CS209862B2 - Method of making the gama-l-glutamyltaurin - Google Patents

Description

The present invention provides a process for the preparation of? -L-glutamyltaurine of the formula

H ₂ N - CH - COOH (CH ₂ ) 2

CO-NH- (CH2) 2-SO3H and its optically active antipodes, characterized by a broad spectrum of therapeutic and preventive efficacy against pathological changes having a direct or indirect cause in damage to the aerobiospheric genetic adaptation system (AG-AS).

To clarify the concept of AGAS, the most important tissues and organs forming this system are listed below:

(a) All biological boundary surfaces in contact with external air as the biosphere (skin, and cutaneous formations, cornea and conjunctiva, mouth and esophagus, airways and lungsJ,

b) skeleton and limbs (hollow tubular bones and spongy bones, spherical joints, lining of the joint cavity, skeleton muscles), ie the organs involved in the regulation of ionic economy (trans-epithelial transport system: intestinal villi, renal ducts),

d) dentition, necessary for grinding food and fixed roots in dental beds,

(e) auditory, olfactory and vocal organs.

Thus, the compound prepared by the method of the invention has a favorable biological or therapeutic effect on the above-mentioned organs or tissues of the AGAS system.

This compound also acts on the following AGAS-related functions: radiation protection, wound healing effect, and generally activating mesenchym, protection against the ever increasing risk of infection and contamination of the skin and mucous membranes (formation of lysozyme of wet mucous membranes, activation of ciliary lining in respiratory tract, etc., increased protection against virus and fungal infections.

The compound prepared by the method of the invention is effective against increasingly and substantially with increasing stress effects (e.g., meteorological effects, considerable differences between day and night temperature, increased risk of injury) on shore by stabilizing adaptation syndrome and at the same time averting damage to peripheral tissues by glucocorticoids (for example, damage to the connective tissues, damage to the underlying bone mass, etc.).

Said compound prepared by the process of the invention acts in part directly, in part by controlling the metabolism of vitamin A, by making vitanrine A metabolites of a more polar nature. This effect is comparable to the effect of parathyroid hormone on the 25-hydroxycholecalferol-1-β-hydroxylase enzyme of the renal tubules. This explanation makes it possible to understand the broad pharmacological, biochemical and therapeutic action of χ-L-glutamyltaurine prepared by the process of the invention. The areas of action of this compound are as follows:

A. Vitamin A effects

(a) Pharmacological and biochemical effects

The sulphates labeled with the radioactive element are increasingly incorporated into the bone cartilage of the rat, optionally into the lens of the liver and lung tissue of the chicken embryo; radiolabeled phosphorus is increasingly incorporated into rat bone cartilage; chondroitin sulfate synthesis enhancing effect; a beneficial effect on wound healing or wound healing, optionally exacerbated by administration of cortisone in rats and dogs; vitamin A potentiating effect of hypovitaminoses or hypervitaminoses, experimentally induced in rats and chickens; sedative effect on stress effects caused by rat ulcer; a beneficial effect on mast cell degranulation; lysozyme production-enhancing effect; effects on the management of trace elements (silicon, zinc, copper, manganese, fluorine); a beneficial effect on epithelial formation; escalating effect on alkaline phosphatase activity; action on granuloma vesicle formation induced by the local action of vitamin A; very flat course of the dose / effect curve or change of the effect sign at large doses; activating effect on Golgi; a favorable effect on the formation of calyx cells; Vitamin A concentration-increasing effect

b) Clinical therapeutic effects

Keratoconjunctivitis sicca; Sjorgren's syndrome; rhinolaryngopharyngitis sicca; ozaena; chronic bronchitis; sinobronchitis; mucoviscidosis; constitutional lung disease in young children; periodontitis; susceptibility of the skin and mucosa to infection by viruses and fungi; cortisone antagonist; beneficial effect on wound healing of skin and mucosa during surgery; erosio colli; reduction of olfactory and taste sense caused by itching ¹ .

B. Vitamin A - free effects

a.) Pharmacological and biochemical effects

Effect on blood sugar in terms of transient reduction; escalating effect on phosphaturia; reducing the effect on serum phosphorus; radio-protective effect, an effect reducing the time required to reach the target in experiments with a labyrinth of inactive animals; a dampening effect on experimentally induced fluorine and cadmium toxicosis; escalating effect on cyclic renal emptying induced by odenosine monophosphate; a mitigating effect on the symptoms of experimentally induced lathyrism; decreased sensitivity to histamine; enhancing effect on the activity of the liver enzyme tyrosinamiinotransferase.

b) Therapeutic effects

Slight radiation damage; local lack of dye in the skin; muscular hypotonia; psychoenergetic effect; a beneficial effect on involutionary and gerontological states as well as on mnestic functions; tendency to scar fibroma formation; spondylosis ankylopoetica; musculoskeletal disorders caused by wear; sclerotic fundus; amyloidosis; morphea; fibrocystic mastopathy.

When administering a compound produced by the method of the invention, the treatment time varies greatly. Many diseases (such as rhinolaryngopharyntis sicca) do not show symptoms after two weeks when given orally three times a day. Symptomatic recovery of other diseases (eg periodontitis, Sjórgren's syndrome) requires one to two months, other diseases (eg in the case of spondylosis ankylopoetica), treatment must be carried out for a quarter to a half year.

Any pharmaceutical preventive, cosmetic or veterinary medicament preparations can be prepared in a simple manner from the compound prepared by the process according to the invention. These preparations may contain the active ingredient in combination with other active ingredients. The daily dose of the active ingredient alone is 50 to 500 pg, based on 1 kg of body weight, and is administered in three divided doses.

One tablet contains 2 to 20 µg, preferably. 10 pg of active ingredient and, in addition, biologically inert carriers, for example milk sugar, starch, as well as conventional tabletting aids (granulating and glidants, for example polyvimylpyrrolidone, gelatin, talc, magnesium stearate, aerosil, etc.).

Because of the extremely low dosage, it is expedient to add the active ingredient to the tablet-forming compound before granulation, in the form of a solution. In this way, an even distribution of the active ingredient is achieved. Moreover, the low content of active substance makes it possible to produce the active substance - even in the manufacture of many millions of tablets a year - only on a laboratory scale, at a price which is available to every patient. The active substance is stable, therefore with mbhou tableϊ

209882 those delivered to the shop without stating the expiration date. Active ingredient content for retarding tablets or spansular capsules; it can be between 10 and 20 μ§. For injectable preparations, the effective dose per vial is 5-10 g and the preparation can optionally be prepared in the form of a powder ampoule in which the active ingredient is mixed with an inert water-soluble filler. Parenteral administration can be performed intramuscularly, subcutaneously or intravenously. At this concentration, the active ingredient has no deleterious effect on either the tissue or the vessel walls. The active ingredient may also be administered as an infusion. Suppositories contain 2 to 20, preferably about 10 µg of active ingredient and are made from cocoa butter or synthetic waxes or fats suitable for this purpose. The active ingredient content of the cosmetic ointment or skin healing ointment is 0.1 to 1 / g / g. The ointment matrix may be hydrophilic or hydrophobic and contains conventional ingredients, for example, cholesterol, paraffin, glycerin, lanolin, cocoa butter, linseed oil, etc. The active ingredient may also be formulated as an aerosol formulation, the content of which may also be in the range 0. 1 to 1 Ag / g · In the form of a perlingual tablet, the tablet contains 10 pg of active ingredient, the disintegration time being 0.5 to 1 hour. The sustained release polymers can be prepared, for example, as a suspension and contain 1 to 5 µg of active ingredient per g of polymer. Delayed injectable preparations may be formulated using either high molecular weight polymers or salts of a compound of the invention formed with high molecular weight organic bases (e.g., histone, protamine), each ampoule containing 10-20 µg of active ingredient. Powders for cosmetic or skin treatment are prepared with conventional carriers, for example talc, and contain 0.1 to 1 µg of active ingredient per g of powder. For ophthalmology, the compound produced by the process of the invention is formulated in the form of drops, optionally in the form of an ointment miscible or not miscible with tears; the active ingredient content of these preparations is 0.1 to 1 [mu] g / g. In children, the compound of the invention should be administered at a dose of about 0.3 µg per kg body weight.

Sterile washes are conveniently produced by sterilization filtration.

The desired preventive pharmacological or cosmetic effect of the above-described preparations can be increased and supplemented by numerous combinations. The biologically active additives that may be considered are primarily the following substances and protect their use:

Vitamins A, C, E and K, trace elements, crcisone and its derivatives, progesterone, thyroid hormones, radiometallic and immunosuppressive agents, psychopharmaceuticals, pre-tranquilizers, timoleptics, organic silicon compounds, gerontological agents, cholesterol lowering agents in the blood, oral antidiabetics, anti-inflammatory agents, antihistamines, etc. The dosage of these active ingredients is generally the same as the usual therapeutic dose when using the agents alone.

The compound produced by the process according to the invention can also be used as an additive in nutritional or medicinal mixtures. It causes weight gain, reduces the need for vitamin A, or improves its metabolism. Further, with the compound of the invention, the resorption of trace elements and their level in the blood increases. When a compound of the invention is used as a feed additive, the oral dose is expediently 200 μ% / kg per day. When mixed with the feed, this dose corresponds to a concentration of about 1 to 2 µg / kg or up to about 2 mg / ton of feed, i.e. a concentration of 0.001 to 0.002 ppm. Because of these very low concentrations, the use of the compound of the invention as an additive in feed is extremely economical. Preferably, the compound is added to the vitamin premixes or used in microcapsules containing the compound of the invention together with other necessary feed additives. The compounds according to the invention can furthermore be used in drinking water for cattle feeding, in licking salts or optionally in the form of a spray.

In veterinary medicine, the compound of the invention has similar fields of use as in human medicine, e.g., in skin diseases, wound healing, bone fractures, etc.

The process of the invention for the preparation of? -L-glutamyltaurine of the formula

................................

H 2 N - CH - COOH I (CH 2) 2 i

CO-NH- (CH2) 2-SO3H is carried out by reacting reduced glutathione with taurine in the presence of γ-glutamyltranspeptidase.

The method according to the invention is illustrated in more detail by the following example:

6.05 g (50 mmol) of 2-amino-2- (hydroxymethyl) propane-1,3-diol [TRIS], 4.38 g (75 mmol) of sodium chloride, 2.50 g (20 mmol) of taurine and 6, 14 g (20 mmol) of reduced glutathione are dissolved in 900 ml of water; The pH of the resulting solution was adjusted to pH 8.0 with concentrated hydrochloric acid and the volume of the solution was made up to 1 liter with water. To the solution was added 100 mg of β-glutamyltranspeptidase ferment isolated from pig kidney according to the procedure described by M. Orlowski, A. Meister in J. Biol. Chem. 240, p. 338 (1965). The solution was allowed to stand at 37 ° C for 1 hour, then 100 ml of a 50% trichloroacetic acid solution was added. The precipitate formed is centrifuged and the trichloroacetic acid solution is extracted for 8 hours in a continuous extractor. The solution is then evaporated at 30 ° C under reduced pressure to a volume of approximately 100 ml and then loaded onto a 2.2-50 cm ion exchange column packed with Dowex 1 in a formate cycle. The column adsorbed was eluted with 1 N hydrochloric acid. Upon appearance of hydrochloric acid, 50 ml of solution are obtained. This solution is evaporated first to dryness at 30 DEG C. under 2 kPa, then at room temperature under 1 mmHg. The residue was recrystallized from 80% ethanol to give 250-300 mg of χ-L-glutamintaurine. The melting point is between 219-220 ° C, _{[a] D} 20 = Y i4 _ívodaj ° _{c =} 1.02).

The relative mobility relative to cysteine acid in paper electrophoresis performed at pH 6.5 is 0.73, at pH 1.8 0.53.

_Rf (n-butanol / pyridine / glacial acetic acid / water 15: 10: 3: 12) - 0.19.

Analysis for C7H11N2O6S (molecular weight 254.27):

calculated:

% C, 33.07;% H, 5.55;% N, 11.02;% S, 37.75;

found:

H, 5.76; N, 10.94.

37.53% O, 12.17% S.

Claims (1)

SUBJECT A process for the preparation of χ-L-glutamyltaurine of the formula H 2 N - CH - COOH (CH 2) 2 CO — NH— (CH2) 2 — SO3H · ' OF THE INVENTION characterized in that the reduced glutathione is reacted with taurine in the presence of χ-glutamyltranspeptidase.