(2.4)
As shown as Table 2.8, a higher free oil recovery from castor seeds, in the range of
37.31–63.23% for the aqueous extraction process involving enzymes was observed
compared to only 9.16% for the control (oil obtained without enzyme treatment).
Among the enzymes studied, nutrase was found to be the best enzyme offering the
highest oil yield (63.23%), which was significantly (p < 0.05) higher than other
enzymes, whereas the oil recovering rate was minimum (37.31%) for the enzyme
of amylase treated seed samples. Overall, the tested enzyme mixtures exhibited
efficacy in the order: nutrase > pectinase > hemicellase > cellulose > amylase. The
high efficacy of proteolytic enzymes for free oil extraction is in agreement with the
findings of soybean and rapeseed. The enzyme action of nutrase during aqueous
extraction can be attributed to the degradation of cytomembrane, breakdown of the
protein networks of cotyledon cells and oleosin-based membranes that surround lipid
bodies, which results in liberating more oils than other enzymes.
The treatments of enzymatic combinations might improve the oil recovering rate,
nutrase, cellulose, and pectinase were chosen to combine with other enzymes to evaluate their cooperative effects on oil yield in this study. As shown in Table 2.9, the
enzymatic combinations obtained were in the range of 48.02–60.62% oil recovering
rate. Among the enzymatic combinations, nutrase coupled with cellulase had the
highest oil recovering rate of 60.62%, whereas the oil recovering rate was minimum
(48.02%) for cellulase combined with hemicellase. Overall, all enzymatic combinations did not increase the oil recovering rate compared to 63.23% for using the enzyme
of nutrase only. Similar results were also observed on peanut seeds oil enzymatic
extraction, which reported that the compounding alcalase with other enzymes did
not improve the peanut seeds oil yield significantly. Therefore, nutrase was chosen
for subsequent enzymatic hydrolysis process optimization in the present study.