IL31964A - Tryptophane derivatives and their use as sweetening agents - Google Patents

Description

TRYPTOPHANE DERIVATIVES AND THEIR USE AS SWEETENING AGENTS The present invention relates to a non-nutritive sweetener which consists of or contains the dextro enantiomorph of a substituted tryptophane and to- novel tryptophane compounds .

Numerous substances have been proposed and/or used Y as non-nutritive agents, affording the usey thereof a sense of sweetness at least desirably comparable to that obtained with natural sugar, but without caloric effect. Such substances, while necessary for some persons in order to limit intake of the natural sugars and thus control various health conditions, including diabetes, still possess several disadvantages, including a bitter aftertaste and toxic side effects. At present, only two classes of non-nutritive sweetening substances are used to any great extent: saccharin-type compounds and cycla-mate-type compounds, both of which classes show the typical disadvantage of bitter aftertaste; and in addition, the cycla-mate-type compounds have only limited activity.

Among other chemicals evaluated for sweetening effect are the amino acids and enantiomorphs thereof as.reported by Vuataz et al., E perientia, Vol. 21, pages 6 2-69^, inclusive (I965) . The report shows that while a number of amino acids are sweet in the d-enantiomorphic form, this is not an absolute correlation. Furthermore, despite the sweet taste of the d-enantiomorph of a given amino acid, the 1-enantiomorph of the same amino acid may be bitter. Resolution of the two enantiomorphs of a given amino acid is often difficult. For these various reasons, despite the contribution of Vuataz et al., no amino acid is being used in the sweetening art.

Moreover, throughout the sweetening art, it is well known that there is no correlation between structure and activity, as even relatively small changes in chemical structure often destroy activity.

It has now been discovered that certain substituted tryptophane compounds, in their d-form, exhibit a sweet taste of a marked degree, at rates below rates at which any undesirable side effect may be noted, and the present invention is directed to the use of an active agent which is the d-enantio-morph of a substituted tryptophane compound of the formula I H or its non-toxic physiologically acceptable salts, wherein R represents halo of an atomic weight of less than 85, lower alkyl, lower alkoxy, or trifluoromethyl; R' represents hydrogen, halo of an atomic weight of less than 85, lower alkyl or lower alkoxy; and R" being hydrogen when R is trifluoromethyl. In the present specification and claims, the term "lower alkyl" and "lower alkoxy" are employed to designate alkyl and alkoxy radicals containing from 1 to 4, both inclusive, carbon atoms. Preferred alkyl and alkoxy radicals are methyl and methoxy, Included within the scope of the generic formula and valuable as non-nutritive sweetening agents are tryptophanes known to the prior art as well as novel tryptophanes.

The compounds to be employed in accordance with the present invention are typically crystalline solid materials, with solubilities varying according to whether the compounds are the alkaline or acid addition salts, or the free amino compounds. As water solubility is desirable in the typical usage of a substance as a sweetening agent, the salts are gen-erally preferred, especially for applications wherein the active agent is to be present in higher concentrations.

The type or identity of salt is not critical except that it be non-toxic and physiologically acceptable. Suitable alkaline salts are the ammonium, sodium, potassium, calcium, and magnesium salts, the sodium and calcium salts being preferred. Acid salt formation occurs at the amino nitrogen atom, the ring nitrogen atom be;lng only weakly basic, making it necessary that the acid be a strong acid, having a pH of, numerically, below about 2, at a concentration of 0.1N. Suitable strong acids are hydrochloric, hydrobromic, hydriodic, sulfuric, and the like.

The free amino compounds of the present invention can be prepared readily by employing art-recognized procedures.

Thus, for example, in one method of synthesis, an indole is caused to undergo a typical annich reaction with formaldehyde and diethylamine to yield a substituted 3- (diethylaminomethyl) -indole which is then condensed with diethyl formamidomalonate according to the general procedure of Rydon et al., J. Chem. Soc, 1955, 3 9 , to form the corresponding diethyl (3-indoly-methyl)formamidomalonate. Hydrolysis and decarboxylation of the foregoing intermediate produces the desired substituted tryptophane .

Alternatively, an appropriately substituted phenyl- hydrazine is condensed with the adduct of acrolein and acet-amidomalonic ester described by Moe et al., J.Am. Chem. Soc., 70, 2765 (1948) . Hydrolysis and decarboxylation affords the desired substituted tryptophane.

Suitably substituted indoles and phenylhydrazlnes which serve as intermediates in the foregoing syntheses are prepared by methods known in the art. The indoles can be prepared either from the appropriately substituted £-nitrotolu-enes, by the method of Rydon et al., J. Chem. Soc., 1955* 3499» or by cyclization of suitably substituted phenylhydrazlnes. The latter can be prepared by the general procedure of Bullock et al., J. Am. Chem. Soc., 78, 585 (1956) .

The compounds obtained as described hereinabove can thereafter be reacted further with a stoichiometric amount of either a base or acid to procure the corresponding salt, the particular reaction to prepare the present salts being carried out in accordance with procedures well known in the art for this type of reaction. Resolution of the compounds serving as active agent in accordance with the present invention is not necessary, the racemic mixture itself being quite active. However, since the d-enantiomorph is the active moiety, resolution may be preferred to lessen the amount of substance needed for the desired sweetening effect.. Moreover, due to the fact that typically only the 1-enantiomorph of amino acids is metabolized by the mammalian body, usage of the d-enantlomorph alone, may be preferred to preclude any opportunity for the mammalian body to incorporate the substance.

When it is desired to employ only the d-enantiomorph, resolution of the racemlc mixture can be achieved in procedures known in the prior art for the resolution of unsubstituted tryptophane . Three such procedures are discussed and exemplified in detail in Chemistry of the Amino Acids , Greenstein et al , , Vol, 3, page 2341 and following (John Wiley and Sons , Inc . , New York, 1961) ; particular attention is dire cted to the first two of these (illustrative procedure 39-5 and illustrative procedure 39-6) .

Representative compounds to be employed as active agent in accordance with the present invention Include the following : 6-chlorotryptophane 6- (trif luoromethyl) tryptophane 6-bromotryptophane -chloro-6-methyltryptophane 6-methylt yptophane 6-f luorotrypt ophane 6-ethyltryptophane , 6-dichlorotryptophane -f luoro-6-methyltryptophane -chloro -6-bromotryptophane -methyl -6 -chlorotryptophane, sodium salt -f luoro-6-chlorotryptophane , 6 -d ime t hy 11 ry pt ophane -bromo-6-methyltryptophane -me t hy 1 -6 -bromo t r pt ophane - bromo -6-chlorotryptophane 6- chlorotryptophane, sodium salt ,6-dichlorotryptophane, sodium salt 6-chlorotryptophane, potassium salt ,6-dibromotryptophane 6- (tri luoromethyl) tryptophane, ammonium salt 6-n-propyltryptophane ,6-dimethyltryptophane, sodium salt 6-lsopropyltryptophane ,6-dichlorotryptophane hydrochloride 6-n-propyltryptophane ,6-dimethyltryptophane nitrate 6-methoxytryptophane -methoxy-6-chlorotryptophane 6-ethoxytryptophane 6-isopropoxytryptophane 6-n^butoxytryptophane ,6-difluorotryptophane ,6-dimethoxytryptophane 6-tert-butyltryptophane -methyl-6- luorotryptophane 6-chlorotryptophane citrate -fluoro-6-bromotryptophane 6-chlorotryptophane hydrochloride -bromo-6=fluorotryptophane 6-bromotryptophane hydroiodide ,6-dimethyItryptophane hydro chloride -chloro-6-fluorotryptophane Mixtures of two or more compounds can also be employed. Any given compound can be employed either as the d-enantlomorph or as a mixture of the d- and 1-enantiomorphs .

The substance of the present invention can be used to sweeten various substances, such as, for example, chewing gum, toothpaste, lip cosmetics, mouthwash, mouthspray, sub-stances used in dentistry for cleansing of teeth, denture treating substances, chewing tobacco and other tobacco products; pet toys, for example, rubber dog bones, as well as other mechanical devices temporarily retained within the mouth; glues and adhesives, as for use on stamps and envelopes; coffee, tea, fruit ades, salad dressings, carbonated and non-carbonated beverages; baked goods such as bread, crackers, pretzels, pastries, or cake, cereal products; milk derived products, such as ice cream, ice milk, sherberts, custards and other puddings; jello and gelatin products; and processed vegetables and fruits, such as, for example, canned tomatoes, frozen vegetables, and the like; and nutritive solid foods, including meat products in which a sweetening substance is incorporated during processing, such as ham and bacon; prepared "mixes" such as mixes for puddings, cakes, pastries, and the like; and confectionary products, for example, popcorn, peanut candies, chocolate candies, jellybeans, gumdrops, candy cigarettes, taffy, licorice, and the like; natural sugar and glycine and other amino acids which are nutritive; a feed, such as a grain-type feed, silage, salt licks; can be used in baits as an attractant; fruit and vegetable juices; alcoholic beverages such as beer, wine, cocktails and cocktail mixes, milk beverages such as milkshakes, "nogs", and the like; medicinal substances such as a tablet, capsule, powder, or lozenge, including cough drops; an elixir, syrup, suspension, and the like.

The sweetener of the present invention can be combined with a flavoring agent. The flavoring agent can be one which is contained in, as an inherent part of, a natural food; or the flavoring agent can be one specifically added to a sub-stance, as, for example, a flavoring agent added to a chewing gum. In this sense, "flavoring agent" is used to describe a substance which has a discernible and desirable flavor at a concentration in liquids of 250 ppm. or less, even though in other specialized applications, such as chewing gum, and high-ly flavored baked goods, higher concentrations may be used.

Representative such flavoring agents include spices and herbs; the essential oils and their extracts; fruit-derived flavorings; plant extracts, as, for example, cola, caffeine, etc.; and synthetic flavorings, including those which simulate or duplicate the effective components of the flavoring agents of the previous categories.

The present non-nutritive agent is conveniently formulated as a tablet, capsule, liquid or powder.

The amount of the present non-nutritive sweetener to be employed is not critical either, it being necessary only that an effective amount is used, an effective amount being that amount which provides a sense of sweetness comparable to that afforded by sucrose at a given usage rate. The amount of the present active agent to be used will also depend upon such variables as the purpose of sweetening and other factors. For sucrose concentrations of from about 6.5 percent to 5 percent, concentrations including most food and even many confectionary applications, the present active agent, when substituted for sucrose, gives approximately equivalent sweetness at concentra tions of from about 0.05 to 30 percent, as the racemic mixture Where the d-enantiomorph is employed alone, these rates can be reduced 'by a factor of about one-half. Higher or lower concen trations of the active agent if the present invention can be used where the degree of sweetness, by a sucrose standard, is greater or lesser. However, where a high degree of sweetness is desired, it is generally preferred to use a combination of the present active agent with one or more other known non-nutritive sweeteners, as is discussed further hereinbelow.

In an initial evaluation, dl-6-chlorotryptophane was administered orally to a group of mice at the rate of 2 grams of compound per kilogram of individual animal body weight. No toxic symptoms of any type were noted. The same results were obtained where the pompound being evaluated was dl-6-methyltryptophane and where the compound being evaluated was dl-6-fluorotryptophane.

The compounds serving as the present active agent can be employed as sole agent; or, alternately, such com-pound or compounds can be employed jointly with other known non-nutritive sweeteners. In particular, one or more of the compounds serving as active agent in accordance with the present invention can be combined with a saccharin-type sweetener; with a cyclamate-type sweetener; with a dihydro-chalcone-type sweetener; or with 5- (3-hydrox phenoxy) -1H-tetrazole. As employed in the present specification and claims, the term "saccharin-type sweetener" means a compound having the essential structure described above and includes non-toxic physiologically acceptable salts thereof, generally, the sodium or calcium salt.

As used in the present specification and claims, the term "cyclamate-type sweetener" means a compound having the es-s ential structure of cyclamic acid, and includes non-toxic physiologically acceptable salts thereof, generally, the sodium or calcium salt, although the magnesium salt has also been used for some purposes. - (3-Hydroxyphenoxy) -lH-tetrazole can, in accordance with the present invention, be employed as one of its non-toxic physiologically acceptable salts. Suitable salts include the sodium, calcium, potassium, and ammonium salts.

The dihydrochalcone-type sweeteners include those of the genus disclosed and claimed in U.S. Patent 3, 087, 821, neohesperidin dihydrochalcone being a preferred compound. A publication by Krbechecks et al (J. Ag. and Food Chem., Vol. 16, No. 1, page 108 /Γ 687) on this genus discloses other related compounds, notably higher alkoxy substituted compounds, which are even more sweet, and such related compounds are also dihydrochalcone-type sweeteners in accordance with the present invention and can be employed jointly with the active agent of the present Invention.

When the active agent of the present invention is used in conjunction with another non-nutritive sweetener, the exact ratio of the components is not critical and can vary considerably, depending upon the particular use.

Representative rates for the present active agent, when employed alone, are set forth hereinabove. When the o active agent is combined with a saccharin-type sweetener, synergistic effects are obtained when combining the substance in a ratio ranging from 300 parts of present active agent to 1 part of saccharin-type sweetener, to 1 part of present active agent to 300 parts of saccharin-type sweetener by weight, a generally preferred range of synergistic combination being that of a ratio ranging from 1: 50 to 50: 1, by weight .

In the combination of the present active agent with 5- (3-hydroxyphenoxy) -lH-tetrazole, synergistic results are ob-tained at a ratio ranging from 1 part of present active agent to 300 parts, of the 5- (3 -hydrox phenoxy) -lH-tetrazole by weight to .1 part of the 5- (3-hydroxyphenoxy) -lH-tetrazole to 300 parts of the present active agent ; a particularly preferred ratio of combination is a ratio of from 1: 50 to 50: 1, by weight .

Synergism with the dihydrochalcone -type of sweetener, synergism is likewise noted throughout mos t of the range , such as a ratio ranging from 1 part of the present active agent to 30 parts of the dihydrochalcone-type sweetener, by weight , to a ratio of 1 part of the dihydrochalcone-type sweetener to 30 parts of the present active agent, by weight . A preferred ratio is from 1: 5 t° 5: 1 by weight .

Synergism with a cyclamate -type sweetener is noted in a ratio ranging from 1 part of cyclamate-type sweetener to 320 parts of the present active agent , by weight , to 1 part of the present active agent to 320 parts of the cyclamate-type sweetener, by weight . A preferred ratio ranges from 1 : 60 to 60: 1, by weight .

It is also possible to combine the present active agent -with sucrose or other nutritive sweeteners so as to obtain a sweetening substance of reduced caloric value. Such a mixture of nutritive sweetener and the present active agent can be formulated as a foam or the like, in accordance with procedures known in the prior art for a combination of sucrose and sodium saccharin. Such manner of formulating results is a composition having both volume and sweetening effect equivalent to sucrose, alone, but with reduced caloric value.

The following examples illustrate the present inven-tion and will enable those skilled in the art to practice the same.

Example 1 An initial series of evaluations was carried out, each in essentially identical procedures. Each evaluation comprised the tasting of a small amount of the given compound, such amount being that which would adhere to a fingertip. Compounds so evaluated were rated for degree of sweetness, and aftertaste, if any. The sweetness ratings were as follows: TABLE I Compound Evaluated Sweetness Rating dl-6- (trlfluoromethyl) tryptophane very sweet d 1 -6-bromotryptophane sweet dl-6-methyltryptophane very sweet dl-6-chlorotryptophane very sweet dl -6-chlorotryptophane hydrochloride very sweet dl-6-fluorotryptophane very sweet dl-5, 6-dimethyltryptophane sweet dl-5, 6-diehlorotryptophane very sweet dl-6-isopropyltryptophane hydrochloride sweet dl-6-methoxytr ptophane sweet dl-6-chlorotryptophane, sodium salt very sweet No aftertaste was observed with any of the compounds evaluated, Example 2 In further operations, dl-6-chlorotryptophane was evaluated In a carbonated beverage; the compound was dissolved at different concentrations in a number of solutions, each having the following composition: sodium citrate 0.025 percent citric acid 0.22 percent alcoholic lemon-lime extract 0.125 percent sodium saccharin 0.01 percent the remainder being carbonated water. The various solutions were judged by a taste panel of five persons to determine which was comparable, in sweetness, to a standard solution.

The standard solution contained sodium citrate, citric acid, and alcoholic lemon-lime extract in the same concentrations as above, but contained 0.1 percent of sodium cyclamate and 0.01 percent of sodium saccharin. This standard solution, therefore, was approximately equivalent to a commercially available carbonated beverage of this type sweetened with a known non-nutritive sweetener. The mixture containing the dl-6-chloro-tryptophane in a concentration of 0.0125 percent was Judged to be equivalent in sweetness to the standard. No off-flavor was noted in any of the solutions containig dl-6-chlorotryp-tophane.

Example 3 In other operations, dl-6-chlorotryptophane was evaluated in hot cocoa. In these operations, the compound was dissolved in a number of mixtures, at varying concentrations, each such mixture comprises 1 cup of milk, a tablespoon of commercially available cocoa powder, 0.02 percent of sodium saccharin, and the dl-6-chlorotryptophane in a given amount. Each such mixture was heated to the boiling temperature and then permitted to cool somewhat. All mixtures were evaluated by a taste panel of five persons, to determine which of the mixtures afforded a degree of sweetness comparable to another mixture prepared as described but containing 1 cup of milk, 1 tablespoon of the same commercially prepared cocoa powder, and 0.2 percent of sodium cyclamate and 0, 02 percent of sodium sac-charin. The consensus of the taste panel was that the mixture containing 0, 0125 percent of the dl-6-chlorotryptophane afforded a sense of sweetness comparable to that of t e standard control solution. No off-flavor was noted in any of the cocoa mixtures containing the dl-6-chlorotryptophane.

Example 4 In another evaluation, dl-6-chlorotryptophane was evaluated in a vanilla-flavored dessert sauce. In the evalua-tion, the compound was dissolved in various mixtures, each such mixture comprising the following ingredients: 1 cup milk 1 tablespoon of cornstarch 1/8 teaspoon of salt sodium saccharin, in a concentration of 0.08 percent dl-6-chlorotryptophane in a given amount The concentration of the dl-6-chlorotryptophane was varied among the several mixtures. Each such mixture was then cooked until gelatinized, and thereafter cooled and 2 table -spoons of butter and £ teaspoon of vanilla added to each mixture. Another such mixture was prepared as a standard, as above, except that in place of the sodium saccharin and dl-6-chlorotryptophane, there was supplied sodium cyclamate in a concentration of 0.4 percent, and sodium saccharin in a con-centration of 0.04 percent. A taste panel of five persons then evaluated the various dessert sauces; the consensus of the panel was that the sauce containing 0.08 percent of sodium saccharin and 0.08 percent of dl-6-chlorotryptophane afforded a sense of sweetness approximately comparable to that of the standard. Some off-flavor was noted by some members of -the taste panel in the sauces containing the dl-6-chlorotrypto-phane and sodium, saccharin; this off-flavor was believed to be due to the sodium saccharin.

Example 5 A liver-flavored dog food comprising dl-6-chloro-tryptophane is prepared. The preparation consists of the mixing and blending of meat and meat by-products, including liver, soy grits, dried whey, propylene glycol, animal fat preserved with butylated hydroxy- anisole and butylated hydroxytoluene, dicalclum phosphate, iodized salt, dried yeast, potassium sorbate, garlic powder, artificial coloring, vitamin A and Ώ oil, vitamin B supplement, vitamin E supplement, as standard ingredients, and, in addition thereto, dl-6-chloro-tryptophane, in an amount to provide 0.075 percent by weight of the ultimate dog food composition. The resulting mixture is compacted in standard procedures as small bite-size chunks.

This composition serves as a complete dog food which is at-tractive to dogs but without the caloric value of sucrose or other nutritive sweetener.

Example 6 A dog treat composition is prepared comprising, as standard ingredients: wheat flour, corn meal, soybean meal, meat meal, wheat germ meal, poultry by-product meal, animal fat preserved with butylated hydroxyanisole, condensed fish solubles, dried skimmed milk, poultry fat stabilized with butylated hydroxyanisole, dried meat solubles, dried vegetable pomace, dehydrated cheese, animal liver and glandular meal, steamed bone meal, salt, leavening, titanium dioxide, vitamin A oil, irradiated yeast, certified food colors, and, as non-nutritive sweeteners, dl-6- (trifluoromethyl) tryptophane in an amount to constitute 0.0175 percent by weight of the total composition, and sodium saccharin in an amount to constitute 0.05 percent by weight of the total composition. The ingredients are mixed together and compacted in the form of bone-shaped snacks which have enhanced palatability without en- hanced caloric content.

The preparation of the novel compound 6- (trifluoromethyl) tryptophane and intermediates in its synthesis, is illustrative of the method of synthesis of the compounds useful in the sweetening process described hereinabove.

Preparation 1 6- (Trifluorometh 1) tryptophane To a solution of 7.53 grams of diethylamlne (0.15 mole) in 22.5 ml. of cold 60 percent acetic acid was added 8.25 ml. of 37 percent aqueous formaldehyde. 6- (Trifluoromethyl) indole (19.0 grams; 0.10 mole) was then added, and the resulting mixture warmed to 60°C. After two hours at this temperature, the solution was poured into 36Ο ml. of 2N sodium hydroxide, and the desired 3- (diethylaminomethyl) -6- (trifluoro-methyl) indole product extracted with ether. The extract was dried, and solvent removed by distillation to separate the product, which was an oil. 3- (Diethylaminomethyl) -6- (trifluoromethyl) indole (12.5 grams; 0.046 mole), prepared as described in the fore-going example, and diethyl formamidomalonate (9.4 grams; 0.041 mole) were mixed in 31 ml. of toluene. Thereafter, Ο.76 gram of powdered potassium hydroxide was added. The resulting mixture was refluxed, while bubbling in nitrogen, for 1.5 hours. It was then cooled, resulting in precipitation of the diethyl (^5"- (trifluoromethyl) -S-indolyJ^meth lJformamidomalonate product, which was separated by filtration and washed with water. A portion of the separated product was recrystallized from ethanol, and the recrystallized portion found to melt at 182 -5°C.

A solution of diethyl (^5"- (trlf luoromethyl) -3-indolyl^methyljformamldomalonate (55.3 grams; 0.138 mole) In 330 ml. of 1.5N hydrochloric acid, I65 ml. of glacial acetic acid, and 50 ml. of ethanol was refluxed for 17 hours. The solution was then concentrated under vacuum, the residue neutralized with ammonium hydroxide, and the resulting 6-(tri-f luoromethyl) tryptophane product separated by filtration. The separated product was washed with water and ethanol, and a sample thereof recrystallized from acetic acid. The re-crystallized portion melted at 267-70°C. (dec.).

Claims (6)

1. A non-nutritive sweetener characterized in that it consists of or contains the d-enant iomorph o^ a substituted tryptophane compound of the formula I H or its non-toxic physiologically acceptable salts, wherein S represents halo of an atomic weight of less than 85, lower alkyl, lower alkoxy, or trifluoromethyl; R' represents hydrogen, halo of an atomic weight of less than 85, lower alkyl, or lower alkoxy, R' being hydrogen when R is tri-fluoromethyl; and the non-toxic physiologically acceptable salts thereof.

2. The non-nutritive sweetener of claim 1 characterized in that the substituted tryptophane compound is 6-chlorotryptophane , 6- (trifluoromethyl)tryptophane or 6-meth ltryptophane .

3. The non-nutritive sweetener of claim 1 or 2 characterized in that it also contains substances selected from saccharin-type sweetener; cyclamate-type sweetener; dihydrochalcone-type sweetener; 5- (£-hydroxyphenyl)-lH-tetrazole or a non-toxic physiologically acceptable salts thereof, and the 1,-enant iomorph of the substituted tryptophane of formula I.

4. . A sweetened composition characterized by a bas ic material and as the sweetener, the non-nutritive sweetener 31964/2 of claim 1, 2 or 3,

5. The composition of claim 4 characterized in that it contains a flavoring agent.

6. A process for sweetening substances characterized by adding to the substance to be sweetened the sweetener of claim 1, 2 or 3, 7, . The substituted tryptophane compound of formula I defined in claim 1 characterized in that R* is hydrogen and R is trifluorom ethyl and the non-toxic physiologic ally- acceptable salts thereof. AG