RU2193891C2 - Method of industrial allergen preparing - Google Patents

Abstract

FIELD: medicine, allergology. SUBSTANCE: invention relates to method of preparing allergens in aim of their following using for detection of allergic response of patient in professional trail selection. Method involves the development of preparing industrial allergen by extraction of air pollutants in industrial compartment. Taking away air is passed successively through two Bobrov's devices containing 70% ethyl alcohol. Mixture is shaken, centrifuged, dried and obtained precipitated is defatted with, for example, ether followed by dilution of allergen with standard dilution liquid. EFFECT: expanded assortment of measures for control of allergic diseases. 3 cl, 1 dwg

Description

 The invention relates to medicine, namely to technologies for the preparation of allergens for the purpose of their subsequent application for testing workers during employment and during work in various fields of production associated with the release of various gases, dust, etc., in particular rubber industry, in the production of plastics.

 For patients sensitized to allergens, as well as for doctors, information on the content of allergic particles of a certain chemical allergen in the air is important. To identify and determine the number of particles in the air, various methods of collection and counting are used [1].

 It is known that specific diagnostics when applying for a job makes it possible to reduce the incidence at the enterprise by 8 times in some industries over the next three years (N.A. Loogna, M.M. Lacey, 1977). The problem of the widespread use of similar diagnostic capabilities is associated, on the one hand, with polymorphism of chemical industries, and on the other, with the lack of standard industrial allergens.

 One of the common causes of professional BraA is production contact with trimellitic anhydride (TIA), which is used in the plastics industry as a plasticizer. A variety of chemical and biological materials of an industrial and professional nature can cause occupational BraA. Air pollution and its role in the pathogenesis of various diseases is one of the most frequently discussed topics. Most pulmonologists note the important role of pollutants in provoking symptoms of allergic and non-allergic BraA and allergic and vasomotor rhinitis.

 Of great importance for the etiological diagnosis of PBA (occupational bronchial asthma) is atopic, i.e. IgE-type, have skin tests. More often, safe and fairly informative scarification and prick testing methods are used. Allergens such as animal dander, bird feathers, plant pollen are usually available in standard kits; some of them can be used in kind, such as flour. However, there are currently no drugs available for testing industrial allergens.

 Some authors (Platkov EM, 1972) used the application method of skin testing for specific diagnosis of “chemical” asthma and obtained a positive result in 76% of cases [2].

 A known method of isolating an allergen from biological raw materials [3], according to which the drug is obtained by ultrafiltration. The culture filtrate of the tuberculosis pathogen grown on a synthetic nutrient medium and inactivated by heating is passed through filter membranes with a retention capacity of 100 kDa, then the obtained enzyme is concentrated on membranes with a retention capacity of 15 kDa and normalized by ME, sodium chloride, and phenol in 1 ml of solution.

 This method is not applicable to industrial conditions when it is necessary to isolate an industrial allergen.

 There is also known a method of producing allergen for detecting sensitization of the body by fungi of the genus CANDIDA [4], according to which the strain Candida tropikalia VSB 928 is grown in a fermenter in a liquid medium in a cyclic cultivation mode at pH 5.5, oxygen saturation рО not lower than 80% with maintenance of glucose level in the environment of 0.6-0.8%. The separated culture fluid is concentrated by passing through HIP8 brand hollow fibers and separated by membrane filtration. The resulting allergic material with a molar mass of 10 to 300 kDa is subjected to fractionation on DEAE cellulose, followed by screening in an allergostat in homologous and heterologous systems.

 This method is intended for the diagnosis of diseases caused by yeast-like fungi Candida tropikalia, percent feed biomass and cannot be used to obtain industrial allergens.

There is also a method of producing an allergen from bee venom [5], according to which bee venom is isolated and processed, the bee is isolated by electrostimulation of bees, the venom is collected and dried, and the dried venom is processed by dissolving it in water and adding albumin at a concentration of 0.03-0.033%, followed by incubation of the mixture for 18-20 hours at 3-5 ^o C, then centrifuged and dialyzed at 3-5 ^o C.

 This method is also specific and applicable only to bee venom and is not applicable to industrial allergens.

 The problem solved by this invention is the creation of diagnostic industrial allergens of various industries.

 The problem is solved by the method of preparing an industrial allergen, which consists in extracting pollutants from the air of the production room, characterized in that the air drawn through the suction type collector is passed sequentially through two Bobrov apparatus containing 70% ethyl alcohol, shaking the mixture, centrifuging it, drying, degreasing the resulting sediment, for example, with ether, followed by dilution of allergen with a standard diluting liquid.

 Air intake in the production room is carried out for 7 days for 15 minutes every 8 hours of a working day at a height of 1.60-1.70 m with a vacuum cleaner.

 The intake air is passed through two Bobrov apparatuses, first through the first vessel, then through the second vessel, while the first device contains twice as much 70% ethyl alcohol as the second.

 The drawing shows a diagram of the implementation of the proposed method, where 1 is an intake air intake type, for example a vacuum cleaner, 2 and 3 - vessels Bobrov apparatus, 4 - connecting channels.

 The air mixture is taken in the production room with a vacuum cleaner 1 at a height of 160-170 cm from the floor for 7 days for 15 minutes every 8 hours for the most complete representation of the content of allergens in the air. The resulting air mixture is passed sequentially through Bobrov apparatuses 2 and 3 with 70% ethanol contained in them, and first, it is passed through a vessel containing 500.0 ml, then through a vessel containing 250.0 ml of 70% ethyl alcohol. Alcohol mixtures in the first and second vessels are thoroughly shaken, after which they are centrifuged at 2000 rpm for 10 minutes. The resulting grayish-yellow precipitate is dried, degreased with ether. To obtain the final product, the powder is diluted with a standard diluting liquid.

 An allergic examination using the obtained allergen was carried out using an application test according to the following procedure: for a single-layer gauze material of 1 square meter. cm, the test preparation was applied and applied to the skin on the inner surface of the forearm. Then this square was covered with a large square of medical oilcloth measuring 4 square meters. cm, the edges of which were fixed with adhesive tape. After a day, the bandage was removed and the skin reaction after 24 and 48 hours was taken into account. All subjects were warned that in case of severe itching or burning under the dressing it must be removed, the applied substance should be washed off with running water.

 In the Kandrinsky workshop of rubber products of the Nizhnekamsk plant, which produces rubber from sodium butadiene synthetic rubber, industrial allergen was isolated, allergen was made for the application of skin test, allergic examination of workers.

 The study was conducted in 23 of 26 employees in this workshop. In this case, positive skin tests were found in 3 examined. All of them had aggravated allergic heredity: one of them suffered from pollenosis - an allergy to plant pollen (in the period of remission at the time of the study) and two had a history of urticaria. Persons with a positive reaction to an industrial allergen had a period of contact with them under industrial conditions from four (two workers) to six (1 working) months. It is important to note that patients did not notice any clinical manifestations in the workplace, which allowed sensitization to be considered as subclinical.

 Thus, we have developed a method for preparing an industrial allergen that enables early (4-6 months of exposure) detection in workers with subclinical sensitization, which allows timely identification of the occupational risk group and ensuring rational employment. The method is simple to implement, universal in relation to different types of allergens and different industries. The implementation of the method does not require sophisticated special equipment, which ensures its efficiency, expands the possibilities of its application.

Sources of information
1. Allergic diseases. Diagnosis and treatment (translation from English). Roy Patterson, Leslie C. Grammer, Paul. A. Greenberg. Publishing house GOEOTAR, Medicine, M., 2000, Page 98.

 2. V.N. Pytsky, N.V. Andrianov, A.V. Artamasova. Allergic diseases, Ed. III, M., "Triad-X", 1999

 3. USSR AS 1573578, IPC 6 A 61 K 39/35, 35/56, 03/27/96.

 4. RF patent 2031657, IPC 6 A 61 K 39/35, C 12 N 1/14, 03/27/95.

 5. AC USSR 1524226, IPC 6 A 61 K 35/64, 05.27.96.

Claims (3)

 1. A method of preparing an industrial allergen, which consists in extracting pollutants from the air of a production room, characterized in that the air drawn through a suction-type collector is passed sequentially through two Bobrov apparatus containing 70% ethyl alcohol, the mixture is shaken, centrifuged, dried, obtained the precipitate is degreased, for example, with ether, followed by dilution of the allergen with a standard diluting liquid.  2. A method of preparing an industrial allergen according to claim 1, characterized in that the air taken from industrial premises is passed through two Bobrov apparatuses, the first of which contains twice as much 70% ethanol as the second.  3. A method of preparing an industrial allergen according to claim 1, characterized in that the air is taken in industrial premises for 7 days for 15 minutes every 8 hours.