Sensory Descriptive Analysis of Soymilk A.V. TORRES-PENARANDA AND C.A. REITMEIER

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1 JFS: Sensory Descriptive Analysis of Soymilk A.V. TORRES-PENARANDA AND C.A. REITMEIER ABSTRACT: A descriptive analysis panel developed terms to describe aroma and flavor of soymilk. Panelists evaluated a commercial soymilk and soymilks processed with beans from the normal or lipoxygenase-free lines. Descriptors used to describe beaniness were raw as hexanal for flavor or aroma, grassy flavor, and sweet as green floral flavor. High correlations were found among these attributes. Judges were consistent in the use of terminology. Beaniness for the normal soymilk was described by the terms raw aroma as in hexanal and raw flavor as in hexanal, while beaniness for soymilk from the lipoxygenase-free line was described by the terms sweet as green floral and grassy flavors. Key Words: soymilk, flavor, descriptive analysis, lipoxygenase, storage Introduction THE FLAVOR OF SOYMILK IS THE MAIN FACTOR LIMITING ITS consumption by the western population. Development of the lipoxygenase-free line of soybeans [Glycine max (L.) Merr.] lacking 3 isozymes (SBL-1, SBL-2 and SBL-3) has been one of the most recent approaches toward the decrease of objectionable flavors. Sensory evaluation of the impact of any breeding or new processing technique on flavor of soy products requires the appropriate terminology as well as appropriate standards to evaluate objectionable flavors. Description of the sensory attributes characterizing aroma and flavor of soymilk has not been thoroughly investigated. Difficulty in describing the attributes has led to a variety of terms to describe the concept related to the flavor characteristics of soybean products. The term beany has been adopted as a general descriptor by many researchers (Kobayashi and others 1995; Liu 1997; Wilson 1995). More specific terms such as green, grassy, painty, rancid, astringent, and bitter have also been used to describe soymilk flavor (Liu 1997; Davies and others 1987; Nelson and others 1976; Oliver and others 1981). More recently, the terms raw beany and cooked beany have been developed by sensory descriptive panels from different nationalities (Torres-Penaranda and others 1998). A significant interaction between panelists and soybean line was found with the term raw beany. This may be attributed to cultural differences in perception or to some confusion in the use of the term beany as a single attribute. Beaniness might rather be a concept involving a variety of different attributes. Ishii and O Mahony (1987) indicated that the use of several standard stimuli may be necessary to define some sensory concepts. Flavor and aroma of soymilk are formed by a complex combination and interaction of chemical compounds, some of which have decreased in concentration with the development of new lines lacking 1 or several lipoxygenase isozymes. Despite some improvements, the lipoxygenase-free lines still contain many of the same chemical compounds responsible for the beany flavors (Kobayashi and others 1995). Aroma constituents of soymilk from normal soybeans and those lacking 2 or 3 of the lipoxygenases have been identified by gas chromatography (GC), and the GC effluents have been analyzed using an odor-sniffing system (Kobayashi and others 1995). Hexanal, which contributes to the odor of soymilk from the normal line, completely disappeared in the lipoxygenase-free line. This was believed to be the main reason for flavor improvement. Storage conditions on farms and elevators may be an additional source of change in flavor from these lines. Soybeans are stored in steel bins or concrete silos after harvest under conditions ranging from very low temperatures in winter to, in some cases, high temperatures in summer. According to Liu (1997), when temperatures are kept at ambient levels, moisture content of 13% or below is considered necessary to ensure stability of soybeans for several mo. Agrawal and Saddiqui (1973) reported that the optimum temperature and percentage seed moisture for storage of soybeans is 5 C and 11%, respectively. Lambrecht and others (1996) reported that soybean storage above 25 C and 50% relative humidity for 3 mo adversely affected the ph and solids content of soymilk produced from SBL-2 and SBL-3 null soybeans. Flavor changes of the lipoxygenase-free lines in storage have not been investigated. The objective of this research was to describe the beaniness concept using descriptive analysis. The judges were exposed to a variety of chemical compounds present in soymilk to find those chemicals suitable for standards. Using the flavor profile developed by the panel, we quantified the differences between soymilks processed with beans from the normal or the lipoxygenase-free lines from the 1996 and 1997 harvests. Materials and Methods Sample Preparation Soybean lines homogeneous for the 3 alleles controlling normal lipoxygenase and lines homogeneous for the 3 null alleles controlling absence of lipoxygenase were grown at the Agricultural Engineering and Agronomy Research Center of Iowa State Univ. during summers of 1996 and The lines were selected and harvested as described by Torres-Penaranda and others (1998). 352 JOURNAL OF FOOD SCIENCE Vol. 66, No. 2, Institute of Food Technologists

2 The treatments evaluated were a commercial soymilk powder and soymilks prepared from: normal line harvested in 1997, normal line harvested in 1996, lipoxygenase-free line harvested in 1997, and lipoxygenase-free line harvested in The commercial soymilk was Soyabean Drink, Instant (Guang XI Cereals Oils and Foodstuffs Import and Export Corporation, Wuzhou, Guang XI, China). The commercial soymilk was prepared by mixing 22 g soymilk powder, which contained 5% sugar cane, with 230 g water and blending at medium speed for 30 sec. Soymilk samples (10 ml) were poured into 4-oz white plastic coded cups with lids 2 h before serving to equilibrate to room temperature. The 1996 harvest of normal or lipoxygenase-free soybean lines was stored at 4 C for 15 mo and compared to the 1997 harvest of normal or lipoxygenase-free soybean lines. The moisture content of the soybeans was 10%. The soybeans from 1997 normal or lipoxygenase-free lines were stored at 4 C after harvest until soymilks were processed 3 mo later in January Soybeans from each treatment were soaked with tap water at room temperature for 12 h, drained, sorted, and washed with fresh water. A sample of 200 g of the hydrated beans was combined with 1.4 L tap water and ground for 1 min at medium speed with a blender. Soymilk was prepared by cooking the slurry on a stove with constant stirring for 7 min until the temperature reached 95 C and for 7 additional minutes at that temperature. Soymilk was filtered through 4 layers of cheesecloth and stored for 20 h at 5 C until 2 h before evaluation. Sensory evaluation Difference Tests A triangle test was performed for each of the 2 lines of soybeans in order to determine differences between soymilks processed from 1996 soybeans stored for 15 mo and beans from the 1997 harvest (stored 3 mo). Thirty-five panelists of Iowa State Univ. students and staff with previous experience evaluating soy products and familiarity with triangle tests performed the evaluation. Panelists received an orientation about the triangle test and the characteristics of the samples. Twenty ml of the samples were presented under red light in 4-oz white plastic cups coded with 3-digit numbers with lids on at room temperature. Panelists were instructed to examine the samples by smelling in 2 short sniffs, to swirl the sample in their mouths for 3 sec, and then swallow. The procedure was repeated with the other 2 samples. Panelists were instructed to select the odd sample. Tests for treatments from each line of soybeans were performed in 2 separate sessions to avoid fatigue. An equal number of the 6 possible combinations of both samples were prepared and presented in random order. Selection of Panelists for Descriptive Analysis Seven judges from Iowa State Univ. students and staff with previous experience evaluating soy products were pre-selected on the basis of interest, availability, and ability to articulate. Pre-screening questionnaires (Meilgaard and others1991) for aroma and flavor were initially used to screen individuals. Three additional screening tests were used: a basic taste test, an odor recognition test, and an intensity ranking test, as described by ASTM (1981a, 1981b) for flavor profile analysis. The interpretation of the screening tests was conducted according to ASTM (1981a, 1981b). Training of Panelists for Descriptive Analysis Panelists were trained for a period of 3 months in 1-hour sessions 3 times a week (36 h total). Judges were exposed to a variety of soymilks including the treatments to be evaluated as well as commercial brands. Initially, they were asked to evaluate sensory differences among samples and make a list of the descriptors for these differences. They considered aroma, flavor, mouthfeel, and aftertaste. The descriptors generated after initial consensus are listed in Table 1. With the purpose of finding descriptors to define the term beany, panelists were presented with a series of chemical compounds that have been found in the headspace of soymilks from normal or lipoxygenase-free lines (Kobayashi and others 1995) or recommended as standards by ASTM (1996). Hexanal, 1-hexen-1-ol, 1-octen-3-ol, 1-octen-3-one, (E)-2-nonenal, (E,E)-2,4-nonadienal, and (E,E)-2,4-decadienal (50 ppm in safflower oil) were each evaluated for aroma. Panelists were asked to indicate whether each compound was related to the sensation of beaniness in soymilk. During the process of defining standards and preliminary evaluation of the samples, the initial list of descriptors was modified. Panelists developed standards for each of the attributes. After further exposure to the products and more familiarity with standards, panelists arrived at consensus for the descriptors and the type of standards to use. Standards were presented in different concentrations until an appropriate dilution was found to anchor the ends of the scale for each attribute. Panelists also defined the type of scale to use for evaluation. A 10-cm line scale anchored from not detectable to intense was defined. Descriptive Analysis Four samples were used for evaluation by descriptive analysis, 1 commercial sample plus the 3 samples that were significantly different according to the triangle test. Sensory evaluation was conducted for 4 wk immediately following training using descriptive analysis procedures described by Meilgaard and others (1991) and ASTM (1981b). Samples were presented in partitioned booths under white light and at room temperature. Fifteen ml soymilk at room temperature were presented in clear plastic cups labeled with 3-digit random numbers. Cups were covered with lids to avoid volatilization of aroma compounds. Six of the attributes ( raw as hexanal aroma, starch as flour aroma, sweet as dairy caramelized aroma, raw as hexanal flavor, grassy flavor, and sweet as green floral flavor) were evaluated for the 4 samples in 8 replications, followed by 8 replications for evaluation of the other 6 attributes ( painty flavor, sweet as dairy caramelized flavor, metallic flavor, bitter flavor, astringency, and mouthcoating). Each replication was a separate session. The 4 samples were presented in a different random order for each judge during each replication. Judges recorded their responses on a 10-cm intensity line scale. Standards were available for comparison to samples; panelists were allowed to change their responses during evaluation. For aroma evaluation, judges smelled the samples in 2 short sniffs. Judges were instructed to swallow the samples and rinse with water between samples. Statistical Analysis Results of the difference test were analyzed using the statistical tables for the critical number of correct answers for Vol. 66, No. 2, 2001 JOURNAL OF FOOD SCIENCE 353

3 Table 1 Descriptors developed by panelists by initial consensus Term Description Aroma Starch flour Pasta cooked pasta sugar Dairy dry milk Dairy sour Flavor Vanilla Dairy Dairy cooked Oxidized Bitter Astringent Mouthfeel Coating Aftertaste Astringent Bitter evaporated milk soy, bean slurry, grass, green peas, earthy, raw beans, pungent sugar legume vanillin dry milk dissolved in water cooked milk aged oil coffee dry, clean tongue soy, green, grass, fresh vegetables, raw beans, pungent, unpleasant film coating the teeth dry, clean tongue coffee raw beans Table 2 Final list of sensory attributes used for description of soymilk samples Standard Descriptor Standard concentration for intense aroma Raw as hexanal Hexanal in safflower oil 50 ppm Other aromas Starch as flour Flour in water 1:10 as dairy Carnation sweetened caramelized condensed milk in water 1:3 flavor Raw as hexanal Hexanal in safflower (illustration by aroma) 50 ppm Grassy 1-hexen-1-ol (illustration by aroma) 50 ppm as green Fresh bean sprouts floral blended in water 1:60 Other flavors Painty Soybean oil aged for 4 yr 25 C, in safflower oil 1:10 as dairy Carnation sweetened caramelized condensed milk in water 1:3 Metallic Iron pills, not dissolved. Bitter Caffeine in water 0.1% Mouthfeel Astringent Alum in water 0.05% Mouthcoating Whole milk 100% difference tests from Meilgaard and others (1991). Descriptive analysis data were analyzed using analysis of variance, and principal component analysis (PCA) of the correlation matrices was performed for the significant attributes (SAS 1988). The interaction of judges with the soymilk factor was divided into 2 parts. One part was a linear component of the soymilk factor with judges and the other a deviations component of the soymilk factor with judges. The soymilk treatment means were used to define quantitative levels for determining linear and deviations components of interaction with judges. 354 JOURNAL OF FOOD SCIENCE Vol. 66, No. 2, 2001 Results and Discussion THE TERMINOLOGY FOR SOYMILK DESCRIPTION GENERATED by panelists by initial consensus consisted of a more common consumer language (Table 1), while the final list of terms (Table 2) developed after judges were extensively exposed to the samples and standards showed the use of more technical vocabulary. As mentioned by Civille and Lawless (1997), the fractionation of a perception into its component sensory impressions may be very difficult. The term beany that appears on the initial list was better defined later using more specific attributes. The use of the terms raw as in hexanal aroma or flavor, grassy flavor, and sweet as in green floral flavor shows the ability of trained panelists to develop a vocabulary that specifically describes a concept, generally known as beany. According to Lawless and Heymann (1998), the terms used for sensory description of products should be orthogonal, that is, not correlated to each other. There was a high positive correlation ( raw as hexanal aroma and raw as hexanal flavor, r = 0.995; raw as hexanal aroma and sweet as green floral flavor, r = 0.832; raw as hexanal flavor and sweet as green floral flavor, r = 0.79; raw as hexanal aroma and grassy, r = 0.783; raw as hexanal flavor and grassy, r = 0.745; sweet as green floral flavor and grassy flavor, r = 0.995) among the terms generated to describe beaniness. This shows the close interrelation of terms and therefore the difficulty of their definition by untrained panelists. The high correlation explains the fact that the principal component 1 accounted for most of the variation (89.7%). On the other hand, the interaction term for judge x soymilk for the terms related to beaniness was not significant (p > 0.05 for raw as hexanal aroma, raw as hexanal flavor, grassy, and sweet as green floral). This shows that judges were highly consistent with regard to the use of the terminology. According to Civille and Lawless (1997), a set of terms in descriptive analysis should enable differentiation among products. Beaniness for the normal soymilk was described by the terms raw aroma as in hexanal and raw flavor as in hexanal, while beaniness for soymilk from the lipoxygenase-free line was described by the terms sweet as green floral and grassy flavors (Table 3). The terms related to beaniness that were developed by a intensively trained panel can also be used as an illustration of the beany concept when evaluation is done by less-trained sensory panels using a series of terms and standards (multiple standards) to define the concept. The use of multiple standards to illustrate the beany concept would enable panelists with less training to evaluate soymilk with better criteria and a broader knowledge of the concept. Multiple standards would also facilitate judgement of beaniness when samples with different kinds of beaniness are tested, as occurs in soymilks from normal and lipoxygenase-free lines. There was no judge x soymilk interaction for any of the attributes with the exception of the term painty (p < 0.001). Even though the term painty was developed by the panelists, and they established the standard, judges were not consistent in the use of the term. The lack of consistency may be

4 Table 3 Sensory evaluation of commercial soymilk and soymilk processed from normal and lipoxygenase-free soybeans and lipoxygenase-free soybeans stored for 15 mo Z Soymilk Lipoxygenase- SensoryAttribute Commercial Normal Lipoxygenase free (1996) free (1997) stored 15 mo aroma Raw as hexanal 3 d 66 a 51 b 36 c Other aromas Starch as flour 82 a 29 c 41 b 22 c as dairy caramelized 92 a 22 b 13 c 7 c flavor Raw as hexanal 2 c 75 a 54 b 44 b Grassy 1 c 38 b 66 a 31 b as green floral 8 c 37 b 53 a 28 b Other flavors Painty 11 b 35 a 39 a 35 a as dairy caramelized 91 a 13 b 4 c 7 cb Metallic 0 d 37 b 54 a 21 c Bitter 0 d 18 c 32 b 47 a Mouthfeeling Astringent 7 b 37 a 42 a 41a Mouthcoating 58 a 13 c 29 b 30 b Z Main effect means within a row are responses of 7 panelists and 8 replications, 0 = not detectable to 100 = intense. P < 0.01 the result of the judges not understanding the term or that most of the samples were not significantly different for painty (Table 3). Replication was not a significant source of variation except for the attribute bitter (p = 0.04). Descriptive analysis showed significant differences across the soymilks for all the attributes (Table 3). The commercial soymilk was significantly different from the others for all the attributes. The attributes related to beaniness were perceived at a very low intensity in the commercial soymilk and lipoxygenase-free soymilk stored 15 mo. The aroma of the commercial soymilk was described by starch as flour and sweet as dairy caramelized. The commercial product had a very intense sweet as dairy caramelized flavor that was expected because sugar was contained in the soymilk powder. The absence of metallic flavor and the very low astringency may be due to the addition of sugar. The absence of bitterness may also be due to the addition of sugar that has been shown to mask bitterness (Roy 1997). The commercial soymilk had a lower painty flavor score as compared to the other soymilks and a higher intensity of mouthcoating. Differences between the commercial soymilk and those prepared in the laboratory may be due to several factors including the processing techniques, which were unknown for the commercial product. Drying and subsequent storage of the soymilk may have caused volatilization and change in the flavor compounds. Soymilks from normal and lipoxygenase-free soybeans were significantly different in all the attributes except for the painty and astringent flavors (Table 3). Soymilk from lipoxygenase-free soybeans had a lower intensity in the attributes raw as hexanal aroma and raw as hexanal flavor. The intensity of the raw as hexanal attributes was lower for the soymilks from the lipoxygenase-free soybeans than for normal soymilk as expected from the breeding modifications of this new soybean line. The decrease of these 2 attributes may have enhanced the grassy and sweet as green floral flavors. Aroma of the soymilk from the lipoxygenase-free soybeans was characterized by the term starch as in flour. Soymilk from the lipoxygenase-free soybeans was significantly higher in the metallic and bitter attributes and equally painty as compared to the soymilk from the normal soybean line. There were significant sensory differences (17 correct judgments out of 35 in the difference test, p < 0.05) between soymilks from the 1996 harvest (15-mo storage) and the 1997 harvest (3-mo storage) for the lipoxygenase-free line. In contrast to the lipoxygenase-free line, soymilks from the normal line did not exhibit sensory differences (7 correct judgments of 35, p < 0.05) between the 1996 (15 mo) and 1997 (3 mo) treatments. These changes in flavor may be due to the fact that the soybeans were produced in different years or that 3- mo storage is sufficient time for lipoxygenase-mediated changes to occur. Lambrecht and others (1996) found that the null lines lacking lipoxygenase-2 and lipoxygenase-3 were more stable to adverse storage treatments than the normal soybeans with regard to changes in ph and solids content of the soymilks produced from the stored beans. They found that the higher the temperature and relative humidity, the greater the reduction in solids of the soymilk. No changes in flavor were analyzed in that study. Soymilk from lipoxygenase-free lines processed after the storage of the beans 15 mo at 4 C exhibited changes in aroma and flavor (Table 3). The intensity of 2 of the aroma attributes decreased significantly, including the starch as flour. The raw as hexanal flavor remained the same as the 3-mo storage treatment for lipoxygenase-free soymilk. The grassy and sweet as green floral flavor attributes decreased to the same intensity as the soymilk from the normal line. The painty flavor and the attributes related to mouthcoating underwent no changes in storage for lipoxygenase- Vol. 66, No. 2, 2001 JOURNAL OF FOOD SCIENCE 355

5 free soybeans. There was a decrease in the metallic attribute and a significant increase in bitterness of soymilk processed after storage of lipoxygenase-free beans. Glycosides such as saponins and isoflavones (Okubo and others 1992) and by-products of fatty-acid oxidation not mediated by lipoxygenase also contribute to soybean flavor. Conclusions THE USE OF THE TERM BEANY FOR SOYMILK CHARACTERIStics comprises a series of different related attributes and must be properly defined when sensory evaluation of soymilk is conducted. Significant differences between the commercial soymilk and the soymilk processed from the lipoxygenase-free line suggest that sugar addition to soymilk would produce important desirable changes in flavor, especially in decrease of bitterness, astringency, and the attributes related to beaniness. Soymilk processed with lipoxygenase-free soybeans exhibited changes in the type of perceived beaniness in comparison to the soymilk from normal soybeans. Despite a decrease in the intensity of the grassy and sweet as green floral flavor attributes, changes in flavor for soymilks processed from stored lipoxygenasefree beans are important considering that the storage treatment significantly increased bitterness. References Agrawal PK, Saddiqui MN Influence of storage temperature and seed moisture on germination, free fatty acids content and leaching of sugars of soybean seeds during storage. Seed Res 1: ASTM. 1981a. Guidelines for the selection and training of sensory panel members: ASTM Special Technical Publication 758. Philadelphia. p ASTM. 1981b. Manual on sensory testing methods: ASTM Special Technical Publication 434. Philadelphia. p ASTM Aroma and flavor lexicon for sensory evaluation: terms, definitions, references, and examples. Civille GV, Lyon BG, editors. West Conshohocken, Pa.: Data Series Publication DS 66. Civille GV, Lawless HT The importance of language in describing percep- tions. In: Gacula MC, editor. Descriptive sensory analysis in practice. p Davies CS, Nielson SS, Nielsen NC Flavor improvement of soybean preparation by genetic removal of lipoxygenase-2. J Amer Oil Chem Soc 64(10): Ishii R, O Mahony M Taste sorting and naming: Can taste concepts be misrepresented by traditional psychophysical labeling systems? Chem Senses 12: Kobayashi A, Tsuda Y, Hirata N, Kubota K, Kitamura K Aroma constituents of soybean [Glycine max (L.) Merrill] milk lacking lipoxygenase isozymes. J Agric Food Chem 43: Lambrecht HS, Nielsen SS, Liska BJ, Nielsen NC Effect of soybean storage on tofu and soymilk production. J Food Qual 19: Lawless HT, Heymann MC Sensory evaluation of foods: Principles and practices. New York: Chapman & Hall, p Liu K Soybeans: Chemistry, technology and utilization. New York: Chapman & Hall. p Meilgaard M, Civille GV, Carr BT Sensory evaluation techniques. 2nd ed. Boca Raton, Fla.: CRC Press, Inc. p Nelson AI, Steinberg MP, Wei LS Illinois process for preparation of soymilk. J Food Sci 41: Okubo K, Iijima M, Kobayashi Y, Yoshikoshi M, Uchida T Components responsible for the undesirable taste of soybean seeds. BioSci Biotech BioChem 56(1): Oliver A-L, Hsieh A-H, Stephen SC Isolation and identification of objectionable volatile flavor compounds in defatted soybean flour. J Food Sci 47: Roy G General ingredient or process approaches to bitterness inhibition and reduction in foods and beverages. In: Roy G, editor. Modifying bitterness: Mechanism, ingredients and applications. Valhalla, N.Y.: Technomic Publishing Co. P SAS Institute SAS/STAT user s guide. Version rd ed. Cary, N.C.: SAS Institute, Inc. Torres-Penaranda AV, Reitmeier CA, Wilson LA, Fehr WR, Narvel JM Sensory characteristics of soymilk and tofu made from lipoxygenase-free and normal soybeans. J Food Sci 63 (6): Wilson LA Soy foods. In: Erickson DR, editor. Practical handbook of soybean processing and utilization. Champaign, Ill., and St. Louis, Mo.: AOCS Press and United Soybean Board. P Ms Journal Paper No of the Iowa Agriculture and Home Economics Experiment Station, Ames, IA, Project No and supported by Hatch Act and State of Iowa funds. The authors wish to acknowledge the support of Dr. Walt Fehr, Dept. of Agronomy and the Center for Designing Foods to Improve Nutrition at Iowa State Univ., the Iowa Soybean Promotion Board, and the Univ. of Costa Rica. Authors are with the Dept. of Food Science and Human Nutrition, Iowa State Univ., Ames, IA Direct correspondence to Dr. Cheryll Reitmeier ( creitmei@iastate.edu). 356 JOURNAL OF FOOD SCIENCE Vol. 66, No. 2, 2001