Evaluation of Soy Sauce Flavor by Stepwise Multiple Regression Analysis of Gas Chromatographic Profiles

The linear correlation was found by the stepwise multiple regression analysis between the sensory test of soy sauce flavor and the gas Chromatographic (GLC) data transformed with arc-sine and logarithm. GLC data will possibly be used for objective evaluation of soy sauce flavor. A multiple correlation coefficient or of a determination coefficient of more than 0.9 was respectively obtained at the 5 th or 10th of 47 steps. The fact that the minimum standard error of an estimate was found at the 24th step suggests the importance of selecting proper peaks from the whole gas chromatogram. High estimated accuracy was acquired by application of GLC data to the calculated multiple regression model.     

Comparison of Headspace and Distillation Techniques for Soy Sauce Aroma in Relation to Analysis by Silica Capillary Gas Chromatography and Sensory Evaluation

A method has been applied to the collection of headspace volatiles from soy sauce at room temperature, using Tenax GC adsorption polymer. Concentrated volatiles obtained from fifty soy sauce samples consisting of genuine- and semi-fermented types using Tenax GC and distillation-extraction methods were analyzed by silica capillary GC analysis in order to compare the suitability to sensory data. From a multiple regression analysis, close relationships between two variously transformed GC data sets and sensory data were found, the square root transformation of peak area from the headspace GC data being the best amongst them. The precise extraction of characteristics contained in the sample volatiles, using either of the two aroma collecting methods, was confirmed by cluster analysis.     

Comparative Volatile Profiles in Soy Sauce According to Inoculated Microorganisms

We compared the volatile profiles in soy sauce according to inoculation with Tetragenococcus halophilus and/or Zygosaccharomyces rouxii. Totals of 107 and 81 volatiles were respectively identified by using solid-phase microextraction and solvent extraction. The various volatile compounds identified included acids, aldehydes, esters, ketones, furans and furan derivatives, and phenols. The major volatiles in the samples treated with T. halophilus were acetic acid, formic acid, benzaldehyde, methyl acetate, ethyl 2-hydroxypropanoate, 2-hydroxy-3-methyl-2-cyclopenten-1-one, and 4-hydroxy-3-methoxybenzaldehyde, while those in the samples inoculated with Z. rouxii were mainly ethanol, acetaldehyde, ethyl propanoate, 2/3-methylbutanol, 1-butanol, 2-phenylethanol, ethyl 2-methylpropanoate, and 4-hydroxy-2-ethyl-5-methyl-3(2H)-furanone. The results indicate that T. halophilus produced significant acid compounds and could affect the Z. rouxii activity, supporting the notion that yeasts and lactic acid bacteria respectively have different metabolic pathways of alcoholic fermentation and lactic acid fermentation, and produce different dominant volatile compounds in soy sauce.     

Objective Evaluation of Soy Sauce by Statistical Analysis of GC Profiles

The relationships between GC data of volatiles and sensory score of the heighest grade of soy sauce (Tokusen shoyu) were analyzed by multivariate analyses. Samples belonging to four brands could be unambiguously classified into the correct brands and the statistical distance among them suggested a close relation with sensory evaluation. The precise predictive equations for the aroma quality were calculated from GC data and sensory evaluation by multiple regression analysis. Eight principal components were extracted from 39 GC peaks as significant factors constituting soy sauce aroma. The second PC can explain 58% of the variation among total variation contained in the sensory score. Discriminant functions on the basis of the 8 PCs can clearly classify all samples.     

Evaluation and Discrimination of Soy Sauce by Computer Analysis of Volatile Profiles

The relationships between gas chromatographic (GC) profiles and sensory data of 72 purely fermented soy sauce samples were analyzed by multiple regression analysis and principal component analysis (PCA). Prior to the analysis, GC data was transformed into 7 different modes in order to compare the fitting to a hypothetical linear model. The result from logarithmically transformed ratio of each peak to the sum of whole peaks showed the best precision of predictability for sensory score (R = 0.978). As the result of PCA, eigen values of 10 PCs were shown to be larger than 1.0 but the 5 major PCs could account for 66% of the variance in the total variance of 39 GC peaks. The first and second PCs showed great importance for aroma quality and similarity or dissimilarity in profiles of extracted PCs showed a similar trend with quality differences evaluated by sensory tests. These results showed the importance of the harmonious balance of each aroma compound to create a preferable soy sauce aroma.     

Non-enzymic Browning of Soy Sauce

Ion exchange resins have been used to separate soy sauce into three fractions of distinctly different composition: a cation fraction, a neutral fraction and an anion fraction. Almost all of the constituents responsible for browning were recovered in these three fractions.

Storage experiments show that when the three fractions were stored separately, only the cation fraction darkened considerably. When they were combined and stored, the color of the mixture increased at nearly the same rate as that of the original soy sauce. Neutral sugars are important constituents of the neutral fraction with respect to browning. The browning rate of a sugar-amino acid mixture (simulated soy sauce), was about 10% of soy sauce. The effect of the anion fraction (mainly caused by organic acids) and the ashed cation fraction on the over-all browning of soy sauce is calculated to be 1O~12% and 20%, respectively.

The sum of the contribution rate of the anion fraction, the neutral fraction, the amino acids and the ashed cation fraction in the browning of soy sauce was concluded to be approximately 40%. Compounds responcible for residual part of 60% should be considered to exist in the cation fraction. It was suggested that such compounds have strong reducing power and 0₂-uptaking ability.     

Persistence of Aflatoxin During the Fermentation of Soy Sauce

Aflatoxin was produced by Aspergillus parasiticus NRRL 2999 but not by A. oryzae during fermentation of soy sauce. Little aflatoxin was degraded within 6 weeks unless Lactobacillus delbrueckii also was present.     

从老抽酱醪中分离耐盐性酵母的研究

以老抽酱醪为实验材料进行耐盐性酵母菌种分离,并做菌种鉴定。分析了在不同盐度条件下耐盐性酵母菌的生长情况和生长过程中培养基总糖的消耗,可以发现实验得到的酵母在22％（质量与体积}E）盐度下依然能够良好生长。结果表明,实验分离出的No．2菌在同级盐度的条件下的生长量要明显高于No．1菌,但在乙醇产率方面,两株菌在相同的含盐量为16％（质量与体积比）的麦芽汁培养基中发酵8d,No．1菌的乙醇产率为3．1％（体积比）,No．2菌的乙醇产率2．9％（体积比）。     

Isolation and Characterization of Neutral Peptides in Soy Sauce

A soy sauce sample was fractionated by gel filtration on a Sephadex G–15 column, then the fractions were subfractionated on the basis of acidity by ion exchange chromatography on a QAE-Sephadex A–25 column. After preliminary fractionation, the components in the neutral subfractions were transformed to copper salts, and the salts were chromatographed on a DEAE-Sephadex A–25 column with a borate buffer and aqueous acetic acid to separate neutral peptide substances from a large amount of free amino acids. The peptide fractions were further fractionated on a preparative amino acid analyzer and by paper chromatography to separate the peptide substances.

Three glycodipeptides and eight dipeptides were isolated and characterized as the major neutral peptide components in soy sauce. However, it was difficult to anticipate any direct contribution of these peptides to the flavor construction in soy sauce on the basis of their taste intensities and contents.     

Isolation and Characterization of Acidic Peptides in Soy Sauce

A soy sauce sample was fractionated by gel filtration on a Sephadex G–15 column, then the fractions were subfractionated on the basis of acidity by ion exchange chromatography on a QAE-Sephadex A–25 column. The acidic subfractions with various acidities were further fractionated, using a preparative amino acid analyzer and by paper chromatography to separate the acidic peptide components.

Four dipeptides and sugar derivatives of ten dipeptides and two tripeptides were isolated and characterized as the major acidic peptides in soy sauce. However, it was difficult to anticipate any direct contribution of these peptides to the flavor construction in soy sauce on the basis of their contents and taste intensities.     

Studies on the Activities of Bacteria in Soy Sauce Brewing

The activity of proteinases in soy masy juice was measured by a modification of the Anson’s method. Compared to proteinases of purely cultured soy koji digest which consisted of only Aspergillus proteinases, no difference was observed with the proteinases of usual soy mash through the studies of pH-proteinese activity curves, the inhibition test, and inactivation by acid and alkali treatment. It is supposed, therefore, that the proteinases in soy mash are mostly of Aspergillus origin. Viable counts on salted media, heat shocking at 80°C for thirty minutes, and the microscopic counts proved that there are very few Bacilli vegetative cells in soy mash. From these facts the author assumed that Bacilli mainly grow in the koji and survive in the soy mash mostly in the form of spores. This work also suggested that the so-called “Aspergillus alkaline proteinase” consists of two distinct proteinases, and further the existence of an inhibitor which only acts at pH8.     

Automated Measurement of L-Glutamate in Soy Sauce Using L-Glutamate Dehydrogenase

An automated method for rapid and convenient measurement of L-glutamate has been developed by using a discrete analyzer, EEL Auto Chemist. It is based on the colorimetric measurement of NADH produced on a mole-mole basis by enzymatic dehydrogenation of L-glutamate using L-glutamate dehydrogenase from bovine liver. The values of L-glutamate obtained by this method were well agreed with those obtained by the routine Waruburg mano-metric method using L-glutamate decarboxylase from Escherichia coli.     

Production of Soy Sauce Koji Mold Spore Inoculum in Plastic Bags

An innovation is described for producing soy sauce koji mold spore inoculum by using inexpensive autoclavable plastic bags and reuseable plastic enclosures to make culture vessels. After growth, the spore mass could be dried and packaged in the same bag after removing the enclosure. Broken rice was used as the substrate for mold cultivation. Viable spore counts of 10⁹ spores per g were obtained under optimal conditions. After drying at 50°C for 6 h, the moisture content of the spore mass decreased from 35.22 to 6.32% with no significant effect on spore viability. The dry spores could be stored in the refrigerator or at room temperature for at least 3 months.     

Smith Degradation and Methylation of an Acidic Polysaccharide from Soy Sauce

APS–I, an acidic polysaccharide isolated from soy sauce, was subjected to Smith degradation and methylation, and each product obtained was examined by paper chromatography, gas-liquid chromatography and mass spectrometry. Some compounds, such as ethylene glycol, propylene glycol, glycerol, D-threitol, D-galactose, L-rhamnose, D-galacturonic acid and D-threonic acid, were identified as Smith degradation products, and their molar ratios were estimated. Several kinds of methylated sugars were detected in the hydrolysate of methylated APS–I and identified by gas chromatograph-mass spectrometry. On the basis of those findings and previous discussion, the constitution of APS–I was discussed.     

Isolation and Analysis of Molds from Soy Sauce Koji in Thailand

Five different isolates of Aspergillus and one of Mucor were compared with a Japanese commercial strain of Aspergillus oryzae for proteolytic activity on wheat bran substrate. One isolate of Aspergillus with superior protease production, identified as Aspergillus flavus var. columnaris, showed no detectable aflatoxin production on glutinous rice or soybean substrate. Preliminary tests using this fungus as a koji mold in a traditionally operated factory resulted in a soy sauce superior in quality to that usually produced.     

Mapping of Peptides in Soy Sauce by an Ion Exchange Chromatography

Pattern analysis of peptides in soy sauce was tried by gel filtration and ion exchange chromatography. In gel filtration through Sephadex G-15, the separation of smaller peptides and amino acids in soy sauce was found to be not so critical. Subsequently the mapping of peptides in soy sauce was done by ion exchange chromatography with Dowex 50-X2. Consequently it was estimated that there existed more than ten kinds of peptides in soy sauce.

Moreover, in the mapping of peptides, the automated measurement by the use of a discrete type analyzer, EEL Auto Chemist, was tried. The method developed here was not fully automated but available and effective for the mapping of peptides eluted from an ion exchange resin column.     

Acid and Enzymatic Hydrolysis of an Acidic Polysaccharide from Soy Sauce

Mild acid hydrolysis of an acidic polysaccharide (APS-I) from soy sauce resulted in a degraded polysaccharide (DPS), the mixture of neutral sugar, D-galacturonic acid, its α-1,4-linked homologous di- and trisaccharides, and acidic oligosaccharides containing residues of D-galacturonic acid and L-rhamnose. Besides the above-mentioned sugars, an aldobiouronic acid containing D-xylose moiety was also yielded in the enzymatic hydrolysates with a crude polysaccharidase preparation. However, only a β-l, 4-galactobiose was isolated from the lower molecular fraction of enzymatic digest of APS-I with a typical hemicellulase preparation. DPS containing 83% of D-galacturonic acid was able to be degraded by endo-polygalacturonase, but APS-I was not because of its highly was discussed on the basis of these results, periodate oxidation study.     

Production of Low-salt Soy Sauce with Enriched Flavor by Freeze Concentration Using Bacterial Ice Nucleation Activity

Soy sauce was frozen in the presence of ice nucleation-active Xanthomonas campestris cells at ?25°C, and the resulting frozen soy sauce was filtered through a 22-mesh screen to remove the ice and eutectic crystals of salt and water. The product retained well its original aroma and taste substances.