What Is Forensic Toxicology?

Forensic toxicology is an applied discipline that studies the harmful effects of exogenous factors (chemical, physical, and biological factors) on biological systems. It is a science that studies the toxicity, severity, frequency, and mechanism of toxic effects of a chemical substance on an organism, as well as the science of qualitative and quantitative evaluation of toxic effects. It is a discipline that predicts its harm to the human body and the ecological environment, and provides a scientific basis for determining safety limits and taking control measures.

Forensic toxicology

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Forensic toxicology is a study of exogenous factors (

main

The currently accepted definition of toxicology is the science that studies the harm of exogenous chemicals to organisms. Because the purpose of toxicology research is to provide a scientific basis for protecting the health or safety of organisms, toxicology belongs to the nature of the discipline

I. Toxicity

Toxicity is the ability of a substance to cause damage to the body. A relatively small amount of a substance can cause certain damage to the body; a relatively low amount of a substance requires a relatively large amount before it becomes toxic. The level of substance toxicity is only of relative significance. In a certain sense, as long as a certain amount is reached, any substance is toxic to the body; in general, if it is lower than a certain amount, no substance is toxic; the key is the amount of such substance in contact with the body. In addition to the amount of contact between the substance and the body, it is also related to the physical and chemical properties of the substance itself and the way it contacts the body.

Second, the dose

Dose is decided

Forensic toxicology concept

The process of foreign compounds undergoing a series of chemical changes in the body and forming their derivatives and decomposition products is called biological transformation, or metabolic transformation. The resulting derivatives are metabolites. After foreign compounds are biotransformed, some can achieve detoxification and reduce toxicity. However, some can increase its toxicity and even produce teratogenic and carcinogenic effects. Therefore, the metabolic transformation should not be regarded only as a detoxification process, but the metabolic process is dual in toxicity to foreign compounds.

Forensic toxicology response types

Nitric oxide

Oxidation can be divided into two types of oxidation reactions catalyzed by microsomal mixed functional oxidase and non-microsomal mixed functional oxidase.

Microsomes are fragments formed by the endoplasmic reticulum during cell homogenization, and are not independent organelles. The endoplasmic reticulum can be divided into two types: rough surface and smooth surface. Therefore, the microsomes formed are also rough surface and smooth surface, but both contain mixed-function oxidase, which is more active.

Micrososmal mixed function oxidase (MFO), also known as mixed function oxidase or microsomal monooxygenase, can be referred to as monooxygenase for short. In this process, NADPH also needs to provide electrons to reduce cytochrome P-450 and form a complex with the substrate to complete this reaction process.

Mixed-function oxidase is an enzyme system on the endoplasmic reticulum membrane of the cell, and its composition is relatively complicated. The cytochrome P-450 oxidase, also known as cytochrome P-450 dependent monooxygenase, is now known There are reduced coenzyme -cytochrome P-450 reductase. In addition, it also contains microsome FDA-monooxygenase, which is characterized by the absence of cytochrome P-450 and the inclusion of flavin adenine dinucleotide instead of cytochrome P-450 to participate in the monooxygenase reaction. During the oxidation of foreign compounds catalyzed by FAD monooxygenase, NADPH and oxygen molecules are also required.

Many foreign compounds can be catalyzed by mixed function oxidases and oxygenated to form various hydroxyl compounds. The hydroxyl compound will be further decomposed to form various products, so the oxidation reaction may have the following various types:

Aliphatic hydroxylation: also known as aliphatic oxidation, is the oxidation of the penultimate or second carbon atom at the end of the side chain (R) of an aliphatic compound and forms a hydroxyl group.

Aromatic hydroxylation: Hydrogen on the aromatic ring is oxidized. For example, benzene can form phenol, and aniline can form p-aminophenol or o-aminophenol. In the measurement of microsomal mixed function oxidase activity, this reaction can be used, that is, aniline is used as a substrate to be hydroxylated by MFO to form p-aminophenol, and the content is measured to indicate the aniline hydroxylase activity. During the hydroxylation process, o-aminophenol can also be formed.

(3) Epoxidation reaction: A bridge structure is formed between two carbon atoms of a foreign compound, that is, an epoxide. Epoxides are generally only intermediate products and will continue to decompose. But after PAHs, such as benzo (a) pyrene, form epoxides, they can covalently bind with cell biological macromolecules, induce mutations and form tumors.

N-dealkylation reaction: The alkyl group on the oxygen N of the amine compound is oxidized to remove an alkyl group to form an aldehyde or a ketone. Carbamate insecticides, such as carbamide, carcinogen azo pigment cream yellow, and dimethylnitrosamine, can all occur in this reaction. Dimethylnitrosamine can also form a free methyl group [CH3 +] after N-dealkylation, which can cause guanine methylation (or alkylation) on nucleic acid molecules in the nucleus to induce mutation or carcinogenesis.

O-dealkylation and S-dealkylation reaction: Similar to N-dealkylation reaction, but the alkyl group connected to oxygen atom or sulfur atom is removed after oxidation.

O-dealkylation can occur in p-nitroanisole. The latter is catalyzed by microsomal mixed function oxidase, and the content of p-nitrophenol formed is determined, which can represent the activity of mixed function oxidase.

N-hydroxylation reaction: The hydroxylation is carried out on the N atom, for example, aniline and carcinogen 2-acetamidinium can occur. Aniline undergoes N-hydroxylation to form N-hydroxyaniline, which can oxidize hemoglobin to methemoglobin.

Alkyl metal dealkylation reaction: Tetraethyl lead can remove one alkyl group under the catalysis of mixed function oxidase to form triethyl lead. With this, tetraethyl lead can exhibit toxic effects in the body.

Desulfurization reaction: Desulfurization reactions often occur in many organophosphorus compounds. In this reaction, sulfur atoms are oxidized to sulfate and fall off. Such as parathion oxidative desulfurization into paraoxon, toxicity is enhanced.

The oxidation reaction catalyzed by non-microsomal mixed functional oxidase. In the cytosol, plasma and mitochondria of liver tissue, there are some less specific enzymes that can catalyze the oxidation and reduction of certain foreign compounds, such as alcohol dehydrogenase. Aldehyde dehydrogenase, catalase, xanthine oxidase, etc.

Hepatocyte cytosol contains monoamine oxidase and diamine oxidase, which can catalyze the oxidation of amines to form aldehydes and ammonia. The oxidation reaction catalyzed by diamine oxidase mainly involves the formation of biological amines in the body, and has less relationship with the metabolism and conversion of foreign compounds .

Co-oxidation during prostaglandin biosynthesis In the oxidation reaction of foreign compounds, in addition to the oxidation reactions catalyzed by the aforementioned microsomal mixed-function oxidase and non-microsomal mixed-function oxidase, an oxidation reaction has been observed in recent years. In the prostaglandin biosynthesis process, some foreign compounds can be simultaneously oxidized, which is called a co-oxidation reaction.

Two reduction reaction

Foreign compounds containing nitro, azo, and carbonyl groups, as well as disulfides and sulfoxides, can be reduced in the body. For example, both nitrobenzene and azobenzene can be reduced to form aniline. Carbon tetrachloride can be catalyzed and reduced by NADPH-cytochrome P-450 reductase in the body to form chloroform free radicals (CCl3 +), which will destroy the lipid structure of liver cell membrane, cause liver steatosis and necrosis. Arsenic in pentavalent arsenic compounds can also be reduced to trivalent arsenic. Trivalent arsenic compounds have higher solubility in water, so they are more toxic than pentavalent arsenic compounds.

Trihydrolysis

Many foreign compounds, such as esters, amides, and phosphate substituents containing ester linkages are extremely prone to hydrolysis. There are many hydrolases in plasma, liver, kidney, intestinal mucosa, muscle, and god tissues, as well as in microsomes. Esterase is a widespread hydrolase. Esterase and amidase can hydrolyze esters and amines, respectively.

The hydrolysis reaction is the main metabolic mode of many organophosphorus pesticides in the body, such as dichlorvos, parathion, dimethoate, and malathion. The toxicity decreases or disappears after hydrolysis. Some insects are resistant to malathion, that is, because of its high carboxylase activity in the body, malathion is easily deactivated. In addition, pyrethroid insecticides are also detoxified by hydrolytic enzyme-catalyzed degradation.

Quadruple reaction

A binding reaction is a biosynthetic reaction of a foreign compound entering the body with certain other endogenous compounds or groups during metabolism. Especially foreign organic compounds and their metabolites containing hydroxyl, amino, carbonyl and epoxy groups are most likely to occur. The products formed by the combination of foreign compounds and their metabolites with certain endogenous compounds or groups in the body are called conjugates. In the binding reaction, coenzyme and transferase are required and metabolic energy is consumed. The source of the so-called endogenous compound or group is the product of the normal metabolic process in the body, and the endogenous compound must be an endogenous compound to participate in the binding reaction, which cannot be performed directly by the in vitro importer.

Foreign compounds can undergo binding reactions directly during metabolism, or they can go through the above-mentioned first-stage biological transformation reactions (first-phase reactions) such as oxidation, reduction, or hydrolysis, and then undergo binding reactions (second-phase reactions). Generally, Under the circumstances, through the binding reaction, on the one hand, certain functional groups on the foreign compound molecule can lose their activity and toxicity; on the other hand, most of the foreign compounds can increase their polarity, reduce their fat solubility, and accelerate their binding reactions. Excretion process by the body.

According to the mechanism of the binding reaction, the binding reaction can be divided into the following types:

Glucuronide binding is probably the most common binding reaction, mainly foreign compounds and their metabolites bind to glucuronide. The source of glucuronic acid is uridine diphosphate glucose (UDPG) generated during the metabolism of sugars. UDPG is oxidized to produce uridine diphosphate glucuronic acid; UCPGA is the donor of glucuronic acid. Under the action of uronic acid transferase, it is combined with the hydroxyl, amino and carboxyl groups of foreign compounds and their metabolites, and the reaction product is -glucuronide. Glucuronic acid must be an endogenous metabolite, and binding reactions cannot be performed directly by an in vitro importer.

The glucuronic acid binding effect is mainly carried out in liver microsomes. In addition, it can also occur in the kidney, intestinal mucosa and skin. Foreign compounds are excreted with bile after a binding reaction in the liver. But sometimes part of it is in the lower part of the intestinal tract, which can be hydrolyzed by -glucuronidase in the intestinal flora, and this foreign compound can be reabsorbed and enter the enterohepatic circulation, so that its residence time in the body is prolonged.

Sulfuric acid binds foreign compounds and their alcohol, phenol or amine compounds in metabolites, which can be combined with sulfuric acid to form sulfate esters. The source of endogenous sulfuric acid is a metabolite of sulfur-containing amino acids, but it must be activated by adenosine triphosphate to become 3'-adenosine-5'-phosphate sulfate (PAPS), and then interact with phenols under the action of sulfotransferase. , Alcohols or amines are combined into sulfate esters. The combination of phenol and sulfuric acid is more common.

The sulfuric acid binding reaction is mostly performed in liver, kidney, gastrointestinal and other tissues; due to the limitation of the source of sulfuric acid in the body, it cannot be provided sufficiently, so it is less than the glucuronic acid binding reaction.

In general, the original toxicity of foreign compounds can be reduced by the sulfuric acid binding reaction. However, some foreign compounds have higher toxicity after sulfuric acid binding reaction. For example, a carcinogen belonging to aromatic amines, 2-acetylaminohydrazone (FAA or AAF for short), undergoes N-hydroxylation reaction in the body to form N-hydroxy-2-acetamidofluorene, and its hydroxyl group can be combined with sulfuric acid to form Sulfate. This AAF sulfate has strong carcinogenicity, which is stronger than AAF itself. This reaction occurs in rats, mice and dogs. However, some animals lack sulfatase in the liver to form sulfate esters.

Glutathione binds toxic metals and epoxides in the body and can be detoxified by binding to glutathione. The glutathione binding reaction is catalyzed by glutathione transferase. Glutathione is contained in the liver and kidney, and the cytosol content of liver cells is large. In recent years, it has been found in liver microsomes. Microsomal glutathione transferase is in direct contact with foreign compounds, which may be more important in the glutathione binding reaction.

The binding of glutathione to epoxide is very important. Many foreign compounds, such as many carcinogens and liver poisons, can form epoxides in the body, and most of these epoxides have a strong damaging effect on cells. For example, brominated benzene is metabolized into epoxide, and bromobenzene epoxide is a strong liver poison that can cause liver necrosis; but after being combined with glutathione, it will be detoxified and excreted. There is a limit to the production and storage of glutathione in the body. If a large number of epoxides are formed in a short period of time, glutathione depletion may occur and still cause serious damage.

Glycine binds to some foreign compounds containing carboxyl groups, for example organic acids can bind to amino acids. The nature of this binding reaction is a peptide binding, and the most common binding with glycine, in fact, other amino acids can also perform this binding. For example, toluene is metabolized in the body to produce benzoic acid, which can be combined with glycine to form hippuric acid and be excreted from the body. Cyanohydrogen is bound by cysteine and excreted by saliva and urine.

Acetyl binds aromatic amines in foreign compounds. For example, aniline can react with acetyl-CoA through its amino group and acetyltransferase catalyzes the aromatic amines to form their acetyl derivatives. In addition, aliphatic amines have a similar response. Acetase A is derived from the metabolites of sugar, fat, and protein.

methyl-binding biological amines in the body and methyl-binding reaction, also known as methylation. The methyl group is derived from methionine. The methyl group of methionine is activated by ATP to become S-adenosylmethionine, which is then catalyzed by methyltransferase to combine biological amines with methyl groups and be detoxified and excreted. In the detoxification of foreign compounds, methyl bonding is not important.

Factors affecting forensic toxicology

One species difference and individual difference

The speed of biotransformation of the same foreign compound can be quite different in different animals. For example, the biological half-life of aniline is 35 minutes in mice and 167 minutes in dogs. The metabolism of the same foreign compound in animals of different species can be completely different. As mentioned earlier, N-2-acetamidinium can be N-hydroxylated in rats, mice, and dogs, and then combined with sulfuric acid to form sulfates, which exhibits a strong carcinogenic effect; while N- generally does not occur in guinea pigs. It is hydroxylated, so it cannot be combined into sulfates, and has no carcinogenic effect or very weak carcinogenic effect.

The individual differences of foreign compounds in the biotransformation process in vivo are also manifested in the activity of certain enzymes involved in metabolism in each body. For example, arylhydrocarbon hydroxylase (AHH) can hydroxylate aromatic hydrocarbon compounds and produce carcinogenic activity, and their activity varies significantly among individuals. With the same amount of smoking, people with higher AHH vitality are 36 times more likely to develop lung cancer than those with low vitality; those with moderate vitality in the body are 16 times more likely to develop lung cancer than those with low vitality.

Inhibition and induction of metabolizing enzymes of two foreign compounds

Inhibition of the biotransformation of one foreign compound can be inhibited by another compound, and this inhibition is related to enzymes that catalyze biotransformation. The enzyme systems involved in biological transformation generally do not have high substrate specificity. Several different compounds can be used as substrates of the same enzyme system, that is, the biological transformation process of several foreign compounds is catalyzed by the same enzyme system. . Therefore, when one foreign compound appears or increases in the body, it can affect the catalytic effect of an enzyme on another foreign compound, that is, the competitive inhibition of the two compounds.

Induction of some foreign compounds can increase the activity of catalytic enzymes or increase the content of enzymes in certain metabolic processes. This phenomenon is called the induction of enzymes. Any compound with an inducing effect is called an inducer. The result of the induction can promote other foreign compounds. The biotransformation process makes it enhanced or accelerated. During the induction of microsomal mixed-function oxidase, proliferation of the endoplasmic reticulum of the slippery surface was also observed; enhanced enzyme activity and the promotion of other compounds' metabolic transformation were related to this.

Triple metabolic saturation

The saturation state of a foreign compound in the body's metabolism has a considerable impact on its metabolism, and therefore its toxic effect. For example, brominated benzene is first converted into brominated benzene epoxide with liver toxicity in the body; if the input dose is small, about 75% of the brominated benzene epoxide can be converted into a glutathione conjugate, and It is excreted in the form of bromophenyl sulfide; however, if a larger dose is entered, only 45% of the side can be excreted in the form described above. When the dose is too large, because the amount of glutathione is insufficient, and even the glutathione is depleted, the binding reaction is reduced, so the reaction of unbound bromobenzene epoxide with DNA or RNA and protein is enhanced, showing Toxic effect.

Four other influencing factors

Mainly manifested in age and gender and nutritional status. The nutritional status of protein, ascorbic acid, riboflavin, vitamin A and vitamin E can all affect the activity of mixed function oxidase in microsomes. If the protein supply is insufficient in animal experiments, the microsomal enzyme activity decreases. When ascorbic acid is deficient, the hydroxylation of aniline is weakened. Lack of riboflavin can reduce azo compound reductase activity and enhance the carcinogenic effect of carcinogen cream yellow. The decrease in the activity of the above enzymes may cause the conversion process of foreign compounds to weaken or slow down.

The effect of age on the metabolic conversion of foreign compounds is manifested in that liver microsomal enzyme function has not yet matured in newborns and adolescents, and has begun to decline after old age. Its functions are lower than those of adults. weak. For example, 30 days after the rat was born, the mixed function oxidase of liver microsomes reached the adult level, and then began to decline after 250 days. The glucuronide-binding response weakened in older animals, but the monoamine oxidase activity of rats increased with age. In general, the ability of young and old organisms to metabolize and convert foreign compounds is weaker than that of adults, so the harmful effects of foreign compounds are also strong.