Functional Properties of Three Soymilk Curds Prepared with

Transcription

1 Agric. Biol. Chem., 52 (5), , Functional Properties of Three Soymilk Curds Prepared with an Enzyme, Calcium Salt and Acid1" Katsumi Murata, Isao Kusakabe,* Hideyuki Kobayashi,* Hiroshi Kiuchi and Kazuo Murakami* Research Development Section, Kibun Food Chemifa Co., Ltd., 2-5, Shinbashi 3-chome, Minato-ku, Tokyo 105, Japan * Institute of Applied Biochemistry, University of Tsukuba, Tennohdai 1-chome, Tsukuba-shi, Ibaraki 305, Japan Received October 19, 1987 A comparative investigation was carried out on functional properties of three kinds of soymilk curds, which were prepared with an enzyme, calcium chloride and an acid. The enzyme-curd showed high emulsion stability over wide ranges of ph and temperature, whereas the Ca- and acidcurds produced lower values at ph 4~6 and temperatures higher than 40 C. The foam stability of each soymilk curd was low, but the enzyme-curds prepared from defatted soymilk had higher foam stability than the others over wide ranges of ph and temperature. The lyophilized enzyme soymilk curd solubility in water was higher than the others, although its water-holding capacity was lower than the others. None of the soymilk curds showed gelation. The experimental results suggest that enzyme-curd could be widely applicable to various food items with its properties of a remarkably high emulsion stability and a very smooth texture. Wereported in the preceding papers1'2) that acid, neutral and alkaline proteinase originating from microorganisms, plants and animals were capable of coagulating soymilk-protein, that alkaline and neutral proteinases from microorganisms were suitable for the industrial production of soymilk curd, and that the soymilk curd made with these enzymes had a smooth texture and "umami". However, no other study has been reported on the functional properties of the soymilk curd, although it has been reported that soy protein was coagulated by acid3) and inorganic salts (calcium and magnesium salts),4) and that acid- and Ca-soymilk curds were made with these coagulants. For this reason, this paper reports a comparative investigation of the functional properties of soymilk curds prepared with an enzyme, calcium chloride and acid. MATERIALS AND METHODS Preparation ofsoymilk curds. The enzyme employed was Subtilisin Carlsberg (Sigma Chemical Co., U.S.A.). "Kibun Tonyu Muchosei" soymilk produced by Kibun Food Chemifa Co., Ltd. was used, the ph value and protein content of the soymilk being 6.8 and 4.4% (w/v), respectively. A mixture of the soymilk (500 ml adjusted to ph 6.1 with a 1 m sodium phosphate buffer solution) and enzyme solution (50ml containing 500units in 5mg of enzyme) was incubated at 65 C for 15min. The resultant coagulum was centrifuged at 1,600xg for lomin, and separated into soymilk curd (tentatively referred to as the enzyme-curd) and whey. A soymilk curd from calcium chloride (Ca-curd) was prepared by adding a calcium chloride solution (30mM) to soymilk according to the conditions already described, and a soymilk curd from acid (acid-curd) was prepared by adjusting the ph of soymilk to ph 4.5 with hydrochloric acid. Chemical analysis. The moisture content was measured by heating at 105 C until the weight reached a constant value. The protein content was measured by the Kjeldahl method, and the fat content by the Soxhlet method with ethyl ether as a solvent5* after lyophilizing the soymilk Offprint requests should be directed to.isao Kusakabe.

2 1136 K. Murata et al. curd. The ash content was determined by heating at 550~600 C until the weight reached a constant value. Measurement of the functional properties. Mcllvaine buffer (ph 2 ~8) and 1,3-propandiamine-HCl buffer (ph 10) solutions were used. The effects of ph and temperature on the functional properties were measuredat room temperature (20 C) and ph 7, respectively. a) Emulsion stability. This was determined by using a modified version of the method of H. Aoki et al.6) A mixture of soymilk curd (5 g) and buffer solution (30ml) was homogenized at 14,000rpm for 1min in a Waring Blendor (ACE AM-1 homogenizer, Nihon Seiki Kaisha Ltd., Japan), and immediately transferred to a measuring cylinder. After standing for 30min, 5ml of the resultant emulsion from the bottom of the cylinder was taken out, and its moisture content was measured. The emulsion stability (ES) was calculated from the equation ^5(%) =(100-Mtest)/(100-Moriginal) x l00, where Mtest was the moisture content of the lowermost 5ml of sample (%), and Moriginal was the initial moisture content of the sample (%). b) Foam stability. This was measured by using a modified version of the method of Kuwata et al.1) A mixture of 5g of soymilk curd or defatted soymilk curd and 20ml of a buffer solution was homogenized at 14,000rpm for 2min, and immediate transferred to a measuring cylinder. After standing for 30 min, the volume of the water phase (drained) was measured. Foamstability (FS) was calculated from the following equation FS=(VT- KD)/20, where VTwas the total volume (ml) after homogenizing, and VD was the volume (ml) drained after 30min. In addition, a 10% (w/w) dispersion of defatted soy flour was prepared with a 0.1% NaH2PO4solution in tap water by stirring at 70 C for 30min. The supernatant liquid obtained after centrifugation at 6,000 x g for 20 min was used as defatted soymilk. The ph and the protein content of the defatted soymilk were 7.4 and 4.6% (w/v), respectively. Defatted soymilk curd was prepared according to the same method as that for soymilk curd. c) Solubility in water. This was measured by using a modified version of the method ofh. Aoki et al.8) Soymilk curd was used, which had been lyophilized after freezing in a dry ice-methanol bath and then passed through a screen (40 mesh). Twenty ml of a buffer solution containing 0.2 g of the lyophilized soymilk curd was stirred for 30 min and then centrifuged at 8000 x g for 20min. The solubility (%) was calculated from the ratio of the protein content of the supernatant liquid to that of the original. d) Water-holding capacity. This was measured by using a modified version of the method quoted in- the Japan Agricultural Standard (JAS).9) Twenty ml of a buffer solution containing 2 g of lyophilized soymilk curd was incubated at 70 C for 30min and then centrifuged at 1000 x g for 5min. The water-holding capacity (WHC) was calculated by the equation WHC=WR/WDx100, where WKwas the weight (g) of the precipitate after centrifugation, and WDwas the weight (g) of the dry matter in the soymilk curd. e) Gelation. This was measured by using a modified version of the method in JAS.9) Soymilk curd, filling a tube made with plastic film (25 mmdiameter), was heated in a boiling water bath for 30min. Gelation was judged by whether the whole soymilk curd coagulated or not. RESULTS AND DISCUSSION Effects of phand temperature on the emulsion s tab ility Figure 1 shows the effect of ph on the emulsion stability of soymilk curd. The stability of the enzyme-curd had a high value (more Fig. 1. Effect of ph on the Emulsion Stability of This experiment was carried out at 20 C. O, enzyme-curd; A, Ca-curd; å, acid-curd. Fig. 2. Effect of Temperature on Emulsion Stability of This experiment was carried out at ph 7. This symbols are

3 Functional Properties of Soymilk Curds Tabl I. Composition of the Soymilk Curds (%) Enzyme-curd E-DC Ca-curd Ca-DC Acid-curd A-DC Moisture Protein Fat Ash E-DC, enzyme-defatted soymilk curd; Ca-DC, Ca-defatted soymilk curd; A-DC, acid-defatted soymilk curd. than 90%) in the range of ph 2~10, whereas that of the Ca- and acid-curds decreased remarkably in ph range 4~6. Figure 2 shows the effect of temperature on the stability. The stability of the enzyme-curd was high (more than 90%) in the range of 20~80 C, whereas that of the Ca- and acid-curds decreased remarkably with increasing temperature, being less than 10% at 80 C. It has been reported that the emulsifying property of soy protein, which was considerably influenced by ph,6) increased when the soy protein was hydrolyzed by an acid or enzymeunder appropriate conditions,10) and also when the soy-protein content was high.n) Moreover, both the emulsion capacity and emulsion stability of soy protein showedminimumvalues in the region of the isoelectric point (ph 4.5^4.8).6) From the results of Fig. 1, it can be seen that the emulsion stability of the enzyme-curd, in spite of a comparatively low protein content (Table I), was higher than that of the Ca- and acidcurds. It seems that the high emulsion stability of the enzyme-curd was due to appropriate hydrolysis of the soymilk-protein by an enzymatic action and that the modification of the protein by enzymatic hydrolysis probably resulted in the formation of some amphiphilic fragments having various isoelectric points. These fragments seem to have played an important role in the stabilization of the emulsion, since amphiphilic proteins were adsorbed onto the surface of oil globules through hydrophobic interaction, and stabilized the emulsion.12) Effects of phand temperature on the foam s tab ility Figures 3 and 4 show the effects of ph and Fig. 3. Effect of ph on the Foam Stability of Soymik Curds and Defatted This experiment was carried out at 20 C. A and B show the results of soymilk curds prepared from soymilk and soymilk curds prepared from defatted soy flour, respectively. O, enzyme-curd; A, Ca-curd; å > acid-curd; #, enzyme-defatted soymilk curd (E-DC); A, Ca-defatted soymilk curd (Ca-DC); å, acid-defatted soymilk curd (A- DC). Fig. 4. Effect of Temperature on the Foam Stability of Soymilk Curds and Defatted This experiment was carried out at ph 7. The symbols are the same as those in Fig. 3. temperature on the foam stability of soymilk curd and defatted soymilk curd. In Figs. 3A and 4A, the stability was low in all curds. It has been reported that the existence of fat decreased the foam stability.11} The soymilk

4 1138 K. Murata et ah curds used in this experiment contained 6 ~ 7% of fat (Table I). Therefore, the measurement of foam stability was carried out by using curd prepared from defatted soymilk to avoid the effect of fat. The moisture contents of the enzyme-defatted soymilk curd (E-DC), Cadefatted soymilk curd (Ca-DC) and acid- defatted soymilk curd (A-DC) were 85.6, 82.3 and 80.9%, respectively. Their protein contents were 12.1, 12.4 and 15.4%, respectively. Figures 3B and 4B show the foam stability of the defatted soymilk curds. The foam stability ofe-dc was higher than those ofca-dc and A-DC in the range of ph 2-10, and had the highest value (2.9) at ph 4. On the other hand, the foam stability ofe-dc had a high value in the range of 20~80 C, and was more than twice as high as that ofca-dc and A-DC. It has been reported that the foam property was improved whensoy protein was hydrolyzed by an enzyme under appropriate conditions.13) Therefore, it seems, as was the case with emulsion stability, that the high foam stability of E-DC was due to appropriate hydrolysis. Effect of ph on the solubility in water Figure 5 shows the effect of ph on the solubility of lyophilized soymilk curd. The solubility of the three curds was a minimumat ph 4 and increased with increasing ph. The solubility of the enzyme-curd was more than twice as high as that of the Ca- and acid-curds in the range of ph 2~10, the maximumand minimum values being 32% (at ph 10) and 19% (at ph 4), respectively. It has been reported that the solubility of soymilk-protein showed a minimumvalue in the region of the isoelectric point.14) The solubility of the soymilk curds also showed a similar tendency. It seems that the comparatively high solubility of enzyme-curd was due to the enzymatic hydrolysis of the soymilk-protein. Effect of ph on the water-holding capacity Figure 6 shows the effect of ph on the water-holding capacity of soymilk curd. The capacity of the enzyme-curd was lower than that of the Ca- and acid-curds in the range of Fig. 5. Effect of ph on the Solubility of This experiment was carried out at 20 C. The symbols are Fig. 6. Effect of ph on the Water-holding Capacity of This experiment was carried out at 70 C. The synibols are ph 2~10. This phenomenon seems to have been due to a dissolution of the enzyme-curd as shown in Fig. 5. On the other hand, it has been reported that the capacity of soy protein, which was precipitated by an acid, showed the lowest value in the region of the isoelectric point.15) The decrease of water-holding capacity of the acid-curd and enzyme-curd (except at ph 10) at the isoelectric point also showed a similar tendency. Gela tion None of the enzyme-, Ca- and acid-curds showed gelation. The enzyme-curd had a smooth texture and "umami", although the Ca- and acid-curds had a rough texture, and

5 Functional Properties of Soymilk Curds 1139 moreover, the Ca-curd had an astringent taste. In conclusion, the enzyme-curd, in contrast with the Ca- and acid-curds, had a remarkably high emulsion stability in wide range of ph and temperature, as well as a very smooth texture. Therefore, these properties indicate that the enzyme-curd could be widely applicable to various kinds of food products. REFERENCES 1) K. Murata, I. Kusakabe, H. Kobayashi, M. Akaike, Y. M. Park and K. Murakami, Agric. Biol. Chem., 51, 385 (1987). 2) K. Murata, I. Kusakabe, H. Kobayashi, H. Kiuchi and K. Murakami, Agric. Biol. Chem., 51, 2929 (1987). 3) F. Yamauchi, New FoodIndustry, 23, 35 (1981). 4) T. Watanabe, H. Ebine and T. Ohota, "Daizu Shokuhin," Kohrin, Tokyo, 1971, p. 75. T. Obara et al., "Shokuhin Bunseki Handbook," Kenpakushya, Tokyo, 1982, p H. Aoki and H. Nagano, GakkaishU 22, 320 (1975). Nippon Shokuhin Kogyo T. Kuwata, Y. Funakawa, H. Watanabe, H. Ohotomo and Y. Yamamoto, Nippon Shokuhin Kogyo GakkaishU 32, 639 '(1985). H. Aoki, N. Orimo', R. Shimazu and K. Wakabayashi, Nippon Shokuhin Kogyo Gakkaishi, 25, 668 (1978). Japan Agricultural Standard, 2601 (1976). H. Aoki and H. Nagano, Nippon Shokuhin Gakkaishi, 23, 26 (1976). Kogyo F. Yamauchi, New Food Industry, 22, 26 (1981). E. Tornberg, "Functionality and Protein Structure,"American Chemical Society, Washington DC, 1979, ACS Symp. Ser. No. 92, pp R. C. Gunther, /. Am. Oil Chemists' Soc, 56, 345 (1979). F. Yamauchi, Nippon Syokuhin Kogyo Gakkaishi, 26, 266 (1979). H. Aoki, Nippon Nogeikagaku Kaishi, 39, 270 (1965).