- Email: [email protected]
Evaluation of the efficacy in cosmetic products safety: Comparison with biochemical substrates
Regulatory Toxicology and Pharmacology 104 (2019) 56–58
Contents lists available at ScienceDirect
Regulatory Toxicology and Pharmacology journal homepage: www.elsevier.com/locate/yrtph
Evaluation of the efficacy in cosmetic products safety: Comparison with biochemical substrates
T
Yaşar Demira,∗, Meryem Uckayab, Nazan Demirb a b
Ataturk University, Faculty of K.K Education, Department of Chemistry Education, 25200, Erzurum, Turkey Mugla Sitki Kocman University, Faculty of Science, Department of Chemistry, 48000, Kotekli-Mugla, Turkey
A R T I C LE I N FO
A B S T R A C T
Keywords: Cosmetic Safety HATKO rules Biochemical substrate Enzyme
Chemicals used in cosmetics must interact with the enzymes for their consumption after entering our bodies. The area at which the interaction realizes on the enzyme is known as the active center. This center is three dimensional and optically active. Considering the properties of the active regions, it is believed that the determination of the geometric properties of the chemicals may contribute to the safety evaluation of the chemical products. Obtainment of toxicological data of chemicals is a long and difficult process. It is an impossible process as the animal experiments have been prohibited. Since there are large number of chemical compounds available, it is not possible to conduct toxicological evaluation on all of them. Therefore, it is important to estimate whether chemicals are toxic through using molecular formulas. In this study, the similarities and differences between the unwanted chemicals used in cosmetic products and the geometric structures of the chemicals used in metabolism were determined by using Group Theory. The aim of the study is to estimate whether the chemicals will be toxic by taking advantage of point-group determinations. Molecular formulae of chemicals prohibited in cosmetics and substrates used in biological systems were used as materials. The point groups of the chemical drawing program was molecules were determined using these formulas. ChemDraw Professional 17 used to draw the formula structures of the molecules. Campus Licensed version of the program provided by application which was developed to examine the molecules acAtatürk University was utilized. An Excel cording to the Group Theory was also used.
1. Introduction Group theory explains the spectroscopic properties and molecular orbitals of the compounds by using the structures of molecules. Group Theory is used to explain the experimental results rather than revealing unknown properties. The catalytic centers of enzymes that catalyze chemical reactions in living systems are three-dimensional. Therefore, molecules with appropriate geometries may approach to active centers and each enzyme has effects on certain functional groups (David L. Nelson, 2013). The chemicals used in cosmetics interact with enzymes after entering our body, in order to be used or destroyed. Reactions catalyzed by enzymes occur in these active centers. The active center is a special region of the enzyme with substrate binding feature. Knowing the geometric characteristics of active centers and chemicals can contribute to the safety assessment of cosmetics. Cosmetic product includes any substance or mixture intended to be placed in contact with the external parts of the human body such as
∗
epidermis, hair system, nails, lips, and external genital organs or with the teeth and the mucous membranes of the oral cavity with a view exclusively or mainly cleaning them, perfuming them changing their appearance, protecting them, keeping them in good condition or correcting body odors (Cosmetics Directive, 2009). Cosmetic product components are any synthetic or natural substance or mixture used in the structure of cosmetic product other than perfume and aromatic substances (Sağlık Bakanlığı, 2005a). The safety assessment is an assessment report that must be made on the finished product, considering the toxicological properties, chemical structure and exposure levels of the components of a cosmetic product, the specific exposure characteristics of the target audience or the area for which the product is to be applied. (Sağlık Bakanlığı, 2005b). Toxicology is a branch of science that studies the negative effects of chemicals on living organisms. All foreign substances i.e. xenobiotics, which are not required for the normal metabolism of the organism rather taken from outside through various ways, are included in the area of interest of toxicology. Toxicity is toxic effect of xenobiotics (Vural,
Corresponding author. E-mail address: [email protected] (Y. Demir).
https://doi.org/10.1016/j.yrtph.2019.03.001 Received 2 October 2018; Received in revised form 14 February 2019; Accepted 1 March 2019 Available online 08 March 2019 0273-2300/ © 2019 Elsevier Inc. All rights reserved.
Regulatory Toxicology and Pharmacology 104 (2019) 56–58
Y. Demir, et al.
2.1. Selection of molecules
Table 1 Quantity and availability percentage of point groups in the prohibited and nonprohibited chemicals, as analyzed according to Group Theory. Group
Prohibited
C1 Cs C∞V D∞V Ci
In the selection of chemicals that are forbidden to use, the list of substances that should not be included in Cosmetic Products is used. Of the 1233 chemicals in this list, 621 chemicals which are not polymeric and inorganic were selected. In order to make a comparison (non-harmful molecular model), 81 molecules used in living systems were selected. These molecules are amino acids, carbohydrates (straight chain structures, α- and βHaworth structures), fatty acids, citric acid cycle molecules, glycolysis pathway molecules and compounds used in other metabolic ways.
Non-prohibited
Count
%
Count
%
252 196 139 30 4
40.51 31.51 22.35 4.82 0.64
18
21.95
61 2
74.39 2.44
2.2. Drawing of molecular structures
Table 2 Appearance Frequencies of the determined point groups according to Group theory (GSx). Point Group
Appearance Prevalence (GSx)
C1 Cs C∞v D∞v Ci
0.542 0.000 3.328 0.506 0.000
Molecules selected for analyses were drawn with ChemDraw Professional 17 program and recorded with JPEG extension. JPEG image files were used in the studies conducted according to Group Theory. 2.3. Rules concerning the examination of molecules While the molecules were analyzed according to Group Theory, a set of rules known as HATKO rules was used to provide standardization (Uçkaya et al., 2016).
2005). It determines the amount of toxicity of a chemical entering the living system, that is to say, the main factor determining the toxicity is the dosage (Borzelleca, 2000). The term LD50 is used as an acute toxicity unit to indicate how toxic a substance is. NO(A)EL, LO(A)EL, SED, LC50 and MoS are other toxicity units. The pharmacological effects, toxicities and pharmacokinetic properties of the chemical substances that are synthesized or isolated from natural sources should be examined in appropriate experimental animals. The basic concept as the basis for these tests and experiments is that the drug would cause the same effects it caused in the experimental animals (Büken Ö., 2000). According to the eighth article of the Universal Declaration of Animal Rights, which was promulgated in Paris on 15 October 1978, conducting experiments causing physical or psychological suffering on animals are violation(s) of animal rights. This shall apply for any and all medical, scientific, commercial and etc. experiments. After 11 March 2013, the sale of any cosmetic and personal care products tested on animals was prohibited in the European Union. This also applies to products imported from the non-member countries (Universal Declaration of Animal Rights, 2016). In our opinion, since these chemicals are forbidden to be tested on animals, these experiments are carried out on humans, especially on women being the main consumers. Therefore, the development of toxicological estimation techniques for human and community health will be very beneficial for public health in the longer term. Cosmetic products became widespread in 1900s. The conscious use cycle of these products started after the Second World War. The green movement, started in Western Europe in the late 1960s, has led to the tendency of natural origin cosmetics in the cosmetic industry. Due to this trend, natural raw materials and finished products were developed. Then, it was seen that natural raw materials or finished products could be harmful to human health. Therefore, it was concluded that safety and efficacy assessment must be performed for all cosmetic products of natural or synthetic origin (Bergişadi et al., 2014; Alğın Yapar and İnal, 2012; Kaymak and Tırnaksız, 2007).
2.4. Programs used in the analysis of molecular structures 2.4.1. Molecular analysis according to group theory used in excel applications The Excel application used in the analysis was developed by Yaşar Demir and used to analyze whether there are inversion point, axes, planes and reflection-rotation points available in the molecules. 2.5. Determination of point groups of molecules In defining the point groups, point group determination table included in Inorganic Chemistry books is used. (Shriver et al., 1994). 3. Results and discussion As a result of the investigations and researches, the point groups of the molecules according to Group Theory were determined separately and the results obtained were discussed. 3.1. Point group analysis A total of 702 molecules were analyzed, 621 of which were prohibited molecules and 81 of them were not prohibited ones. At the end of these studies, the following data have been obtained. In a review of this table to see the difference between the groups, it is seen that the prohibited molecules are clustered in five different point groups and the non-prohibited molecules are clustered in three different point groups. Cs and Ci groups are seen as groups unique to the prohibited molecules. C1, C∞V and D∞V point groups are seen in both the prohibited and non-prohibited groups. According to intra-group analysis, the most covalent group of molecules in the prohibited group is the C1 group while it is C∞V group in the non-prohibited group. Cs point groups have never been found in non-prohibited molecules. This result indicates that there are no intramolecular symmetries of the substrates. Substrate molecules do not have mirror planes. The absence of the point group Ci indicates that the non-prohibited molecules do not have symmetry points in the molecule. The fact that the C1 point group was present in both clusters indicates that all the molecules examined have at least one axis of rotation.
2. Materials and methods In this study, 621 molecular formulae which are prohibited to be used in cosmetic products and 81 formulae of chemical substances used in living metabolism have been used. Structures of these formulas are drawn and point groups are determined according to Group theory. 57
Regulatory Toxicology and Pharmacology 104 (2019) 56–58
Y. Demir, et al.
order to determine the point groups.
The fact that the D∞v element is available in both groups shows that some of the prohibited and non-prohibited molecules have one main axis, one horizontal axis and one horizontal plane perpendicular to the main axis. Other point groups were never seen in both the prohibited and nonprohibited groups. When we compare the presence quantity of these point groups to each other (Prohibited/Non-Prohibited), we get the Prevalence of Appearance (GSx) of these properties. The appearance frequencies of the point groups are given in the table below. In the five groups observed, only the prevalence of C∞V is significantly higher in the non-prohibited group. The prevalence of other groups of points is higher in the prohibited cluster. The Cs and Ci group are seen only as a feature of the prohibited cluster. The Cs property is important because it defines a plane and the Ci property is important since it defines the intramolecular symmetry. The fact that there is a plane of the molecule and that its symmetries (i.e. the Ci property) are noticeably high mean that identifiable symmetrical geometric structures of the prohibited molecules may be present. In addition, non-presence of these characteristics in the non-prohibited cluster supports the idea that the active regions of the enzymes work as a result of the interaction of the functional groups around the active center (compatibility model as a result of interaction) rather than the suitability of the geometric structure (key lock model).
Transparency document Transparency document related to this article can be found online at https://doi.org/10.1016/j.yrtph.2019.03.001. References Alğın Yapar, E., İnal, Ö., 2012. Nanomaterial sand cosmetics. J. Fac. Pharm. Istanbul 42 (1), 43–70. Bergişadi, N., Erdem, O., Çadırcı, E., Alğın Yapar, E., Aktaş, Y., Erdal, M.S., Demirel Özel, A., 2014. Critical analyze of safety cosmetic components' safety evaluation carried pursuant to European standards. In: Kadıoglu, Y. (Ed.), Safety Evaluation of Cosmetics in Europe. Cosming.com. Publications, İstanbul, pp. 64–109. Borzelleca, J.F., 2000. Profiles in toxicology paracelsus. Her. Mod. Toxicol. 23, 2–4. Büken, N.Ö., 2000. Clinical Drug Researches in Term of Experimental Subject and Patient. A.U. Unpublished Doctoral Thesis of Institute of Medical Sciences, Ankara. Kaymak, Y., Tırnaksız, F., 2007. Unwanted effect of cosmetic products. Dermatose 6 (1), 39–48. Nelson, David L., Cox, Michael M., 2013. Lehninger Biyokimyanın İlkeleri, (5. Baskı). Palme Yayıncılık, Ankara, pp. 183–186. Sağlık Bakanlığı, T.C., 2005a. Kozmetik Yönetmeliği (Resmi Gazete Sayısı: 25823), Birinci Bölüm, File:. Sağlık Bakanlığı, T.C., 2005b. Kozmetik Yönetmeliği (Resmi Gazete Sayısı: 25823), Üçüncü Bölüm. Shriver, D.F., Atkins, P.W., Langford, C.H., 1994. Molecular shape and symmetry. In: Inorganic Chemistry, second ed. Oxford University Press, Oxford, pp. 105–145. The European Parliament and The Council, 2009. Regulation (EC) No 1223/2009 of 30 November 2009 on Cosmetic Products Section One Article 2. 30, 11. Uçkaya, M., Uçkaya, F., Demir, N., Demir, Y., 2016. Evaluation of the efficiency and safety in cosmetic products. Int. J. Pharm. 499, 295–300. Universal Declaration of Animal Rights, 2016 Article 8. Vural, N., 2005. Toxicology. Ankara University, Publications of Pharmacy, Ankara Faculty No:73.
4. Conclusion In accordance with the results of the study we conducted, it is seen that it is possible to determine the toxicological characters of the chemicals. In this determination, it made sense to compare the harmful chemicals with the chemicals (substrates) used in the living systems in
58
Contents lists available at ScienceDirect
Regulatory Toxicology and Pharmacology journal homepage: www.elsevier.com/locate/yrtph
Evaluation of the efficacy in cosmetic products safety: Comparison with biochemical substrates
T
Yaşar Demira,∗, Meryem Uckayab, Nazan Demirb a b
Ataturk University, Faculty of K.K Education, Department of Chemistry Education, 25200, Erzurum, Turkey Mugla Sitki Kocman University, Faculty of Science, Department of Chemistry, 48000, Kotekli-Mugla, Turkey
A R T I C LE I N FO
A B S T R A C T
Keywords: Cosmetic Safety HATKO rules Biochemical substrate Enzyme
Chemicals used in cosmetics must interact with the enzymes for their consumption after entering our bodies. The area at which the interaction realizes on the enzyme is known as the active center. This center is three dimensional and optically active. Considering the properties of the active regions, it is believed that the determination of the geometric properties of the chemicals may contribute to the safety evaluation of the chemical products. Obtainment of toxicological data of chemicals is a long and difficult process. It is an impossible process as the animal experiments have been prohibited. Since there are large number of chemical compounds available, it is not possible to conduct toxicological evaluation on all of them. Therefore, it is important to estimate whether chemicals are toxic through using molecular formulas. In this study, the similarities and differences between the unwanted chemicals used in cosmetic products and the geometric structures of the chemicals used in metabolism were determined by using Group Theory. The aim of the study is to estimate whether the chemicals will be toxic by taking advantage of point-group determinations. Molecular formulae of chemicals prohibited in cosmetics and substrates used in biological systems were used as materials. The point groups of the chemical drawing program was molecules were determined using these formulas. ChemDraw Professional 17 used to draw the formula structures of the molecules. Campus Licensed version of the program provided by application which was developed to examine the molecules acAtatürk University was utilized. An Excel cording to the Group Theory was also used.
1. Introduction Group theory explains the spectroscopic properties and molecular orbitals of the compounds by using the structures of molecules. Group Theory is used to explain the experimental results rather than revealing unknown properties. The catalytic centers of enzymes that catalyze chemical reactions in living systems are three-dimensional. Therefore, molecules with appropriate geometries may approach to active centers and each enzyme has effects on certain functional groups (David L. Nelson, 2013). The chemicals used in cosmetics interact with enzymes after entering our body, in order to be used or destroyed. Reactions catalyzed by enzymes occur in these active centers. The active center is a special region of the enzyme with substrate binding feature. Knowing the geometric characteristics of active centers and chemicals can contribute to the safety assessment of cosmetics. Cosmetic product includes any substance or mixture intended to be placed in contact with the external parts of the human body such as
∗
epidermis, hair system, nails, lips, and external genital organs or with the teeth and the mucous membranes of the oral cavity with a view exclusively or mainly cleaning them, perfuming them changing their appearance, protecting them, keeping them in good condition or correcting body odors (Cosmetics Directive, 2009). Cosmetic product components are any synthetic or natural substance or mixture used in the structure of cosmetic product other than perfume and aromatic substances (Sağlık Bakanlığı, 2005a). The safety assessment is an assessment report that must be made on the finished product, considering the toxicological properties, chemical structure and exposure levels of the components of a cosmetic product, the specific exposure characteristics of the target audience or the area for which the product is to be applied. (Sağlık Bakanlığı, 2005b). Toxicology is a branch of science that studies the negative effects of chemicals on living organisms. All foreign substances i.e. xenobiotics, which are not required for the normal metabolism of the organism rather taken from outside through various ways, are included in the area of interest of toxicology. Toxicity is toxic effect of xenobiotics (Vural,
Corresponding author. E-mail address: [email protected] (Y. Demir).
https://doi.org/10.1016/j.yrtph.2019.03.001 Received 2 October 2018; Received in revised form 14 February 2019; Accepted 1 March 2019 Available online 08 March 2019 0273-2300/ © 2019 Elsevier Inc. All rights reserved.
Regulatory Toxicology and Pharmacology 104 (2019) 56–58
Y. Demir, et al.
2.1. Selection of molecules
Table 1 Quantity and availability percentage of point groups in the prohibited and nonprohibited chemicals, as analyzed according to Group Theory. Group
Prohibited
C1 Cs C∞V D∞V Ci
In the selection of chemicals that are forbidden to use, the list of substances that should not be included in Cosmetic Products is used. Of the 1233 chemicals in this list, 621 chemicals which are not polymeric and inorganic were selected. In order to make a comparison (non-harmful molecular model), 81 molecules used in living systems were selected. These molecules are amino acids, carbohydrates (straight chain structures, α- and βHaworth structures), fatty acids, citric acid cycle molecules, glycolysis pathway molecules and compounds used in other metabolic ways.
Non-prohibited
Count
%
Count
%
252 196 139 30 4
40.51 31.51 22.35 4.82 0.64
18
21.95
61 2
74.39 2.44
2.2. Drawing of molecular structures
Table 2 Appearance Frequencies of the determined point groups according to Group theory (GSx). Point Group
Appearance Prevalence (GSx)
C1 Cs C∞v D∞v Ci
0.542 0.000 3.328 0.506 0.000
Molecules selected for analyses were drawn with ChemDraw Professional 17 program and recorded with JPEG extension. JPEG image files were used in the studies conducted according to Group Theory. 2.3. Rules concerning the examination of molecules While the molecules were analyzed according to Group Theory, a set of rules known as HATKO rules was used to provide standardization (Uçkaya et al., 2016).
2005). It determines the amount of toxicity of a chemical entering the living system, that is to say, the main factor determining the toxicity is the dosage (Borzelleca, 2000). The term LD50 is used as an acute toxicity unit to indicate how toxic a substance is. NO(A)EL, LO(A)EL, SED, LC50 and MoS are other toxicity units. The pharmacological effects, toxicities and pharmacokinetic properties of the chemical substances that are synthesized or isolated from natural sources should be examined in appropriate experimental animals. The basic concept as the basis for these tests and experiments is that the drug would cause the same effects it caused in the experimental animals (Büken Ö., 2000). According to the eighth article of the Universal Declaration of Animal Rights, which was promulgated in Paris on 15 October 1978, conducting experiments causing physical or psychological suffering on animals are violation(s) of animal rights. This shall apply for any and all medical, scientific, commercial and etc. experiments. After 11 March 2013, the sale of any cosmetic and personal care products tested on animals was prohibited in the European Union. This also applies to products imported from the non-member countries (Universal Declaration of Animal Rights, 2016). In our opinion, since these chemicals are forbidden to be tested on animals, these experiments are carried out on humans, especially on women being the main consumers. Therefore, the development of toxicological estimation techniques for human and community health will be very beneficial for public health in the longer term. Cosmetic products became widespread in 1900s. The conscious use cycle of these products started after the Second World War. The green movement, started in Western Europe in the late 1960s, has led to the tendency of natural origin cosmetics in the cosmetic industry. Due to this trend, natural raw materials and finished products were developed. Then, it was seen that natural raw materials or finished products could be harmful to human health. Therefore, it was concluded that safety and efficacy assessment must be performed for all cosmetic products of natural or synthetic origin (Bergişadi et al., 2014; Alğın Yapar and İnal, 2012; Kaymak and Tırnaksız, 2007).
2.4. Programs used in the analysis of molecular structures 2.4.1. Molecular analysis according to group theory used in excel applications The Excel application used in the analysis was developed by Yaşar Demir and used to analyze whether there are inversion point, axes, planes and reflection-rotation points available in the molecules. 2.5. Determination of point groups of molecules In defining the point groups, point group determination table included in Inorganic Chemistry books is used. (Shriver et al., 1994). 3. Results and discussion As a result of the investigations and researches, the point groups of the molecules according to Group Theory were determined separately and the results obtained were discussed. 3.1. Point group analysis A total of 702 molecules were analyzed, 621 of which were prohibited molecules and 81 of them were not prohibited ones. At the end of these studies, the following data have been obtained. In a review of this table to see the difference between the groups, it is seen that the prohibited molecules are clustered in five different point groups and the non-prohibited molecules are clustered in three different point groups. Cs and Ci groups are seen as groups unique to the prohibited molecules. C1, C∞V and D∞V point groups are seen in both the prohibited and non-prohibited groups. According to intra-group analysis, the most covalent group of molecules in the prohibited group is the C1 group while it is C∞V group in the non-prohibited group. Cs point groups have never been found in non-prohibited molecules. This result indicates that there are no intramolecular symmetries of the substrates. Substrate molecules do not have mirror planes. The absence of the point group Ci indicates that the non-prohibited molecules do not have symmetry points in the molecule. The fact that the C1 point group was present in both clusters indicates that all the molecules examined have at least one axis of rotation.
2. Materials and methods In this study, 621 molecular formulae which are prohibited to be used in cosmetic products and 81 formulae of chemical substances used in living metabolism have been used. Structures of these formulas are drawn and point groups are determined according to Group theory. 57
Regulatory Toxicology and Pharmacology 104 (2019) 56–58
Y. Demir, et al.
order to determine the point groups.
The fact that the D∞v element is available in both groups shows that some of the prohibited and non-prohibited molecules have one main axis, one horizontal axis and one horizontal plane perpendicular to the main axis. Other point groups were never seen in both the prohibited and nonprohibited groups. When we compare the presence quantity of these point groups to each other (Prohibited/Non-Prohibited), we get the Prevalence of Appearance (GSx) of these properties. The appearance frequencies of the point groups are given in the table below. In the five groups observed, only the prevalence of C∞V is significantly higher in the non-prohibited group. The prevalence of other groups of points is higher in the prohibited cluster. The Cs and Ci group are seen only as a feature of the prohibited cluster. The Cs property is important because it defines a plane and the Ci property is important since it defines the intramolecular symmetry. The fact that there is a plane of the molecule and that its symmetries (i.e. the Ci property) are noticeably high mean that identifiable symmetrical geometric structures of the prohibited molecules may be present. In addition, non-presence of these characteristics in the non-prohibited cluster supports the idea that the active regions of the enzymes work as a result of the interaction of the functional groups around the active center (compatibility model as a result of interaction) rather than the suitability of the geometric structure (key lock model).
Transparency document Transparency document related to this article can be found online at https://doi.org/10.1016/j.yrtph.2019.03.001. References Alğın Yapar, E., İnal, Ö., 2012. Nanomaterial sand cosmetics. J. Fac. Pharm. Istanbul 42 (1), 43–70. Bergişadi, N., Erdem, O., Çadırcı, E., Alğın Yapar, E., Aktaş, Y., Erdal, M.S., Demirel Özel, A., 2014. Critical analyze of safety cosmetic components' safety evaluation carried pursuant to European standards. In: Kadıoglu, Y. (Ed.), Safety Evaluation of Cosmetics in Europe. Cosming.com. Publications, İstanbul, pp. 64–109. Borzelleca, J.F., 2000. Profiles in toxicology paracelsus. Her. Mod. Toxicol. 23, 2–4. Büken, N.Ö., 2000. Clinical Drug Researches in Term of Experimental Subject and Patient. A.U. Unpublished Doctoral Thesis of Institute of Medical Sciences, Ankara. Kaymak, Y., Tırnaksız, F., 2007. Unwanted effect of cosmetic products. Dermatose 6 (1), 39–48. Nelson, David L., Cox, Michael M., 2013. Lehninger Biyokimyanın İlkeleri, (5. Baskı). Palme Yayıncılık, Ankara, pp. 183–186. Sağlık Bakanlığı, T.C., 2005a. Kozmetik Yönetmeliği (Resmi Gazete Sayısı: 25823), Birinci Bölüm, File:. Sağlık Bakanlığı, T.C., 2005b. Kozmetik Yönetmeliği (Resmi Gazete Sayısı: 25823), Üçüncü Bölüm. Shriver, D.F., Atkins, P.W., Langford, C.H., 1994. Molecular shape and symmetry. In: Inorganic Chemistry, second ed. Oxford University Press, Oxford, pp. 105–145. The European Parliament and The Council, 2009. Regulation (EC) No 1223/2009 of 30 November 2009 on Cosmetic Products Section One Article 2. 30, 11. Uçkaya, M., Uçkaya, F., Demir, N., Demir, Y., 2016. Evaluation of the efficiency and safety in cosmetic products. Int. J. Pharm. 499, 295–300. Universal Declaration of Animal Rights, 2016 Article 8. Vural, N., 2005. Toxicology. Ankara University, Publications of Pharmacy, Ankara Faculty No:73.
4. Conclusion In accordance with the results of the study we conducted, it is seen that it is possible to determine the toxicological characters of the chemicals. In this determination, it made sense to compare the harmful chemicals with the chemicals (substrates) used in the living systems in
58