FUNCTION IN FOODS

III. FUNCTION IN FOODS

A. Basic Qualities

1. Umami Taste The theory of four basic tastes (sweet, sour, salty, and bitter) was proposed by a German

psychologist (Henning, 1916), and was accepted for a long time without sufficient scien- tific data to support it. He explained that all tastes experienced could be made up from the mixture of the four basic tastes, located at the corners of a tetrahedron and located

‡ Preparation of ninhydrin solution: Dissolve 136 g of sodium acetate trihydrate and 25 mL of acetic acid in water and adjust to 250 mL. Add to this solution 750 mL of ethyleneglycol monomethyl ether, pass nitrogen gas for 20 min with stirring. Add 20 g of ninhydrin, pass nitrogen gas for 15 min, and stir well to clear solution. Add 1.7 mL of titanium trichloride, pass nitrogen gas for 10 min, stir well to completely clear solution, press ‡ Preparation of ninhydrin solution: Dissolve 136 g of sodium acetate trihydrate and 25 mL of acetic acid in water and adjust to 250 mL. Add to this solution 750 mL of ethyleneglycol monomethyl ether, pass nitrogen gas for 20 min with stirring. Add 20 g of ninhydrin, pass nitrogen gas for 15 min, and stir well to clear solution. Add 1.7 mL of titanium trichloride, pass nitrogen gas for 10 min, stir well to completely clear solution, press

Multidimensional scaling analyses of human sensory tests demonstrated that the umami taste is located outside the tetrahedron of the traditional four basic tastes, and the taste quality is distinctly different from those of the other basic tastes (Yamaguchi, 1987). The taste physiological studies in mice also supported that umami taste quality is different from those of the other basic tastes (Fig. 4) (Ni- nomiya and Funakoshi, 1987).

The taste quality of umami is not produced by mixing any of the other four basic tastes (Yamaguchi, 1987). Recent electrophysiological studies in primates suggest that a single taste nerve in the chorda tympani is responsible for the taste stimuli of umami substances such as MSG and GMP (Hellekant and Ninomiya, 1991). A study of the monkey cortex demonstrates that umami achieves independence as a taste quality at higher levels of neural processing (Rolls et al., 1994).

Electrophysiological studies suggest that the taste bud receptor site for glutamate is different from those for the traditional four basic tastes (Ohno et al., 1984; Kuma- zawa and Kurihara, 1990). These electrophysiological data are supported by the

Figure 4 Locations of each of the 17 test stimuli in three-dimensional space obtained through multidimensional scaling based on the data obtained from glossopharyngeal fibers in mice. Suc, sucrose; Sac, Na saccharin; Fru, fructose; Glu, glucose; Mal, maltose; Gly, glycine; Gln, L-gluta- mine; Arg, L-arginine HCl; MAG monoammonium L-glutamate; Asp, monosodium L-aspartate; MSG, monosodium L-glutamate; IMP, disodium 5 ′-inosinate; GMP, disodium 5′- guanylate; M⫹G,

a mixture of 0.1M MSG with 0.5mM GMP; Na, NaCl; H, HCl, Q, quinine-HCl. (From Ninomiya

Table 4 Detection Thresholds for Five Taste Substances (in g/dL)

Quinine MSG

0.000049 Source : Yamaguchi and Kimizuka (1979).

most recent molecular biological study on glutamate receptor sites in taste bud cells (Chaudhari et al., 1994).

Umami is one of the primary tastes, independent of the four traditional basic tastes.

2. Taste Thresholds

a. Glutamate. The detection threshold for MSG was as low as 0.012 g/100 mL, or

6.25 ⫻ 10 ⫺4 M. It was higher than that of quinine sulfate or tartaric acid, lower than that of sucrose, and almost the same as that of sodium chloride at isomolar concentrations

(Table 4). Several umami substances have lower thresholds than MSG (Yamaguchi et al., 1979).

b. Nucleotides. The threshold values of IMP and GMP are 0.025 and 0.0125 g/100 mL, respectively (Table 5). The taste threshold for 50:50 blends of GMP and IMP has been reported to be 0.0063%. When they were used in combination with 0.8% MSG, however, the resulting threshold was lowered to 0.000031%, which represents a dramatic synergistic effect.

3. Taste Intensity The relationship between MSG concentration and the taste intensity of MSG is found

to follow a straight line. The slope for MSG is not as steep as for the four basic tastes ( Fig. 5 ) (Yamaguchi, 1979). Moreover, the taste intensity of IMP increases hardly at all, even when its concentration is increased considerably.

The relative taste intensities of amino acid–based derivatives and nucleotides are given in Tables 6 and 7 , respectively.

Table 5 Taste Thresholds of Basic Taste Substances

Absolute threshold a

Quinine sulfate Glutamate Solvent

Sodium chloride

Tartaric acid

(bitter) (umami) Pure water

Sucrose (sweet)

(salty)

(sour)

6.25 ⫻ 10 ⫺7 M 6.25 ⫻ 10 ⫺4 M (8.6 ⫻ 10 ⫺2 %)

6.25 ⫻ 10 ⫺7 M — 5 ⫻ 10 ⫺3 M

IMP, 5 ⫻ 10 ⫺3 M 1.25 ⫻ 10 ⫺3 M

(2.0 ⫻ 10 ⫺4 %) a Significant at 5% level.

Figure 5 Relationship between concentration and taste intensity.

B. Taste Synergism

1. The Synergistic Effects of Umami Substances Figure 6 shows that taste intensity of mixtures (IMP and MSG) increases exponentially

with their concentration and that the degree of synergism depends upon the ratio of IMP to MSG.

Figure 7 shows the relationship between the intensity of umami and the proportion of IMP in the mixture of MSG and IMP. Because the umami intensities of the samples on both extremes are very weak and almost the same, the curve would have been horizon- tally linear if the synergistic effect had been absent. The symmetric curve illustrates this remarkable synergistic effect. In this curve, the intensity of umami at its maximum is equivalent to that of 0.78 g/100 mL of MSG alone. The mixture is thus 16 times as strong as that of MSG. This amplification factor is concentration dependent and becomes higher with increasing concentration.

The synergistic effect between MSG and IMP can be expressed by means of the following simple equation:

y ⫽ u ⫹ 1200uv (1)

Table 6 Taste Intensities of Amino Acid Based Derivatives Relative umami

Momosodium L-glutamate ⋅H 2 O

Monosodium DL-threo- β-hydroxy glutamate ⋅ H 2 O

Monosodium DL-homocystate ⋅H 2 O

Monosodium L-aspartate ⋅H 2 O

Monosodium L- α-amino adipate ⋅ H 2 O

L-Tricholomic acid (erythro form) a 5–30 L-Ibotenic acid a 5–30

a From Terasaki et al. (1965a,b).

Table 7 Taste Intensities of Nucleotides Having Synergistic Effect on Glutamate Taste

Relative potency of umami Substance (disodium salt)

( β) 5 ′-Inosinate ⋅ 7.5 H 2 O

1 5 ′-Guanylate ⋅ 7 H 2 O

2.3 5 ′-Xanthylate ⋅ 3 H 2 O

0.61 5 ′-Adenylate

0.18 Deoxy-5 ′-guanylate ⋅ 3 H 2 O

0.62

2.3

2-Methyl-5 ′-inosinate ⋅ 6 H 2 O

2.3

2-Ethyl-5 ′-inosinate ⋅ 1.5 H 2 O

3.6

2-Phenyl-5 ′-inosinate ⋅ 3 H 2 O

8.0

2-Methylthio-5 ′-inosinate ⋅ 6 H 2 O

2-Ethylthio-5 ′-inosinate ⋅ 2 H 2 O

7.5 2-Ethoxyethylthio-5 ′-inosinate a 13 2-Ethoxycarbonylethylthio-5 ′-inosinate a 12 2-Furfurylthio-5 ′-inosinate ⋅ H 2 O a 17

2-Tetrahydrofurfurylthio-5 ′-inosinate ⋅ H 2 O a 8 2-Isopentenylthio-5 ′-inosinate (Ca) a 11 2-( β-Methallyl)thio-5′-inosinate a 10 2-( γ-Methallyl)thio-5′-inosinate a 11

2-Methoxy-5 ′-inosinate ⋅ H 2 O

4.2 2-Ethoxy-5 ′-inosinate a 4.9 2-i-Propoxy-5 ′-inosinate a 4.5 2-n-Propoxy-5 ′-inosinate a 2

6.5

2-Allyloxy-5 ′-inosinate (Ca) ⋅ 0.5 H 2 O

3.1

2-Chloro-5 ′-inosinate ⋅ 1.5 H 2 O

2.3

N 2 -Methyl-5 ′-guanylate ⋅ 5.5 H 2 O

2.4

N 2 ,N 2 -Dimethyl-5 ′-guanylate ⋅ 2.5 H 2 O

0.74

N 1 -Methyl-5 ′-inosinate ⋅ H 2 O

1.3 N 1 -Methyl-2-methylthio-5 ′-inosinate

N 1 -Methyl-5 ′-guanylate ⋅ H 2 O

8.4

6-Chloropurine riboside 5 ′-phosphate ⋅ H 2 O

3.4

6-Mercaptopurine riboside 5 ′-phosphate ⋅ 6 H 2 O

2-Methyl-6-mercaptopurine riboside 5 ′-phosphate ⋅ H 2 O

7.9 2 ′,3′-o-Isopropylidene 5′-inosinate

2-Methylthio-6-mercaptopurine riboside 5 ′-phosphate ⋅ 2.5 H 2 O

0.21 2 ′,3′-o-Isopropylidene 5′-guanylate

0.35 a From Imai et al. (1971).

Source : Yamaguchi et al. (1971).

where u and v are the respective concentrations of MSG and IMP in the mixture, and y is the equi-umami concentration of MSG alone (Yamaguchi, 1967).

The synergistic effect can be demonstrated between any combination of substances in Tables 6 and 7, and the intensity of umami can also be expressed by an equation essentially equivalent to Eq. (1). The taste intensities of all substances in Table 6 are always proportional to that of MSG. Therefore, u ′ g/100 mL of any substance cited in

Figure 6 Taste equivalency of mixture of IMP and MSG to MSG alone. t represents IMP content (percent) in mixtures.

ity; values for various substances are listed in Table 6 . On the other hand, the strength of umami taste of nucleotides in Table 7 is consistently proportional to that of IMP. Hence, v ′ g/100 mL of any nucleotide is replaceable with βv′ g/100 mL of IMP. The constants β for all nucleotides are given in Table 7. Therefore, the umami intensity of the mixture of any combination of substances in Tables 6 and 7 can be calculated by substituting αu′ for u and βv′ for v. Because the interrelationships within each series of substances are additive, the intensity of umami of the mixture of two or more different L- α-amino acids

Figure 8 Three-dimensional configuration for five taste stimuli.

and two or more nucleotides can be calculated by substituting the product sums ∑α i u i and ∑β j v j for u and v, respectively, in Eq. (1).

2. The Synergistic Action of Umami in Foods Multidimensional analysis has shown umami to be present in the taste of natural foods

(Yamaguchi, 1987). Umami is definitely located outside the tetrahedron of the four basic tastes and is an independent basic taste (Fig. 8). The broths made from animal and fish stocks fall outside the area of the four basic tastes and lie nearer to umami (Fig. 9). This demonstrates that umami is a vitally important element in broth taste composition.

In contrast, broth made from vegetables also contains umami, but some of the taste factors are sweetness or sourness. Thus these broths are distributed widely over the five

Figure 10 Three-dimensional configuration for vegetable stocks and the five taste stimuli.

taste areas (Fig. 10). However, if a small amount of IMP is added, the tastes of all the broths move in the direction of umami (Fig. 11). This shows the synergistic effect of umami that is brought into existence between the glutamate contained in the vegetables and the added inosinate (Yamaguchi et al., 1967).

C. Taste Physiological Data

The biological significance of the chemoreception of amino acids was reviewed by Kawa- mura (1987).

Figure 11 Three-dimensional configuration for vegetable stocks with 0.005% disodium inosinate

Amino acids are one of the most important classes of nutrients and are potent attract- ants for many living organisms; organisms as diverse as bacteria, protozoa, annelids, gas- tropods, crustaceans, insects, fish, amphibians, and mammals all have chemoreceptors sensitive to amino acids. The treatment of carp olfactory epithelium, bullfrog tongue, and rat tongue with pronase E obliterated responses to amino acids in these animals; this showed that receptor molecules for amino acids, including glutamate, are protein. Cross- adaptation experiments utilizing various combinations of amino acids in olfactory and taste systems of the carp explored the receptor sites for various amino acids. Results showed that there are multiple types of receptors corresponding to different amino acids and that the receptor for glutamate is different from those for other amino acids. Evidence suggesting that there is a specific receptor protein for glutamate in chemoreceptor mem- branes is consistent with reports on receptor potentials in bacteria.

Taste buds on the front of the tongue receive inputs from the chorda tympani (CT) branch of the facial nerve, whereas those on the posterior part of the tongue receive inputs from the glossopharyngeal (GP) nerve. Single-unit nerve responses to glutamate and 5 ′- nucleotides were recorded in the taste pore, chorda tympani nerve, glossopharyngeal nerve, geniculate ganglion, petrosal ganglion, and cortical neurons in the rat, hamster, mouse, cat, goat, and dog. Results varied somewhat depending on the nerve fiber recorded and the species of experimental animal studied. For example, although some units of the cat geniculate ganglion responded strongly to nucleotides, these same units failed to respond to MSG. In the hamster, MSG-sensitive units of cortical neurons also responded to NaCl; however, the responses of MSG-sensitive units recorded within the hamster taste pore were independent of the responses to any of the four basic taste stimuli.

Differences in responsiveness of CT and GP nerves to various taste stimuli have been observed in mice. Some of the glossopharyngeal nerve fibers of the mouse are espe- cially sensitive to MSG, and these fibers are only slightly sensitive to NaCl or to sucrose. Patterns of suppression of licking in denervated mice suggested that the CT nerve is impor- tant for the sodium component of MSG and for discrimination among sugars, whereas inputs from the GP nerve play an important role in discrimination between salty and umami taste components of MSG. Patterns of suppression of licking across various taste stimuli also showed that MSG and GMP elicit similar patterns but that MSG shows the highest similarity to NaCl, especially at high concentrations.

The mechanism of the taste synergism between glutamate and nucleotides has been examined both electrophysiologically and biochemically. Recordings from the rat CT nerve demonstrate the remarkable synergism between MSG and purine-based 5 ′-nucleo- tides, which is consistent with and corroborates the psychophysical data. Analysis of data from the rat has suggested that the presence of 5 ′-nucleotides induces an increase in the affinity (i.e., strength of binding) of glutamate for the receptor sites while not increasing

the amount of glutamate bound. However, measurements of binding of L-[ 3 H]glutamate to bovine circumvallate papillae showed that in this system the presence of nucleotides causes a remarkable increase in the maximal capacity of glutamate binding to the papillae without affecting binding affinity. The results were explained in terms of allosteric regula- tion.

In the dog, large synergism between MSG and GMP or IMP, similar to that in humans, was observed in the chorda tympani nerves. Amiloride, an inhibitor of Na re- sponse, suppressed the response to MSG similarly to that observed with NaCl, suggesting that the response to MSG is primarily a salt response. Amiloride did not, however, suppress In the dog, large synergism between MSG and GMP or IMP, similar to that in humans, was observed in the chorda tympani nerves. Amiloride, an inhibitor of Na re- sponse, suppressed the response to MSG similarly to that observed with NaCl, suggesting that the response to MSG is primarily a salt response. Amiloride did not, however, suppress

Brain neuronal responses to umami taste stimulation were recorded in the rat and monkey. In the nucleus of the solitary tract of the rat, there were no specific responses to umami taste substances. These results were similar to those observed with the CT nerves. Using macaques, recordings were made from 190 taste responsive neurons in the taste cortex and adjoining or orbitofrontal cortex taste area, single neurons were found that were tuned to respond best to MSG. Across the population of neurons, the respon- siveness to MSG was poorly correlated with the responsiveness to NaCl, suggesting that the representation to MSG was clearly different from that of NaCl. In addition, cluster analysis and multidimensional scaling analyses confirmed that the neuronal representation of MSG fell outside the space defined by sweet, salt, bitter, and sour. Therefore, in primate taste cortical areas, MSG is as well represented as are the tastes produced by glucose, NaCl, HCl, and quinine (Baylis and Rolls, 1991).

D. Influence on Food Consumption

1. Taste Preferences and Nutritional Status The perinatal human infant can communicate feelings and emotions by a repertoire of

nonverbal communications of facial expressions and actions. When a sour or bitter taste or an unflavored vegetable soup is given, infants respond with expressions and actions indicating displeasure and dissatisfaction. If glutamate is added to the unflavored vegetable soup, however, infants not only detect its presence but also indicate, with their facial expression, enjoyment of the taste in a manner similar to their responses to sweet solutions (Steiner, 1987).

Similar behavioral manifestations have been observed in rats given dietary glutamate (Torii et al., 1983). There is a reproducible correlation between protein level in the diet and selection of taste. If the protein content of the taste material is insufficient for growth, rats do not choose glutamate even when it is available. The preference for glutamate returns when the protein level of the diet is sufficient for rat growth. Even if the quantity of protein is sufficient, the rats do not grow, and they choose sodium chloride rather than glutamate (as in the case of the low protein diet) when lysine, one of the essential amino acids, is deficient. When lysine is supplemented and the diet becomes well balanced both in quality and in quantity, the rats begin to grow and their preference for glutamate appears. It has therefore been suggested that the change in taste preference could be a reaction to an altered physiological state depending upon the nutritional status of the animal (Torii et al., 1985).

To relate neural activity to preference for amino acids, the electrical responses of lateral hypothalamus (LHA) neurons were observed during the application of amino acids to the tongue. The data demonstrated that during an essential amino acid deficiency, more LHA neurons responded to the taste of the deficient amino acid (Tabuchi et al., 1991).

Spontaneously hypertensive rats (SHR) have a tendency to ingest a large amount of sodium chloride and become hypertensive. However, sodium intake declined as the protein content in the diet was increased. When an umami substance (MSG and MSG plus GMP) was available as an aqueous solution, rats selected it, and a drastic decrease in total sodium intake—even up to 70%—was observed when the solutions were offered in conjunction with the diet of higher protein content. Although the predisposition of SHR Spontaneously hypertensive rats (SHR) have a tendency to ingest a large amount of sodium chloride and become hypertensive. However, sodium intake declined as the protein content in the diet was increased. When an umami substance (MSG and MSG plus GMP) was available as an aqueous solution, rats selected it, and a drastic decrease in total sodium intake—even up to 70%—was observed when the solutions were offered in conjunction with the diet of higher protein content. Although the predisposition of SHR

2. Physiological Responses to Umami Taste Sham-feeding experiments with dogs have shown that oral stimulation by MSG produces

significant and dose-dependent stimulation of both pancreatic flow and protein output in conscious dogs (Naim et al., 1991).

In the rat, the effects on metabolic parameters such as total and background metabo- lism and respiratory quotient (RQ) of umami-flavored meal was compared with an unfla- vored one. MSG added to the meal modified the metabolic parameters closer to those for protein metabolism (Viarouge et al., 1992).

These studies along with a nutritional study by Torii et al support that umami taste stimuli could be a marker of protein intake.

3. MSG and Sodium in the Diet MSG contains 12.3% sodium, or one-third that of common table salt; as the usual MSG

use level is around one-tenth that of salt, the contribution of MSG to dietary sodium is usually one-twentieth to one-thirtieth of that of NaCl. In general, as the salt level in food is reduced, there is a corresponding reduction in food acceptability. By using a small amount of MSG, more than 30% of sodium content may be reduced while maintaining

a very palatable and acceptable level of taste (Yamaguchi and Takahashi, 1984b; Chi and Chen, 1992; Altug and Demirag, 1993).

A recent report states that calcium glutamate may be used to improve the acceptabil- ity of sodium free diets of children suffering kidney disease (Bellisle et al., 1992).

4. Role of Umami in Food The role of umami substance in food tastes has been examined with snow crab extract,

which was analyzed for amino acids, nucleotides and related compounds, organic bases, sugars, organic acids, and minerals. The extract from the meat of snow crab was recon- structed with about 40 pure chemicals in accordance with the analytical data. A series of organoleptic tests of the reconstructed extract, using omission test methods, revealed that the unique flavor of boiled crab is derived from a rather limited number of components

(Gly, Ala, Arg, Glu, IMP, NaCl, KH 2 PO 4 ). The relationship between the taste of a compos- ite extract and characteristics in snow crab meat is shown in Table 8. Fairly high correla-

Table 8 The Relationship Between the Taste of a Composite Extract and Characteristics in Snow Crab Meat

Umami (Gly ⫹ Ala)

Sweet

Salty

(Glu ⫹ IMP) Continuity

(NaCl ⫹ K 2 HPO 4 )

Seafoodlike flavor

Overall preference

* P ⬍ 0.05, **P ⬍ 0.01.

tions were observed between umami and five descriptors of flavor characteristics and over- all preference. Thus when umami was omitted, not only did the continuity, complexity fullness, and mildness diminish, but the characteristic taste of crab itself disappeared, and so did the overall preference. This fact suggests the function of umami substances in flavor-enhancing properties (Konosu and Yamaguchi, 1987).

The role of umami taste in meat flavor was also studied (Kato and Nishimura, 1987). The increase in strength of umami and brothy tastes obtained through conditioning is more evident in pork and in chicken than in beef, which contains less glutamate. The addition of glutamate or glutamate plus IMP to broths enhanced the umami and brothy tastes and showed that umami plays an important role in meat taste.

There is large natural variability in the free glutamate levels in cheese of different varieties. It has been well established that free amino acids content of cheese, including glutamic acid, increases during maturation. The free glutamic acid content of cheese is used as an indicator of maturation (Weaver, 1978) Similarly, the free glutamic acid content of tomato increases during ripening. Glutamate value in green tomato increases during the ripening period (Inaba, 1980; Skurray, 1988). The free glutamate content of ripened tomato is about seven times as high as unripend green tomato.

5. Hedonic Functions and Self-Limiting Properties Psychometric studies on aqueous solutions of the four basic tastes revealed that three

(salty, bitter, and sour) are rated as unpalatable over a wide concentration range in that they received unpleasantness ratings. Only sweetness was given a pleasantness rating. In

a similar fashion to salty, sour, or bitter stimuli, umami (MSG in aqueous solution) also had an unpleasantness rating or was rated neutral in acceptability at all concentrations studied (Yamaguchi and Takahashi, 1984a).

Of additional importance is the fact that there is an optimal concentration for MSG added to food. Beyond this most palatable concentration, the palatibility of food decreases. Thus the use of MSG is self-limiting in that overuse decreases palatability ( Fig. 12 ) (Yama- guchi and Takahashi, 1984a).