Ascorbic Ac id Derivatives and Processes

This invention relates to novel derivatives of L-ascorbic and dehydroascorbic ac ids .

0 The compounds of the invention have the structure

5

wherein

OH

20 X is - C

I

H

Y is a group selected from the class consisting of

25

OH OH

I I - C = C - and

30 0 0

- C - C - groups, and n is a positive integer.

- - ^> The compounds are prepared by the self-esterifi- cation of L-ascorbic acid and dehydroascorbic acid, accordin to the following equations:

(1) Self-esterification of L-ascorbic acid

(L-ascorbic acid)

H (I )

( 2) Self-esterif ication of dehydroascorbic acid

(dehydroascorbic acid)

H = O (II)

Reactions (1) and (2) can be carried out by carefully heating the acids or mixtures thereof in the presence of water in sealed glass tubes at 100°C until the esterification reaction is complete. Water in the esteri- fication mixture shifts the equilibrium between the open- chain and cyclic lactone forms of ascorbic acid toward the cyclic form, i.e.,

(3) Equilibrium of straight chain and cyclic lactone forms of ascorbic acid

which, in turn, favors reactions (1) and (2). The reaction continues as long as there is unreacted acid present, yielding products (I) and (II) of increasingly higher molecular weights as the reaction continues. Broad infrared absorbance peaks in the region 1600-1700 cm , character¬ istic of the cyclic rings of the acids, diminish as the reactions proceed and are at very low levels at molecular weights of the reaction products corresponding to the tetramer.

Ultimate analysis of the products eluted from the reaction mixture by gel chromatography at MW 700 and higher confirms the predicted empirical formulas for products (I) and (II) at n _ 3. Product (I) is colorless and product (II) is yellow.

Products (I) and (II) and mixtures thereof are useful in treatment of idiopathic movement disorders in laboratory animals (dogs) and are confirmed non-toxic by rat-feeding studies.

An alternative preparation of the compounds of the invention involves the partial conversion of an aqueous L-ascorbic acid solution to a metal salt, advantageously the calcium salt to provide a non-toxic product, heating

the aqueous acid-salt mixture in the absence of oxygen at a temperature in the range 80-95°C for a length of time sufficient to attain the desired degree of esterification, completing the conversion of unconverted acid in the esterification mixture by reaction with excess salt conversion reagent, e.g., calcium carbonate and drying the reaction mixture. The esterified product including metal salts thereof is separated from the unreacted reagent, e.g., by water-leaching the soluble product out of the dried reaction mixture and drying the leach solution to obtain a product containing the ester derivatives and salts thereof, which may be used as such or subjected to gel separation to isolate the higher molecular weight self- esters and salts thereof from the lower molecular weight species and the ascorbate salts.

If the dehydroascorbate self-ester product is desired, all or a portion of the L-ascorbic acid can be oxidized by bubbling air through the reaction mixture after the partial salt conversion step described above, followed by the esterification, drying and separation steps.

The following examples are presented to illustrate the practice of the invention, and not as limitations on the scope thereof:

Example 1

L-ascorbic acid- is moistened with distilled water and placed in sealed and open glass tubes in an oven preheated to 100°C. After six hours, the tubes are with- drawn and examined by infrared spectroscopy. The materials in the sealed tubes do not show infrared absorption bands in the 1600-1700 cm ^-1 band while those in the open tubes clearly show such bands characteristic

i

of the lactone ring structure of L-ascorbic acid. The dry residue in the open tubes is dark brown. The ascorbic 5 acid self-ester product, isolated from the material in the sealed tubes by gel chromatography using Sephadex G-10 eluting solvent, contains molecular weight species varying from 425 to 700.

0 Example 2

To a 2-liter, 3-neck flask equipped with an agitator, thermometer and vent tube is added 300 ml distilled water and 440 g. (2-5 moles) L-ascorbic acid. 5 To this stirred slurry is added incremental additions of finely divided calcium carbonate, while maintaining the reaction temperature at about 20°C and at a rate to produce constant evolution of carbon dioxide (reaction byproduct) . Calcium carbonate addition is suspended 0 after about 25 g. to about 37.5 g. are added (representing between 20% to about 30% of that required for complete reaction with the L-ascorbic acid charge) .

At this point, the temperature is raised to 80°C 5 while maintaining a carbon dioxide atmosphere in the reactor. At intervals, samples of the reaction mixture are removed and analyzed by infrared spectrometer to determine the degree of L-ascorbic acid esterification which has taken place, further additions of calcium carbonate are 0 begun, while maintaining the temperature in the range of

60°C and about 70°C, and while still maintaining an oxygen- free atmosphere in the reactor. The total quantity of calcium carbonate added is 125 g. (1.25 moles).

5 The reaction mixture is transferred to a covered shallow container in an essentially oxygen-free atmosphere, maintained at a temperature ranging between 60° and 80°C for a period of from 12 to 36 hours, during which time the

pH of the mixture rises to a pH range of 6.0-7.0. At this point, the excess water is removed under vacuum.

5

The dry products are light tan in color and readily soluble in water, except for unreacted calcium carbonate, to produce neutral solutions. Analysis indicates that the products possess molecular weights averaging

_] _o about 460. By gel filtration techniques using Sephadex G-10 eluting solvent, a product which is eluted at the solvent front is isolated which has molecular weights in excess of 700. This corresponds to self-ester species having at least four ascorbic acid units. Some units

_] _5 contain calcium.

Example 3

The procedures of Example 2 are followed, with 20 the exception that air is not excluded from the reaction mixture throughout the operation. This leads to the intermediate formation of dehydroascorbic acid and resul¬ tant self-esterified dehydroascorbic acid and salts thereof.

25 The quantities of these oxidation products are regulated by the extent of air contact within the reactor and during the subsequent heating and drying periods.

The dry products generally have similar physical 2 _Q characteristics to those described in Example 1. They are usually darker in color, being light brown.

Example 4

35 This example illustrates the separation of the polymeric products of the invention from the reaction mixture of Example 1 by gel permeation techniques.

The 160 Waters protein-packed gel permeation columns are calibrated using 6.5 K and 1.6 K W sulfonated polystyrene standards (commercially available from Pressure Chemical Co.) and using sucrose and glucose as lower molecular weight standards. Further calibration runs are made along with a chromatogra of L-ascorbic acid as a reference point.

The dry residue from the open tubes of Example 1 is disolved in 0.05 amonium acetate in a 10% (wt) methanol- water solution. The aceta e-methanol-water solvent has a pH of 7.2.

15 ml of a 1 wt% solution of the dried reaction product of Example 1 is injected at a rate of 1.5 ml/minute into the 160 Waters protein columns. These columns are monitored at 8X on a Waters Model 401 refractometer and -simultaneously at 254 nm 2.0 AUFS on a Waters Model 440 absorbancy detector. The temperature of the columns is maintained at 40°C.

Using the above-described techniques it is determined that the dried residue product of Example 1 contains molecular species having a molecular weight distri¬ bution, as follows:

Molecular Weight Lowest 326

Mean 454

Highest 554

The polymeric molecular species of the dried residue of Example 1 are separated from unpolymerized monomer (which is eluted last) using the same gel separation technique. The polymeric species are identified by molecular weight and are present in the following amounts:

Species Weight %

Dimer 70.4

Trimer 17.4

Tetramer 12.2

Each of the dimer, trimer and tetramer products is separated from the other polymer product by repetition of the gel-separation techniques described above.

Infrared spectroscopy reveals that the C=0 stretch in the lactone ring of the ascorbic acid progres¬ sively disappears as the molecular weight of the self- ester increases. At the same time a new straight-chain ester carbonyl appears, confirming the self-ester structure.

Example 5

^Tne product of Example 2 which contains, molecular species with molecular weights in excess of 700 is further subjected to the gel-separation procedure of Example 4. It is determined that this product contains molecular species having the following distribution:

Molecular Weight Lowest 189

Mean _ _. 418

Highest 710