FR2566663A1 - MEBENDAZOLE-BASED VERMICIDE AND PROCESS FOR THE PREPARATION OF THIS VERMICIDE - Google Patents

Abstract

THE VERMICIDE, WHICH IS BASED ON METHYL-5-BENZOYL-2-BENZIMIDAZOLE CARBAMATE, HAS A LIPOSOMIC FORM CONTAINING TOTAL EGG YELLOW LIPIDS (0.2 TO 0.6G) AND AN AQUEOUS SOLUTION AT 0.9 OF SODIUM CHLORIDE (20 TO 80ML). CARBAMATE IS PRESENT IN QUANTITIES RANGING FROM 0.5 TO 2.0G. THE PROCESS FOR THE PREPARATION OF THIS VERMICIDE INCLUDES THE FOLLOWING STEPS: A SOLUTION OF TOTAL EGG YELLOW LIPIDS IN CHLOROFORM-METHANOL, METHYL-5-BENZOYL-2-BENZIMIDAZOLE CARBAMATE, EVAPORATES TO DRY TO A DRY RANGING FROM 20C TO 40C, THE AQUEOUS 0.9 SOLUTION OF SODIUM CHLORIDE IS ADDED TO THE DRY RESIDUE WITH CONTINUOUS STIRRING UNTIL LIPOSOMES FORM, AFTER A FREEZING AND THAWING CYCLE INCLUDING 1 TO 12 PERIODS OF - 196C A 35C. VERMICIDE IS USED TO TREAT HYDATICAL DISEASE AND ALVEOCOCCOSIS IN HUMANS AND ANIMALS.

Description

Mebendazole vermicide and process for the preparation thereof

vermi

cide

  The invention relates to a vermicide, in particular for treating

  of the hydatid disease (echinococcosis) and alveococcosis and a process for the preparation of this vermicide, finding application in medicine

human and veterinary.

  The use in the form of pills of mebendazole (methyl-5-benzoyl-2-benzimidazole carbamate) for the treatment of hydatid disease and alveococcosis is known. The disadvantages of mebendazole as a vermicide are of low efficacy in humans. as well as for animals, even during a

  daily treatment for 1 to 5 years; toxicity to the recipient organism

  tor; this preparation causes disturbances in the last months

of pregnancy.

  The methods already known for preparing liposomes which contain

  biologically active substances include the dissolution of

  phospholipids in an organic solvent with a specific concentration, the

  poration, addition of water or buffer and subsequent treatment with ultrasound.

  The liposomes thus prepared are predominantly monolithic

  which renders them of low efficiency to include quantities

  optimum of hydrophobic pharmaceutical substances such as mebendazole.

  The object of the invention is to develop a vermicide, more

  particularly for the treatment of hydatid disease and alveolar

  coccosis, from mebendazole as well as to develop a process for the preparation of this vermicide, allowing intravenous application, and to achieve high efficacy against parasites, as well as a low

  toxicity to the recipient organism.

  This object is achieved by the development of a vermicide, pre-

  liposomal form containing 0.5 to 2.0 g of mebendazole (methyl-5-benzoyl-2-benzimidazole carbamate), 0.2 to 0.6 g of total lipids of egg yolk, and 20 to 20 g of to 80 ml of a 0.9% aqueous solution of

sodium chloride.

  The liposomes thus prepared are homogeneous with respect to their average diameter, which is determined by electron microscopy and is on average 0.4 microns. Liposomes having a

  diameter of 0.3 and 0.5 microns. Liposomes are multilamellar,

  that is to say constituted by several bimolecular layers.

  The process for preparing the vermicide according to the invention comprises the following steps: The chloroform-methanolic solution is added to

  total lipids of egg yolk and methyl-5-benzoyl-2-benzimidazole car-

  Bamate. The mixture is stirred and evaporated to dryness at 20 ° to 40 ° C. Then the 0.9% aqueous solution of sodium chloride is added at room temperature, stirring continuously to hydrate the lipids at the final formation of liposomes. These are subjected to a cycle of freezing and defrosting by passing them

  from 1 to 12 times at temperatures of -196 C and +35 C.

  The aqueous phase: mebendazole: lipid ratio should be: 0.6: 0.1 to 25: 0.5, 0.2. This gives the liposome form of vermicide,

  which allows intravenous application.

  The benefits of the vermicide for the treatment of

  that and alveococcosis according to the invention are its high efficiency, the

  short duration of treatment, its safety. This results in a period of

  vermicide, which allows the application intervals to be spaced

  cation and decrease the total dose. This vermicide is biologically decomposed

  posable. The advantages of the process according to the invention are as follows: it does not require special equipment and rare raw materials; the process is fast and easy to carry out; he ensures the

stable liposomes.

  The invention is illustrated in more detail by the following examples:

Example 1

  The vermicide according to the invention has the following composition: phase

  aqueous (0.9% aqueous solution of sodium chloride) -40 ml; mébenda-

  zole (substance) -1 g; lipid phase (total lipids of egg yolk) -0.4 g.

  Electron microscopy gives the possibility of very precise characterization of liposomes. The liposomes thus obtained represent vesicular, multilamellar structures with double layers of lipid membranes (FIG. It is clearly seen the multitude of lamellae from which the liposome is made. Each coverslip represents a lipidic bimolecular layer having a thickness of about 50Å (50 × 10 m m).

  which corresponds to the thickness of natural biological membranes.

Example 2

  The vermicide according to the invention is prepared in the following manner: 3.3 ml of a solution of the total lipids of the egg yolk, dissolved in a sterile condition, are pipetted under sterile conditions. chloroform and methanol (1%). The initial lipid solution has a concentration of 129 mg / ml. Another 10 ml of chloroform is added and the mixture is stirred vigorously (vortex). Then, 1.0 g of mebendazole is added to the lipids, progressively with continuous stirring. It is evaporated in a rotary evaporator for 5 to 6 min at room temperature. Another 5 ml of chloroform are added and the mixture is evaporated for 1.5 h in the rotary evaporator at a temperature of 35 ° C. (water bath). Any traces of chloroform are removed by nitrogen blowing for 5 minutes. To the substance thus dried, 40 ml of sterile saline (0.9% aqueous solution of sodium chloride) are added. After additional nitrogen insufflation for 5 min, incubate for one hour at room temperature, stirring continuously (vortex) to hydrate the lipid. Now, the freezing is carried out with liquid nitrogen for 2 to 3 minutes, then thawing in a water bath at 500C for 2 to 3 min, with continuous and strong stirring, such as

  so that the temperature of the mixture does not exceed 400C.

Example 3

  The proposed vermicide was tested on ex-

  perimentally by hydatid disease. Animal treatment (sheep A and B) begins 20 months after infection. In order to monitor the effect of the treatment, a blood sample was taken before each sting, in order to study the dynamics of the antibodies according to the method of

  the passive reaction of hemoagglutinationlfig.2Y.

  The general condition of the animals during treatment, the macroscopic image, the pathohistological changes in the parasite and the internal organs of the receptor organism, as well as the ultrastructural changes in the parasite and affected organs were evaluated. Just after the preparation of the mebendazole liposomes included, which are in the form of a white suspension comparable to the milk having a salty taste and a specific odor, they are injected in a dose of 20 ml containing 10 mg / kg of mebendazole, once a week intravenously (vein ugularis). The injection speed of 4 to 5 min. until 5-6 sec. had no influence on how to support the preparation. No effects were observed

  during the entire treatment period. The treatment course

  includes six bites. Treated animals were killed three weeks after the last sting; however an hour before their death, the animals

  underwent an additional sting to follow using the micros-

  electronic copy the localization of liposomes in the organs. A group of control animals, contaminated with the same parasite but which have not been

  treated patients underwent histological and electron microscopic examinations.

  At the end of the experiment, the animals treated with the vermicide were

  in a very good general condition and had gained weight. The title in anti

  body (fig.2) began to decrease gradually after the second sting until the sixth to stabilize then and remain constant until

  the end of the study period (9 weeks). In animals of the group

  Lesser examined under the microscope, parasite cysts of 10 to mm were established, having a characteristic macroscopic image and a specific ultrastructure. In the treated animals, the cysts were 1 to 5 mm and pathohistologically there was complete destruction of the germinal layer and considerable changes in the cuticular membrane (fig.3). In the periocystic wall and sometimes in the hydatid vesicle, the presence of calcium deposits has been established. The internal organs of the treated animals do not deviate from the norm. In terms of ultrastructure, a well-preserved parenchyma was found. A multitude of secondary liposomes with autophagosome features have been observed

  and residual corpuscles. In some cases, residences have been identified

  of liposomes subjected to destruction and phagocytosis. The germinal membrane demonstrates complete destruction and there are residues in the form of vesicles, membranous formations, osmiophilic granules

  etc. (Fig.4). In areas of necrosis, inclusions with

  of liposomes subjected to decomposition.

Claims (2)

  1. Vermicide, especially for the treatment of   Hydatid disease and alveococcosis, based on methyl-5-benzoyl-2-   benzimidazole carbamate, characterized in that it has a lipo-form   containing total lipids of egg yolk and 0.9% aqueous sodium chloride solution in the following quantitative proportions: Methyl-5-benzoyl-2-benzimidazole carbamate 0.5 to 2.0 g Total lipids egg yolk 0.2 to 0.6 g aqueous solution 0.9% sodium chloride 20 to 80 ml, 2. Process for the preparation of the vermicide according to claim 1, characterized in that in a solution of the total lipids of the day of egg in chloroform-methanol is included methyl-5-benzoyl-2-benzimidazole carbamate, evaporated to dryness. at a temperature ranging from 20 ° C. to 40 ° C., the 0.9% aqueous sodium chloride solution is added to the dry residue while stirring continuously until the liposomes are formed, after which a freezing and thawing cycle comprising from 1 to 12 passages from -196 C to +35 C.