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Effect of Calcium Sulfate Concentration in Soymilk
Effect of Calcium Sulfate Concentration in Soymilk
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The effects of calcium sulfate concentration in soymilk on the microstructure of tofu and the
constitutions of protein in tofu whey were investigated. The firm tofu made with 0.4% CaSO4‚2H2O
was found to be most uniform and continuous in the microstructure. This tofu gave the maximal tofu
yield, maximal solid and protein recoveries, and the best water retention ability. The results of gel
electrophoresis and the ratio of amino nitrogen to total organic nitrogen indicated that the low molecular
weight proteins in tofu whey were at their lowest when the tofu was made with 0.4% CaSO4‚2H2O.
The SEM observations suggested that the tofu made with 0.4% CaSO4‚2H2O has the most uniform
and homogeneous microstructure and, consequently, can most efficiently retain soybean proteins
and water in the tofu gel.
2H2O, and this was similar to that obtained with CaSO4‚1/2H2O Table 1. Effect of Calcium Sulfate Concentration in Soymilk on
by deMan et al. (2). When the CaSO4‚2H2O concentration was Moisture Content, WRA, Tofu Yield, Solid Recovery, and Protein
increased to 0.5%, the network of tofu became too porous and Recovery of Firm Tofua
too compact (Figure 1D) and not as uniform and smooth as CaSO4‚ moisture tofu yield solid protein
that obtained with 0.4% CaSO4‚2H2O. 2H2O content WRAb (g tofu/ recovery recovery
Effect of CaSO4‚2H2O Concentration on Yield, WRA, (%, w/v) (%) (%) 100 g soybean) (%) (%)
Protein, and Solid Recoveries of Firm Tofu. In this study, 0.20 80.2 a 67.5 c 205 e 45.9 c 62.6 e
we investigated the effects of CaSO4‚2H2O concentrations, from 0.25 80.2 a 67.7 c 230 d 49.7 b 67.0 cd
0.2 to 0.5%, on the microstructure and physical properties and 0.30 80.9 a 70.3 b 237 cd 51.2 b 67.9 bcd
0.35 80.8 a 75.6 a 243 bc 51.9 b 68.4 bc
found that 0.4% CaSO4‚2H2O in soymilk produced tofu with a 0.40 80.5 a 77.2 a 253 a 55.2 a 70.9 a
maximum tofu yield (253 g tofu/100 g soybean), maximum 0.45 80.7 a 76.9 a 247 ab 53.7 ab 69.5 ab
protein recovery (70.9%), maximum solid recovery (55.2%), 0.50 81.0 a 76.0 a 240 bc 51.5 b 68.3 bc
and a maximum WRA (77.2%). All four parameters increased
significantly as the CaSO4‚2H2O concentration increased from a Mean scores bearing the same letters among the same column are not
0.2 to 0.4% but decreased as the CaSO4‚2H2O concentration significantly different (p < 0.05). b WRA of tofu.
increased from 0.4 to 0.5% (Table 1). At the lower calcium
sulfate concentrations, the interaction of the protein molecules and other soluble substances. These events would seem to
with calcium ion was not strong enough to form a firm tofu account for the lower yield of tofu (240 g tofu/100 g soybean),
gel. There were many discontinuous fragments and large holes lower protein recovery (68.3%), and lower solid recovery
in the microstructures of tofu made with 0.2 or 0.3% CaSO4‚ (51.5%) at this concentration. At the optimum coagulant
2H2O (Figure 1A,B). The exclusion of noncoagulated proteins concentration (0.4% CaSO4‚2H2O), a uniform, continuous tofu
and other soluble solids with the water during the pressing step network was formed (Figure 1C), and it was able to trap more
resulted in a lower tofu yield and the lower protein and solid water and soluble substances. Therefore, it could obtain the
recoveries (Table 1). On the contrary, at a high concentration highest tofu yield, protein and solid recoveries, and WRA (Table
of 0.5% CaSO4‚2H2O, too many cross-linkages were formed 1). These observations were in agreement with the findings of
and resulted in a too compact and too porous structure (Figure Puppo and Añón (17), who showed that a protein gel with a
1D). This structure was less uniform and less homogeneous than homogeneous and fine structure gave high WRA, when
that with 0.4% CaSO4‚2H2O. This, in turn, resulted in an compared to the gel with a nonhomogeneous structure, that had
increased syneresis and induced the loss of water, whey proteins, a high degree of syneresis.
Table 2. Effect of Calcium Sulfate Concentration in Soymilk on the
Textural Characteristics of Firm Tofua
CaSO4‚
2H2O hardness springiness gumminess chewiness
(%, w/v) (kg) (mm) cohesiveness (kg) (kg mm)
0.20 0.40 b 7.84c 0.35 d 0.14 c 1.11 d
0.25 0.45 b 8.70 b 0.39 c 0.18 c 1.52 c
0.30 0.66 a 9.74 a 0.40 bc 0.26 b 2.57 b
0.35 0.68 a 9.76 a 0.41 b 0.28 b 2.72 b
0.40 0.71 a 9.76 a 0.45 a 0.32 a 3.11 a
0.45 0.73 a 9.84 a 0.45 a 0.33 a 3.21 a
0.50 0.74 a 10.32 a 0.45 a 0.33 a 3.39 a
a Mean scores bearing the same letters among the same column are not
trends (Table 2). significantly different (p < 0.05). b TON, subtraction of ammonium nitrogen from
Effect of CaSO4‚2H2O Concentration on the Constitution TN.
of Tofu Whey Proteins. The effect of CaSO4‚2H2O concentra-
tions on the constitution of tofu whey proteins was analyzed The increase of CaSO4 concentration from 0.2 to 0.4% in
by TSDS-PAGE (Figure 2), a method that had been shown to tofu making resulted in a decrease in TN and AN in tofu whey
have an improved resolution for LMW proteins (22). The and an increase in TN and AN when beyond 0.4% CaSO4‚2H2O
TSDS-PAGE pattern of soymilk protein showed that the protein (0.04 and 0.013%, respectively). The results supported the
bands with molecular masses in the range of 16-40 and 42- previous finding that tofu with a homogeneous and uniform
80 kDa were the two major proteins of soybean, i.e., 11S network gave the highest protein recovery at the optimum
(glycinin) and 7S (β-conglycinin) globulins. Glycinin is com- coagulant concentration (Table 1). Among the four tofu whey
posed of acidic (34-45 kDa) and basic (17-20 kDa) polypep- samples, the percentages of LMW proteins (Figure 2, lane 4)
tide chains paired by a disulfide bond (27, 28). β-Conglycinin and the ratio of AN to TON (Table 3) were the lowest in tofu
consists of R, R′, and β subunits, wherein the molecular masses whey obtained from the soymilk with 0.4% CaSO4‚2H2O (29.0
of R and R′ subunits are reported to be in the range of 57-83 and 39.4%, respectively). This suggested that a homogeneous
kDa, while that of the β subunit is 42-53 kDa (29). network can more efficiently retain the LMW proteins in tofu
As compared with soymilk protein composition, the tofu whey matrixes.
protein contained higher amounts of LMW proteins with a A possible gelation mechanism for firm tofu in the presence
molecular mass below 16 kDa (Figure 2). The protein bands of CaSO4 is schematically represented in Figure 3. It has been
on the gel were densitometrically scanned, and the relative ratio suggested that a Ca2+ gel is induced in the denatured protein
of bands was estimated. The percentages of LMW proteins in solution to form a filamentous gel structure (30-35). Heating
soymilk and tofu whey that were discharged from the pressed a soybean protein solution causes the proteins to unfold and
tofu made with 0.2, 0.3, 0.4, and 0.5% CaSO4‚2H2O were 1.8, form protein filaments due to the formation of disulfide bonds
41.1, 81.0, 29.0, and 53.7%, respectively. A similar trend was and hydrophobic interactions. A filamentous gel structure is
found in the ratio of AN to TON in soymilk and tofu whey formed when CaSO4 is added, and this reduces the electrostatic
(Table 3), with the values of 0.93, 43.1, 66.7, 39.4, and 57.5%, repulsion between the charged filaments, causing them to
respectively. These results confirmed the suggestion that the aggregate with the Ca2+ ions acting as bridges between the
proteins remaining in tofu whey were mainly LMW proteins adjacent charged carboxylic groups on neighboring protein
(15). molecules. At the end of pressing, the ratio of tofu whey volume
Figure 4. Representation (highly schematic) of likely states of constitution
of protein filaments and LMW proteins in the tofu network and in the tofu
whey obtained with different concentrations of CaSO4‚2H2O in soymilk;
(O) representing HMW protein; (b) representing LMW protein. The eight
Figure 3. Gelation mechanism of firm tofu in the presence of CaSO4; states were as follows: (A) 0.2% (tofu network), (B) 0.3% (tofu network),
(O) representing HMW protein; (b) representing LMW protein. Each (C) 0.4% (tofu network), (D) 0.5% (tofu network), (E) 0.2% (tofu whey),
square area represents a relative volume. (F) 0.3% (tofu whey), (G) 0.4% (tofu whey), and (H) 0.5% (tofu whey).
The left-hand square areas represent the relative volumes of tofu, and
to firm tofu volume was in the range of 2.4-3.2. An increase the right-hand square areas represent the relative volumes of tofu whey.
in the CaSO4 concentration can promote gelation of the protein
filaments. At a low CaSO4 concentration (0.2% CaSO4‚2H2O), and consequently, resulted in the maximal tofu yield, maximal
the protein filaments failed to form a firm tofu gel (Figure 4A), protein and solid recoveries, and the best WRA.
with many individual protein filaments and LMW proteins
dispersed in tofu whey (Figure 4E). When the CaSO4‚2H2O ABBREVIATIONS USED
concentration was increased to 0.3%, the extent of aggregation SEM, scanning electron microscopy; GDL, glucono-δ-lac-
of the protein filaments in tofu gel increased remarkably. tone; LMW, low molecular weight; HMW, high molecular
However, the gel was still not homogeneous and exhibited a weight; WRA, water retention ability; TSDS-PAGE, tricine
fragmentary network with a relatively low retention of water sodium dodecyl sulfate-polyacrylamide gel electrophoresis; TN,
and LMW proteins (Figure 4B) and resulted in a maximum total nitrogen; FN, formol nitrogen; AN, amino nitrogen; TON,
percentage of LMW proteins dispersed in tofu whey (Figure total organic nitrogen; ANOVA, analysis of variance.
4F). As the CaSO4‚2H2O concentration further increased to the
optimum concentration (0.4% CaSO4‚2H2O), the LMW proteins ACKNOWLEDGMENT
were well-dispersed in the spaces (pores) of the homogeneous
network structure (Figure 4C) with only a few protein filaments We are grateful to Miss Chien Lin for technical assistance with
and LMW proteins dispersed into tofu whey (Figure 4G). As picture drawing.
the CaSO4 concentration was further increased to 0.5%, the
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