RU2108580C1 - Method for determining fish blood toxicity - Google Patents

Abstract

FIELD: medicine. SUBSTANCE: method involves mixing fish blood serum with alpha amino acetic acid solution and holding it for some time. Then crystallographic examination is carried out on obtained changes. Crystallization centers of various forms, volumes and length as well as formations in the form of deposit being produced, toxicity is diagnosed. EFFECT: simplified method. 4 dwg, 1 tbl

Description

 The invention relates to the field of toxicology and allows you to set the toxicity of the liquid media of the fish organism.

 A known method for detecting the toxicity of liquid media of an organism of fish by determining the histamine content in fish products, which is based on measuring the intense fluorescence of a derivative obtained by the interaction of histamine with o-phthalaldehyde [1].

 The disadvantage of this method is its technical complexity of use (it takes a long time to prepare the sample, highly qualified personnel, sophisticated equipment and severely deficient reagents).

 A known method for determining the content in the blood serum of fish of total protein, protein fractions, which are separated by electrophoresis on paper. A change in the ratio of albumin and globulin and their content in the blood serum indicates fish poisoning [2].

 The disadvantage of this method is the technical complexity of execution (it takes a long time to separate proteins into fractions, as well as acute deficiency reagents).

 The crystallographic method has found application in biology and medicine, in particular for forensic chemical analysis, qualitative determination of drugs, diagnosis of the degree, malignancy of the tumor process, cerebrovascular accident, etc.

 The objective of the invention is to develop a method for determining the toxicity of blood fish by providing a higher detection sensitivity. This result is achieved by the incubation of blood serum with a crystal-forming substance followed by crystallization of the mixture, including the use of an alpha-aminoacetic acid solution, and, when crystallization centers of various shapes, volumes and lengths, as well as plaque formations appear, determine the toxicity of the blood of fish.

The method is as follows. To 5 ml of filtered aqueous solution of alpha-aminoacetic acid / glycine / add 0.2 ml of blood serum of fish, mix thoroughly and transfer the mixture to Petri dishes, which are placed in a thermostat at 37 ^o C or left in air until the contents of the cup are completely dried. After incubation for 5-6 hours, a crystallographic examination of the obtained changes is carried out, crystallography of pure glycine is used as a control, and when crystallization centers of various shapes and volumes, lengths, and also formations in the form of plaque appear, toxicity of fish blood is determined.

 Examples of the method. For comparison, in FIG. 1 shows the crystallography of pure glycine (scale 1: 1). It consists of soft polyhedral single crystals with sizes of 4-6 mm, distinguishable visually, and there are also single crystals of the parallelepiped type up to 30 mm long. Figure 2 presents crystallography obtained from serum of carp, caught on 05.25.94, in the Chardarya reservoir. To 0.2 ml of fish blood serum add 5 ml of alpha-aminoacetic acid solution, after which they are incubated in air until the mixture crystallizes. Three fields are clearly visible in the crystallographic picture, consisting of: the first part consists of crystalline formations; the second part is like sand; the third part has the form of plaque, which is a patterned lattice up to 40 mm in size.

 In the first part, there are 4–5 crystallization centers in the form of a leaf with veins, dividing these formations into two parts and veins extending from them to the sides. Characteristic is the presence of single crystals of a polyhedral shape with sizes of 4-5 mm and crystalline formations up to 20 mm long, as well as groups of crystals in the form of spears or swings and the presence of formations in the form of branches.

 Figure 3 presents crystallography obtained from serum of carp, caught on 24.09.94, weighing 1450 g in the Chardarya reservoir. To 0.2 ml of blood serum add 5 ml of a solution of alpha-aminoacetic acid, followed by incubation in air until the mixture crystallizes. Three regions are observed in the crystallographic picture. In the first section, the formation of crystals of various shapes is clearly visible, in the second - the formation of sand, as it were; on the third - plaque in the form of a patterned lattice up to 65 mm long. In the first section, 2–3 crystallization centers are observed, having the form of half-radials, up to 5–6 mm long, and crystal formations having the form of a swing or spear, up to 8–10 mm. The single crystals have a size of 4-6 mm in the form of a parallelepiped with a length of 8-10 mm. One can see many formations of amorphous form.

 Figure 4 presents crystallography obtained from the blood serum of crucian carp caught on 05.28.94 in the Chardarya reservoir. To 0.2 ml of fish blood serum add 5 ml of alpha-aminoacetic acid solution, after which they are incubated in air until crystals appear. A large number of single crystals of a polyhedral shape of 4-6 mm, as well as single crystals of the parallelepiped type up to 40 mm in length, are visible in the field of view. The height of individual single crystals reaches 2–3 mm, a separate section of up to 50 mm is a lattice built of crystals intersecting at different angles. At the same time, crystallization centers in the form of plaque, which are a formation in the form of "frost", are clearly visible. Crystallization centers built from crystals in the form of a “block” or semi-radial are clearly visible. Separate crystalline formations are in the form of "spears" and "swings". A high degree of "structuring" of the crystalline picture is noted.

 Based on a comparison of the crystalline pattern of alpha-aminoacetic acid used as a control, we can draw a conclusion about the toxicity of the studied fish blood samples.

 To prove the revealed toxic changes in the crystal picture of the liquid media of the fish organism, a study was made of the content of average molecular peptides (SM) of fish blood. It is known that with endogenous intoxication of the human and animal body, a wide range of metabolites with a toxic effect is detected. An integral indicator of endogenous intoxication is considered the level of compounds of groups of medium molecules (SM) [3].

 The content of SM in the blood of carp caught from the reservoir of the Chilik pond farm on March 22, 1994 was taken as a conditional norm. The content of cm in the blood of 5 examined adult fish weighing 950 ± 109 g was 0.051 ± 0.003 U.E. (see table). In contrast to the control, the content of medium-molecular peptides in the blood of carp (adult fish weighing 1510 ± 129 g /), caught in the spring, averages 0.14 ± 0.02 U.E. The level of SM peptides in carp (adult fish weighing 1846 ± 140.0 g) examined in the fall (September 24, 94) is 0.283 ± 0.032 U.E. In the blood of crucian carp (average weight 517 ± 13.5 g), captured on 05/28/94, the magnitude of the peptides was 0.143 ± 0.037 U.E.

 It is seen that the content of SM peptides in the blood of fish increases in the fall (the water level in the reservoir drops sharply) and depends on the level of pollution of the reservoir with toxic substances that accumulate in the organs and tissues of the fish and cause endogenous intoxication of the fish organism. When comparing with the conventional norm, the content of SM peptides shows that their level increases 3 times when examining fish in the spring, and in the autumn it increases almost 6 times. Based on a comparison of the serum crystallograms of the examined fish with the detected level of endogenous intoxication, determined by the content of SM peptides, it can be concluded that the presented method for determining toxicity reveals endogenous intoxication of the fish organism, which manifests itself in detectable blood toxicity by crystallogram.

 The method was tested in the laboratory of fish diseases of the Kazakh Research Institute of Fisheries of the Academy of Agricultural Sciences of the Republic of Kazakhstan on 113 fish.

 The method for determining the toxicity of blood fish is highly sensitive, fast (2 - 5 hours), indicative and informative, can be used to protect the environment.

Sources:
1. Temporary hygiene standards and a method for determining the content of histamine in fish products. Ministry of Health, M. 1986.

 2. Metelev V.V., Kanaev A.I., Dzasokhova N.G. Water toxicology M .: Kolos, 1971, p. 72.

 3. Nikolaychik V.V., Bychko G.N., Pilotovich V.S. and others. On the use of gel filtration to determine the severity of intoxication and the effectiveness of detoxification procedures. Method. letter - Minsk, 1983.

Claims (1)

 A method for determining the toxicity of fish blood by analyzing the blood serum of fish, characterized in that during the blood analysis, the serum is mixed with a solution of alpha-aminoacetic acid and the presence of blood toxicity is judged by the appearance of crystallization centers of various shapes, volumes and lengths, as well as formations in the form of plaque.

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