Flavor Potentiation: The Preparation, Structure,

and

Chemical Interactions of Monosodium L-Glutamate

Chemistry Seminar paper presented by David R. Gardner
University of Guam, Mangilao, Guam. November 18, 1977.

Summary:

Although monosodium glutamate (MSG) was first prepared in a German chemistry laboratory in 1867, its flavor enhancing properties were not discovered until investigations of traditional Oriental spices led to its isolation as the active agent. Since this accomplishment in 1908, and the immediate commercial production of MSG, food flavor research has become an important application of chemistry. This paper covers the chemical aspects of monosodium glutamate's preparation, structure, and some of its synergistic relationships with other flavor potentiators. Research in MSG's biochemical activities and its possible implications in human biochemistry are also discussed.

Preparation

Monosodium glutamate (MSG) was first prepared in a German chemistry laboratory in 1867 (Benard, 1971) but due to stringent laboratory safety precautions its flavor and flavor-enhancing properties remained obscure until 1908. At this time, the investigation of the chemical properties of traditional Japanese spices led Kikunae Ikeda at the University of Tokyo to discover MSG's flavor potentiating characteristics (Kuninaka, 1967). Ikeda chemically fractionated extract from the edible Japanese seaweed, kombu or "sea tangle", and isolated the sodium salt of L-glutamic acid as the active flavor-potentiating ingredient.
 
Figure: Monosodium L-Glutamate (MSG) (Pryde, 1973)

Occuring naturally in cereal, meat, poultry, fish, and other proteinaceous foods (Smith and Minor, 1974), MSG was produced on a large scale in Japan as a food-flavor-enhancing additive for the general consumer immediately after its taste effect was discovered (Kuninaka, 1967). Production of MSG was made relatively simple due to the availability of proteinaceous glutamic acid sources such as soybeans, sugar-beet wastes (Steffen's wastes), wheat and corn (Markley, 1950); and the high insolubility of glutamic acid hydrochloride in HCL solution (Greenstein and Winitz, 1961). Glutamic acid hydrochloride was so highly insoluble in HCl solution as compared to the high solubility of all the other amino acid hydrochlorides that it was readily isolated from protein hydrolysates.
 

Figure: Glutamic acid--shown in its zwitterion (dipolar ion) form, the proton being attached to the amino group. (Giese, 1973)

Glutamic acid occurs in all proteins, and for the best yield, proteins with the highest glutamic acid content are used in the preparation of MSG. Wheat gluten, for example, contains approximately 36 percent glutamic acid (Meyer, 1960). In general, the protein is degraded  to the amino acid stage by prolonged hydrolysis with boiling concentrated acids or bases (Markley, 1950), or with enzymes or microbiological agents (Smith and Minor, 1974). Once the amino acids have been separated, excess HCl is added. When the solution has been allowed to stand for a few hours, crystals of glutamic acid hydrochloride are removed from the mother liquour by filtration. Dissolving this crude hydrochloride in water and adjusting the pH to 3.2 causes the pure glutamic acid to slowly chrystallize. This product is recovered by filtration and is neutralized to a pH of 7 with either NaOH or Na2CO3, resulting in the sodium salt of glutamic acid, which can then be decolorized and recrystalized (Meyer, 1960).
 

Figure: The conversion of protein to monosodium L-glutamate. (Meyer, 1960).

Specific procedure for isolation of L-glutamic acid from a protein hydrolysate obtained from Greenstein and Winitz (1961) is as follows:

Reflux 300ml HCl and 100g of gluten flour for 6 hours. The resulting solution is allowed to cool to about 80 degrees centigrade, 20g Norit is added and the mixture is again brought to boil for a few minutes. The solution is then cooled to room temperature and is filtered. The residue is washed with 50ml of 5N HCl. The combined filtrate and washings are evaporated under aspiration to about 100ml. The condensate is chilled to about 0 degrees centigrade, and HCl gas is admitted to saturation. On standing for several hours at -5 degrees centigrade, crystals of L-glutamic acid hydrochloride separate. This resulting mixture is filtered, the mother liquour is evaporated under aspiration to about half its original volume, and a second much smaller crop is obtained. The combined crude products are then dissolved in about 50ml of hot water, about 1g of Norit is added and the mixture is brought to boil for a few minutes. The mixture is filtered, the filtrate chilled to about 0 degrees centigrade,  and HCl gas is admitted to saturation. After standing for several hours at -5 degrees centigrade, the crystals of pure L-glutamic acid hydrochloride are filtered, washed with 20ml of ice cold 5N HCl, and dried in a vacuum desicator over P2O5 and NaOH. The yield here is about 26g of the hydrochloric salt with a melting point of 208 to 209 degrees centigrade.

The hydrochloric salt is converted to free L-glutamic acid by dissolving 36.7g of the salt in 200ml of hot water, adding 20g of redistilled aniline, and quickly cooling the solution to room temperature. Ethanol is added to 80 percent while stirring and the mixture is allowed to stand at 5 degrees centigrade for 20 hours. The crystals of pure L-glutamic acid are filtered over suction, washed with 95 percent ethanol until chloride-free, and then are finally washed with ether. When dry, the yield should be about 27g of L-glutamic acid with a melting point of 211 to 213 degrees centigrade.

Final neutralization of this product with NaOH or Na2CO3 to a pH of 7 results in the formation of monosodium L-glutamate.

There are other ways to prepare MSG by chemical process only, but they are tedious, involve expensive chemicals, and generally result in low yields.

Enzymatic and microbiological processes are the most used in the MSG industry, and their specific techniques, apparatus, and reaction agents are well-guarded secrets. It is assumed that the proteins are hydrolyzed by the enzymes or microorganisms in a type of "fermentation" process and that the glutamic acid is converted to the hydrochloric salt and finally MSG by the previously mentioned conventional procedures.

An interesting feature of the preparation of MSG is that it is reversible. In other words, glutamic acid can be isolated from MSG. Using Ajinomoto as a starting source of MSG, Greenstein and Winitz (1961) described this method of isolation. Ajinomoto MSG is brought into suspension in about four times its weight of hot water. Norit is added and the mixture is filtered. To the clear solution brought to the boiling point, 4N HBr is added to a pH of 3.0 and the solution is cooled. The solution is allowed to stand at 5 degrees centigrade for several hours. The crystalline L-glutamic acid is filtered and washed successively with ice water, ethanol, and ether. This product should now be halogen-free. The yield from 100g of Ajinomoto containing some 85g of monosodium L-glutamate may be approximately 50g.
 

Structure

The structure of glutamic acid was first elucidated by H. Ritthausen, who was also the chemist to first prepare glutamic acid and monosodium glutamate in 1866 to 1867.  He obtained a hydroxy acid from the nitrosation of glutamic acid. This product was reduced by a 57 percent HI in water solution (hydriodic acid) resulting in a product identical to the product of the acid hydrolysis of trimethylene cyanide (glutaronitrile). The hydroxy acid was therefore a hydroxyglutaric acid; and, since it was different from the known beta-hydroxyglutaric acid, it must have been the alpha-acid. Thus, glutamic acid was alpha-aminoglutaric acid (Greenstein and Winitz, 1961).
 
Figure: Reaction flow diagram for the determination of the basic glutamic acid structure.

With a water-free molecular weight of 169, MSG is usually commercially available as the hydrate with a molecular weight of 187. MSG is the conjugate base of glutamic acid, a dicarboxyl amino acid, and has an extremely high melting point.

Many of MSG's characteristics can be deduced from glutamic acid's characteristics. With the modification of the sodium ion, glutamic acid salt retains the chiral center possessed by glutamic acid itself. This indicates that D and L configurations of MSG also exist. Kirimura and colleagues (1974) also confirmed that the biochemical effects of the dextro and levo forms of MSG were significantly different. These forms can be racemized by excessive heating.
 
 

Flavor Interactions

MSG's effect on flavor is still in dispute. In a strictly chemical sense, MSG's chemical interactions are what one would expect them to be according to theory and proven facts. Chemical syntheses to and from MSG are readily applicable and the results are generally what would be expected. Reactions such as the Hell-Volhard-Zelinsky alpha-halogenation reaction combined with amination, or the pthalimidomalonic ester synthesis method (a combined malonic ester-Gabriel synthesis) can lead to glutamic acid which can then be neutralized with NaOH or Na2CO3 to produce MSG (Morrison and Boyd, 1973).

In turn, MSG has a variety of uses that rely on it solely for its chemical properties--such as using it to prepare glutamic acid by the previously mentioned procedures, or as a biologically compatible glutamic acid source in solution.

In a study of glutamine synthetase enzyme (known also as L-glutamate: ammonia ligase <ADP>) and its role in the detoxification of ammonia in biological systems, Dierks-Ventling and associates (1975) used MSG in their enzyme incubation solutions of brain tissue mitochondrial microsomal homogenates. MSG was the source of glutamic acid needed for the formation of glutamohydroxamic acid in this experiment. Greenstein and Winitz (1961) also cited reports where MSG itself was used to reduce high blood ammonia levels. Although results varied, they concluded that in theory, MSG, as a glutamic acid source, should be at least partially effective.

Thus, in the above chemical and biochemical interactions, MSG behaves as it should chemically. But MSG's taste effects cannot be explained on a chemical or biological basis alone. MSG's taste enhancing properties also enter the theoretical realm of genetics, physiology, and psychology. Its taste effect, although chemically the same, may be physiologically and psychologically different for each individual partaking in it.

Much of the work carried out has been an investigation of the preferences of tasters for a food with and without added MSG. Physiological and biochemical effects of the taste sensation have also been studied. MSG's effect is most evident in the pH range of 3.5 to 7.2, which also is the range that most foods possess (Meyer, 1960). MSG is generally effective in intensifying the flavor of high-protein foods such as meat, fish, eggs, and cheese. It will not work with fruits, fruit juices, sweet spicy foods, or any other foods rich in sugar and carbohydrates (Pryde, 1973). Bad flavors are also intensified by MSG.  Meat products made from old and tainted meat do not have MSG in their formulation. Peppery and spicy seasoning is instead added to anesthetize taste buds, masking the flavor of spoilage (Smith and Minor, 1974). Food characteristics such as fats, oils, and high viscosity also modify MSG's influence (Meyer, 1960).

Beef extract, hydrolyzed vegetable proteins, casein digests, and yeast autolyzate extracts are flavorings that have characteristic amino acid, peptide, protein, nucleotide, and "browning reaction" flavorings (Smith and Minor, 1974). Through hydrolysis and neutralization reactions, these crude flavor conglomerations can be purified and be brought to yield flavor potentiators such as MSG. MSG is to these crude components that sugar is to molasses. MSG is synergistic with these components in that it enhances their total effect in a way that also expresses itself.

Not only is MSG synergistic with its mother liquor components, but it is synergistic with other flavor potentiators. Flavor potentiation (as defined by Kuninaka, 1967) refers to the "action of a compound which, in small quantities, has by itself no sensory effect, but exaggerates the effect of other agents on the system."

There are two groups of known flavor potentiators (Kuninaka, 1967):

Figure: General structures for MSG, tricholomic acid,  ibotenic acid, 5'-GMP, 5'-IMP, and 5'-XMP.

The nucleotides are usually used in the form of sodium salts--therefore, they can also be termed as disodium guanylate (DSG), disodium inosinate (DSI), and disodium xanthylate (DSX) (Smith and Minor, 1974). A specific synergistic action in flavor exists between a compound of the first group and a compound of the second group (Kuninaka, 1967). The ribonucleotides (derived from muscle and yeast RNA by acid hydrolysis, then neutralized with NaOH) have essentially the same flavor enhancing property as MSG but are ten times as powerful. They are even more effective when mixed with MSG (Birch, et al, 1972).

Through extensive studies, Kuninaka (1967) and associates have determined that for the nucleotide potentiators, the ribosidic and 5'-phosphomonoester linkages are necessary for taste potentiation. Further studies have indicated that the two phospho-hydroxy groups are also necessary for this effect.

Tricholomic acid and ibotenic acid (both isolated from the fungi Tricholoma muscarium and Amanita strobiliformis respectively) have the same general structure as MSG and have essentially the same flavor potentiating effect. But unlike MSG. DSI, and DSG, which are available to the consumer, tricholomic acid and ibotenic acid are undergoing closer scrutiny than the majority of flavor effect substances.

This is because it is well known in Japan that the fungi that they are isolated from are deadly to flies. Amanita sp. in particular are well known in the U.S.A. as extremely toxic for human consumption. Consequently these last two amino acids will be exhaustively studied before they are allowed on the consumer's table.

Theories on these flavor potentiators' mechanisms are many. Pryde (1973) stated that MSG could increase taste bud sensitivity, stimulate greater saliva flow, and combine with trace metals present, thus freeing additional taste bud receptor sites to react with the taste stimulating compounds present. Majtenyi (1974) also mentioned the above theories, but added that it was nucleotide binding of metal ions that sensitized tasting sites otherwise not active in the enhancement of flavors.

MSG is used at a level of one-tenth the concentration of salt used in food. Its use is restricted to protein food. From this, the impact of MSG on the American diet can be realized. In the United States alone about 30 billion pounds of protein foods are seasoned with MSG annually (approximately 150 pounds per person per year) (Smith and Minor, 1974). This use is increasing rapidly.

In 1950, at least one company in the U.S. produced 3500 tons and the other companies also produced substantial amounts (Markley, 1950). In Japan, where the main production of MSG occurs, the 1961 national production topped 27,571 tons. The country exported 7,663 tons (Harada and Motoyama, 1964). The world's supply of MSG was reported in 1974 to be 75,000 tons (150,000,000 pounds) per year valued at $80 million. Yet production in the United States is less than one-fourth the total amount produced (Smith and Minor, 1974).

Convenience foods particularly benefit from the addition of MSG and there may be some preservative qualities associated with its use. MSG is often poured quite liberally on restaurant foods (Pryde, 1973). Soups, stews, and other related foods use MSG extensively (Birch, et al, 1972). Artificial sake (rice wine) consumed in Japan is made by blending known ingredients found in natural sake, including MSG.  It is made by blending ethanol, glucose, succinic acid, lactic acid, inorganic phosphates, sodium chloride, MSG, glycine, and alanine. Dipeptides of Gly-L-Leu, L-Val-L-Glu were also added to improve body, increase the complexity and balance of taste, and serve as buffering agents (Kirimura, 1969). Even certain drugs contain MSG, for example, Glutavine (Turner, 1970).

Being an amino acid, MSG has been regarded as harmless. But recent evidence suggests that there may be unpleasant effects when large amounts are ingested (Birch, et al, 1972). Some people have low thresholds of sensitivity to MSG (Majtenyi, 1974) and a "Chinese Restaurant Syndrome" (CRS) has been medically recognized as their reaction to ingestions of MSG. CRS includes these symptoms: burning sensation in the back of the neck spreading to the forearms and to the anterior thorax, accompanied by a feeling of infraorbital tightness, pressure, substernal discomfort (Bernarde, 1971), headaches, temporary weakness and numbness, loss of breath, fainting, and other symptoms resembling heart disease (Pryde, 1973).

Since MSG is derived from glutamic acid, the most abundant protein building block in nature, those who are allergic or sensitive to proteins may also be sensitive to MSG (Smith and Minor, 1974). But avoiding MSG is extremely difficult. Even a careful reading of all labels will not uncover the products to which MSG may be legally added. More than ten thousand processed foods contain MSG, and many, such as mayonaise, French dressing, and salad dressing do not have to list MSG if it is used. Although glutamate is present in all proteins in the body, an excess amount can become toxic and destroy nerve cells (Pryde, 1973). This has caused a dilemma in the food industry. Food companies cannot be relied upon to police themselves. They have acquired the assumption that a product can remain on the market until definitely proven dangerous (Turner, 1970).

Some experiments involving feeding MSG to infant rats and mice were done by Dr. John W. Olney of Washington University. The rats incurred brain damage (lesions), stunted skeletal development, marked obesity and sterility in the females (Benarde, 1971). Another study revealed that high doses caused brain lesions in infant primates (Majtenyi, 1974). Pryde (1973) stated that continuing research has shown other effects of MSG in experimental animals. Parts of the brain were numbed and temporarily inhibited and teratogenic effects (birth defects) were also demonstrated. Yet tests still need to be carried out on MSG. Cytogenic changes, primarily chromosome breaks in the cells of MSG-treated animals or bacteria, should be investigated (Turner, 1970).
 

References Cited

Benarde, Melvin A. The Chemicals We Eat. New York: American Heritage Press, 1971.

Birch, Gordon G.; Spencer, Michael; and Cameron, Allan G., Food Science. New York: Pergamon Press Ltd., 1972.

Dierks-Ventling, Crista; Cone, A.L.; and Bessman, S.P., "The Attachment of Glutamine Synthetase to Brain Membranes." Biochemical Medicine. July 1975.

Giese, Arthur. Cell Physiology. Philadelphia: W.B. Saunders Co., 1973.

Greenstein, Jesse P. and Winitz, Milton. Chemistry of the Amino Acids. New York: John Wiley and Sons, Inc., 1961.

Harada, Zo and Motoyama, Hagifune. "Ajinomoto."  in Sekai Daihyakka Jiten. Vol. 1 A-isumo. Kunihiko Shimonaka ed. Tokyo: Kabushiki Kaisha Hebonsha,
1964.

Kirimura, Jiro et al. "The Contribution of Peptides and Amino Acids to the Taste of Foodstuffs." Journal of Agricultural and Food Chemistry. July/August, 1969.

Kuninaka, Akira. "Flavor Potentiators." In: Symposium on Foods: The Chemistry and Physiology of Flavors. H.W. Schultz, ed. Westport, Connecticut: The Avi Publishing Company, Inc., 1967.

Majtenyi, Joan Z. "Food Additives--Food for Thought." Chemistry. May 1974.

Markley, Kare S. ed. Soybeans and Soybean Products. New York: Interscience Publishers, Inc., 1950.

Meyer, Lillian H. Food Chemistry. New York: Reinhold Publishing Corporation. 1960.

Morrison, Robert Thornton, and Boyd, Robert Neilson. Organic Chemistry. 3rd ed. Boston: Allyn and Bacon, Inc., 1973.

Pryde, Lucy T. Environmental Chemistry: An Introduction. Menlo Park, California: Cummings Publishing Company, 1973.

Pyle, James L. Chemistry and the Technological Backlash. Englewood Cliffs, New Jersey: Prentice Hall, Inc., 1974.

Smith, Laura, and Minor, Lewis, ed. Food Service Science. Westport, Connecticut: Avi Publishing Co., Inc., 1974.

Turner, James. The Chemical Feast. New York: Grossman Publishers, Inc., 1970.