UDC 615.21: 615.076.9 APPROVED

General Director CJSC “Saint Petersburg Institute of Pharmacy” Doctor of Medical Sciences, Professor V.G. Makarov [Signature] 13.09.2021 

It is carried out in accordance with the GLP OECD Principles

STUDY REPORT

Study of pharmacological safety of substance Lithium Ascorbate (Normopharm LLC, Russia) in rats following a single intragastric administration 

(final)

Study Leader V.A. Vavilova 13.09.2021 [Signature]

Leningrad Region, 2021

ABSTRACT

CONCLUSION Study code 2.21/21, Test system: Wistar rats, 10 males in each group, route of administration – intragastric

SUMMARY

Throughout the experiment, the animals were in satisfactory condition, the behavioral reactions of the rats that had received the substance did not differ from those of the control group.

The effect of the test object in doses of 10 mg/kg (HTD), 50 mg/kg (5 HTD) and 150 mg/kg (15 HTD) on the dynamics of body weight and body temperature was not shown.

Based on the results of evaluation of the pharmacological safety of substance Lithium Ascorbate (Normopharm LLC, Russia), it was established that the test object had no effect on the respiratory, cardiovascular and central nervous systems.

Keywords: lithium ascorbate, single administration, pharmacological safety, rats, intragastric administration

The report is presented on 176 pages, including 8 tables and 17 sources

STUDY DATES

LIST OF EXECUTORS

STUDY DECLARATION OF GLP COMPLIANCE 
FROM THE STUDY LEADER

This study was carried out in accordance with the standard operating procedures of the institution, and the Study Plan mutually agreed with the Sponsor (Normopharm LLC, Russia).

The study complies with the GLP requirements, except for the fact that no identity, purity and stability of the test object were not determined. The values were determined by the Sponsor by standard methods without regards to GLP requirements.

There were no deviations from the Study Plan that would affect data interpretability or the scientific integrity and results of the study.

I, the undersigned, hereby confirm that I take overall responsibility for the technical conduct of the study; analysis, interpretation, documentation and presentation of results, as well as archiving of the study-related materials.

The objectives set out in the study plan have been achieved. There were no unforeseen circumstances that could affect the quality or integrity of the study.

This report presents the results reliably. I am fully responsible for the accuracy of the data obtained, as well as the confidentiality of the preclinical study.

I guarantee that after the study completion, the study plan, the final report, source data and all relevant documentation will be transferred to the archives of the research institution.

STATEMENT BY THE MANAGEMENT OF THE RESEARCH INSTITUTION ON THE PROVISION OF RESOURCES TO CONDUCT THE STUDY IN ACCORDANCE WITH THE GLP PRINCIPLES, REGULATORY REQUIREMENTS AND STANDARDS FOR THE ETHICAL HANDLING OF ANIMALS

The management of the research institution shall ensure that the following has been provided for the proper conduct of the study:

• availability of a sufficient number of qualified and experienced personnel with a clear understanding of their responsibilities, as confirmed by training data;

• equipping the research institution with the necessary equipment, facilities and materials;

• availability of a quality service responsible for the quality assurance system;

• availability of approved standard operating procedures, as well as access to them by all personnel involved in the conduct of the study;

• appointment of a study leader in accordance with the established procedure, who has qualifications appropriate to the study objectives;

• interaction of the study leader, employees of quality service and personnel involved in the study.

Compliance with the Principles of Good Laboratory Practice

This study was carried out in accordance with the principles of GLP OECD (GOST 33044-2014 “Principles of Good Laboratory Practice”; Decree of the EEC Council № 81 “On Approval of the Rules of Good Laboratory Practice of the Eurasian Economic Union in Circulation of Medicines” dated 03.11.2016). The manipulations were performed in accordance with the standard operating procedures of the institution and the Study plan.

Regulatory Compliance

The design of this study was based on the selection of the study goal, in accordance with the regulatory legal acts and guidelines laid down in “Regulatory Standards” section of this Report.

Compliance with the Standards of Ethical Handling of Animals

This study was reviewed at a meeting of the Bioethics Commission for compliance of the draft study with the “Three R’s” principles and Directive 2010/63/EU. The study was approved for conduct (№ BEC 1.28/21 dated 12.05, 2021, 11 persons voted).

General Director of CJSC “Saint Petersburg Institute of Pharmacy”

V.G. Makarova         [Signature]   13.09.2021

       /Name/     /Signature/     /Date

STATEMENT OF THE QUALITY SERVICE ON CONDUCTING AND DOCUMENTING THE INSPECTION OF THE KEY STAGES OF THE STUDY

The Quality Service conducted and documented all stages of the study inspection. The results were reported to the study leader and the management of the research institution.

The study was inspected to ensure that the procedures and manipulations performed were in accordance with the standard operating procedures of the institution, the study plan, and the regulatory requirements of the Good Laboratory Practice standards.

The final study report was reviewed by a quality officer and found to be an accurate statement of the data obtained and the procedures applied. The results presented in the final report accurately and fully reflect the data obtained during the study.

The conclusion of the quality service on this study is an Annex to the final report.

S.S. Sapynov                 [Signature]             13.09.2021

/Name/             /Signature/ /Date/

TABLE OF CONTENTS

LIST OF ABBREVIATIONS AND ACRONYMS 12

INTRODUCTION 14

STUDY GOAL AND OBJECTIVES 15

1. Materials and methods 16

1. Study Objects  16

2. Animals 16

3. Method of administration and selection of doses 19

1. Method and duration of administration 19

2. Selection and calculation of doses 20

3. Dosing procedure 20

4. Methodology 20

1. Study design 20

2. Feed deprivation 21

3. Body weight recording 21

4. Recording of the timing of development of intoxication and clinical examination of animals…………..…….. 22

5. Body temperature recording 23

6. Study of pharmacological safety in relation to the respiratory system ……………….. 23

7. Study of pharmacological safety in relation to the cardiovascular system 23

8. Study of pharmacological safety in relation to the central nervous system (CNS) 24

9. Euthanasia 26

10. Data analysis 26

11. Study quality assurance and control……………………………………………..…………26

2. Study results 27

1. Clinical picture of intoxication and clinical examination of animals 27

2. Effect of the test object on the body weight of animals 28

3. Effect of the test object on the body temperature of animals 28

4. Evaluation of the pharmacological safety of study drugs in relation to the respiratory system ……………………………………………………………………………………………………………...28

5. Evaluation of pharmacological safety of study drugs in relation to the cardiovascular system.. 29

6. Evaluation of the pharmacological safety of study drugs in relation to the central nervous system 29

FINDINGS 31

CONCLUSION 32

TABLES 33

DATA ARCHIVING 36

REGULATORY DOCUMENTS 37

REFERENCES 38

ANNEX A 40

ANNEX B 44

ANNEX C 68

ANNEX D 71

ANNEX E 75

ANNEX F 79

ANNEX G 80

ANNEX H 173

In this R&D report, the following abbreviations and acronyms are used:

INTRODUCTION

The test object is Lithium Ascorbate, substance (Normopharm LLC, Russia).

Organic lithium salts are most often used as neuroprotective agents for various affective disorders and vascular cognitive disorders [1, 2]. There are data on the efficacy of lithium not only in endogenous mental disorders, but also in patients with organic psychoses, epilepsy and various psychopathies. In experiments, the neuroprotective effect of lithium salt preparations was established in various models of cerebral ischemia [3]. However, the mechanism of action of lithium salts is not well understood. At the same time, tests on rats have shown a low cumulative effect and low toxicity of lithium ascorbate [4]. In this regard, the study of lithium preparations, in particular the complex of lithium and ascorbic acid, remains relevant. It has been established that this salt has anti-stress and antidepressant activity [5].

This study is aimed at evaluating the pharmacological safety of the test object substance lithium ascorbate following a single intragastric administration to mature rats, and is regulated by normative documents [6-8].

The present study was carried out in accordance with the principles of good laboratory practice, approved by the study plan [Appendices A and B], reviewed and approved by the bioethics commission [ANNEX C].

The information obtained during the study did not duplicate the results of previous studies.

STUDY GOAL AND OBJECTIVES 

Study goal:

Evaluation of pharmacological safety of lithium ascorbate substance (Normopharm LLC, Russia) following a single intragastric administration to rats.

Study objectives:

1. Evaluation of the effect of a single administration of different doses of the test object on the functional condition of the respiratory system;

2. Evaluation of the effect of a single administration of various doses of the test object on the functional condition of the cardiovascular system;

3. Evaluation of the effect of a single administration of different doses of the test object on the functional condition of the central nervous system.
   

1. Materials and methods

1. Study objects

Table 1.1.1.1 and 1.1.2 provide information about the study objects.

Table 1.1.1 – Test object

Table 1.1.2 – Control substance

The forwarding documents of the test object are given in Annex D.

The research institution has not carried out any identification, purity and stability studies of the test object. These values were determined by the Sponsor according to standard methods. The Sponsor of the study is responsible for the reliability of the submitted data on the identification, purity and stability of the test object.

1.2 Animals

Age of animals at group formation   Up 12 weeks:

Group allocation:  To exclude the influence of the investigator’s preferences on the

formation of experimental groups, animals were selected with the method of modified block randomization [8]. To do this, all animals submitted to the study were randomly placed in the cages of the randomization block (the number of cages of the randomization unit is a multiple of the number of groups in the experiment). Then, using a random number generator, a list of data was obtained, containing the numbers of the cages with animals and the corresponding numbers of the groups where the animals were subsequently placed [9] [Annex F].

The animals were kept under standard conditions in accordance with Directive 2010/63/EU of the European Parliament and of the Council of 22 September 2010 on the protection of animals used for scientific purposes [11].

3. Method of administration and selection of doses

1. Method and duration of administration

The test object and control substance were administered to animals intragastrically since this method is an analogue of oral, which is to be used in clinical practice. The objects were injected administered once with further observation for 14 days.

1. Selection and calculation of doses

According to Patent № RU2617512C1 in the study of pharmacological efficacy of lithium ascorbate in rats, animals received lithium ascorbate in the dose range of 30-120 mg/kg, intragastrically. The maximum efficacy was observed when administered at the lowest dose studied [12].

On the Sponsor’s recommendation, doses of 10 mg/kg (HTD), 50 mg/kg (5 HTD) and 150 mg/kg (15 HTD) were studied to evaluate the pharmacological safety of lithium ascorbate when administered intragastrically.

The control substance was administered once intragastrically to animals of group № 1, in an amount equivalent to the volume of the maximum dose of the test object.

2. Dosing procedure

The test object (as a suspension) and the control substance were administered to the animals once intragastrically using a tube.

The test object was administered in various concentrations. Dosing volumes which were calculated on the basis of animal body weight data and did not exceed the recommended ones [13], are presented in Table 1.3.3.1.

Table 1.3.3.1 – Dosing volume of the test object

The control substance was administered to the animals once intragastrically in a volume corresponding to the volume of the maximum dose of the test object (1 ml/kg).

4. Methodology

1. Study design

The total number of animals involved in the experiment was 40 Wistar rats (10 males in each group).

The characteristics of the experimental groups and the scheme of the experiment are presented in Tables 1.4.1.1 and 1.4.1.2.

Table 1.4.1.1 - Characteristics of experimental groups

Table 1.4.1.2 - Manipulation scheme

1.4.2   Feed deprivation

The rats were deprived of food for 16 hours before the study subjects were administered. Water was given ad libidum throughout the experiment.

3. Body weight recording

The body weight of the animals was recorded the day before the administration of the objects, then weekly [ANNEX G].

The rats were weighed on a Vibra AJ-1200CE balance (Shinko Denshi, Japan). The maximum weighing limit is 1200 g, the minimum weighing limit is 0.5 g. Calibration mark is 0.1 g. Accuracy class is 2 [ANNEX B].

4. Recording of the timing of development of intoxication and clinical examination of animals

The animals were continuously monitored for 30 minutes from the administration of the test subjects and then daily.

The following was recorded:

• Behavior: distress/agitation;

• Response to stimuli: decreased/increased;

• Skin: redness / paleness / cyanosis / jaundice;

• Mucous membranes: redness/pallor/cyanosis/jaundice;

• Discharge: from the eyes / from the nose / from the anus / from the urethra;

• Muscle tone: decreased/increased;

• Motor coordination disorders: ataxia/hyperkinesis;

• Dyspnoea;

• Death.

The animals were clinically examined before administration of the objects, 30 minutes after administration, on day 2 day, then weekly. A detailed examination of the animal was carried out in the cage, in the hands and in the open area. The manifestation and severity, where acceptable, of signs of intoxication were observed.

Examination in the cage:

• Behavior: normal/distress/agitation/aggression;

• Attitude towards other animals: normal/aggression.

Outdoor examination:

• Position of the body in space: normal/forced position of the body/impaired coordination of movement (ataxia/hyperkinesis);

• Breathing Type: normal/change in breathing pattern/change in breathing rhythm/dyspnea;

• Bowel movements: normal/diarrhea/presence of blood in the stool/change in stool color;

• Urination: normal/discoloration.

Examination of the animal when taking it in hand:

• Response to stimuli: normal/decrease/increase;

• Hair condition: normal/ruffled/shedding hair/dull/dirty/discolored;

• Body condition: normal/dystrophic/obese;

• Muscle tone: normal/decrease/increase;

• Skin: normal/discoloration/hemorrhage/inconsistency/palpable masses/turgor;

• Eyes and conjunctiva: normal/abnormal/ptosis/exophthalmos/discharge/conjunctival discharge/discoloration;

• Nasal cavity: normal/discharge/discoloration of the mucosa/violation of integrity;

• Oral cavity: normal/drooling/mucosal discoloration;

5. Body temperature recording

Body temperature was recorded rectally in all animals using the IM-10 patient monitor (CJSC East Medical, Moscow) before administration, 30 minutes after administration, on day 2 and then weekly.

6. Study of pharmacological safety in relation to the respiratory system

The effect of the test object on the functional condition of the respiratory system was evaluated on anesthetized animals by recording the respiratory rate (RR) using a pneumograph with a piezo sensor (AD Instruments, Australia) 24 hours after the administration of the test object or control substance immediately before ECG recording.

7. Study of pharmacological safety in relation to the cardiovascular system

Pharmacological safety in relation to CVS was evaluated by recording an electrocardiogram (ECG) and systolic blood pressure (SBP) on day 2 of the experiment.

To record the ECG, any animal was preliminarily anesthetized with a mixture of Zoletil 100 (Virbac Sante Animale, France) and Xila (Interchemie werken De Adelaar B.V., the Netherlands) during the intravenous administration, after which it was fixed on the operating table. ECG was recorded out using a computer electrocardiograph for veterinary medicine “Poly-spectrum-8E” (Neurosoft LLC, Russia). Steel terminals were used as electrodes. The following parameters were evaluated: heart rate (HR), RR interval (ms), P (ms), PQ (ms), QRS (ms), QT (ms).

Systolic blood pressure (SBP) was recorded and evaluated in anesthetized animals immediately after ECG recording. In animals, SBP was measured non-invasively using a device for measuring blood pressure in laboratory animals (Blood pressure measurement system Systola, Neurobotics LLC, Russia).

8. Study of pharmacological safety in relation to the central nervous system (CNS)

Pharmacological safety for the central nervous system was evaluated using a battery of functional tests proposed by Irwin S. [14] and recommended for the evaluation of pharmacological safety by the S7A ICH guideline and GOST R 56700-2015 [15, 16] and “open field” test to evaluate individual behavior. It was evaluated 24 hours after administration of the test object or control substance in all animals.

The following components were used for testing: an open area - an “arena” with a wire mesh, bounded on four sides; transparent container; the path is a “corridor”.

The procedure included an evaluation of the initial behavioral characteristics and a number of subsequent manipulations, during which the animal was sequentially exposed to provoking stimuli. The evaluation was carried out in stages, except for items ”pelvic elevation”, “ataxia”, “diarrhea” (the presence or absence of this sign was noted).

The evaluation of the animal’s condition began with the observation of “natural” behavior in a transparent container (the following parameters were noted: body position, locomotor activity, abnormal behavior, breathing, the presence of tremors, convulsions). At the end of the examination inside the container, the animal was quickly lowered to the level of the “arena” to evaluate the response to movement and values such as ptosis, piloerection and startle reaction.

Next, the animal was fixed by the tail and transferred to the “corridor” installation. The investigator recorded the distance (in centimeters) traveled by the animal (movement along the “corridor”). Also at this stage, the elevation of the pelvis and tail during movement, the severity of adynamia, the reaction to the approach of the rod and the reaction of avoidance to the rod were tested.

At the end of testing in the corridor installation, the animal was transferred back to an open area to evaluate its response to touch, posture passivity, and the presence of ataxia.

To evaluate visual orientation, the animal was fixed by the middle of the tail and, after contact with the net, it was released to allow it to stand in a normal position. During this test, the grasping reflex was also evaluated. To evaluate the values of body muscle tone, corneal reflex, pineal reflex, response to squeezing of the fingers of the pelvic limb with tweezers, and behavior on a stretched cord, the animal was held by the base of the tail.

When fixing the animal in a backward condition, such values as skin color, the presence of diarrhea, stiffness of the limbs, muscle tone of the anterior abdominal wall, the condition of the pupils, the presence of salivation, lacrimation, and bite reaction were evaluated.

At the next stage, the investigator released the animal into an open area, where he evaluated its reaction to the pinching of the tail and the attitude reflex.

During the entire sequence of manipulations, the possible desire of the animal to harm the investigator was also evaluated, and forced freezing was noted. After the completion of these manipulations, the animal was returned to the cage.

The battery of tests made it possible to evaluate the complex effect of the test drug on the central nervous system of experimental animals, including behavior, neurological disorders and the functional status of the autonomic nervous system.

The orientation and exploratory activity of the animals was studied 24 hours after the administration of the test object or control substance in all animals. Based on the behavioral atlas for rodents [17], a number of elementary motor acts and postures were selected, the totality of which characterized holistic behavior when tested in the “Open Field” installation [ANNEX G].

In the “Open Field” test, the following values were evaluated:

• the number of crossed squares and center visits (horizontal activity);

• number of free and wall racks (vertical activity);

• number of urination and defecation boluses;

• autogrooming.

9. Euthanasia

In accordance with Directive 2010/63/EU of the European Parliament and of the Council of the European Union on the protection of animals used for scientific purposes of 22 September 2010, the animals were euthanized with the use of carbon dioxide (CO2), followed by the cutting of the main blood vessels [11]. This type of animal euthanasia was accompanied by a minimum of pain, suffering and distress and was carried out by competent personnel.

10. Data analysis

Descriptive statistics was applied to all data: the data were checked for compliance with the normal distribution law using the Shapiro-Wilk’s W test. In the case of a normal distribution, the mean value and the standard error of the mean were calculated, which, together with the value of n (number of cases), are presented in the summary tables. In cases where the data did not conform to the normal distribution law, the median and quartile range were calculated. For evaluation of data with signs of normal distribution univariate analysis of variance (ANOVA) was used, if a significant effect of the studied factor was found, subsequent intergroup comparisons (post hoc analysis) were carried out using Tukey’s test analysis. To compare the two groups, the Student’s test was used. For the analysis of data that did not conform the normal distribution law, the Kruskal-Wallis test was used, followed by the use of the nonparametric method of average ranks for multiple comparisons in case of a significant effect of the studied factor. To compare the two groups, the Mann-Whitney U-test was used. Differences were determined at the significance level of p<0.05.

Statistical analysis was performed using licensed software Statistica 10 (StatSoft, USA).

11. Study quality assurance and control

The quality service of the research institution was carried out [ANNEX I]: 

- verification of the study design;

• checking the study schedule;

• incoming audit of the preclinical study;

• audit of the experimental part of the study;

• checking the chronology of the study and the completeness of the study report;

• verification of the final study report.

2 Study results

The pharmacological safety of substance Lithium Ascorbate (Normopharm LLC, Russia) was studied in rats following a single intragastric administration. The data obtained during the experiment are presented in full in the source charts [ANNEX G].

1. Clinical picture of intoxication and clinical examination of animals

Throughout the experiment, no deaths of animals were recorded.

During the experiment, the animals were observed daily.

Prior to the administration of the objects, 30 minutes after the administration, on the 2nd day and thereafter, a clinical examination of the animals in the cage, in the hands and in the open area was carried out weekly, their general condition and behavior were evaluated.

The results of clinical observations and clinical examinations did not show any deviations in the appearance and behavior of the experimental animals.

The response to stimuli and to pick-ups was standard, moderately pronounced in all animals.

Visually, the body condition of all animals was satisfactory. The hair is smooth, shiny, without patches of baldness.

Muscle tone in all animals was moderate. There were no disorders of coordination of movements and gait.

Turgor and integrity of the skin were preserved, palpable masses were not noted.

The visible mucous membranes are pale pink in color, shiny, without impaired integrity.

In all animals, exophthalmos, swelling or hyperemia of the mucous membranes of the eyes, lacrimation were not observed.

The nose was moderately moist, there was no pathological discharge. Ears of normal temperature, suppuration, inflammation, impurities were not noted in any animal. All of them have their teeth preserved. No salivation disorders were observed.

Respiration was normal in all experimental animals.

There was no pathological discharge during urination and defecation. No unusual behavior was reported.

The results of clinical observations and examinations are presented in full in the source charts.

Thus, throughout the experiment, the animals were in a satisfactory condition, the behavioral reactions of the rats that received the substance did not differ from those of the control group.

2. Effect of the test object on the body weight of animals

Table 2.2.1 presents the body weight of males during the experiment.

The data corresponded to the normal distribution law. The body weight of the animals before the beginning of the experiment did not differ between the groups (univariate ANOVA, p>0.05) and did not deviate from the mean for all groups before the beginning of the experiment by more than 20%.

Analysis of variance with repeated measurements found a statistically significant effect of the factor “repeatability of measurements” (p<0.05), while the “group” factor was insignificant (p>0.05). A subsequent intergroup comparison using the Tukey test showed that the animals of all groups showed physiological weight gain during the experiment (Table 2.2.1).

The effect of the test object on the body weights of males has not been established. There were no statistically significant differences from the control group.

3. Effect of the test object on the body temperature of animals

Table 2.3.1 presents data on the body temperature of males during the experiment.

The data corresponded to the normal distribution law. The body temperature of the animals before the experiment did not differ between the groups (univariate ANOVA, p>0.05).

Analysis of variance with repeated measurements did not show statistically significant effects of the factor “repeatability of measurements” and the “group” factor (p>0.05) (Table 2.3.1).

The effect of the test object on the body temperature of male rats has not been established. There were no statistically significant differences from the control group.

4. Evaluation of the pharmacological safety of the test object in relation to the respiratory system

Table 2.4.1 presents the results of measuring the respiratory rate (RR) of experimental animals on day 2 of the experiment.

The data were consistent with the normal distribution law (the Shapiro-Wilk test). Univariate analysis of data variance (ANOVA) did not show the effect of the group factor on respiratory system in male rats on day 2 of the experiment (p>0.05, Table 2.4.1).

The effect of the test object on the respiratory system was not found.

5. Evaluation of the pharmacological safety of the test object in relation to the cardiovascular system

Table 2.5.1 presents the data of ECG parameters of experimental animals on day 2 of the experiment.

The data were consistent with the normal distribution law (the Shapiro-Wilk test). Univariate analysis of data variance (ANOVA) did not show statistically significant differences between the experimental groups in the studied ECG values in male rats on the 2nd day of the experiment (p>0.05).

Table 2.5.2 presents the results of systolic blood pressure (SBP) measurements in experimental animals on day 2 of the experiment.

The data were consistent with the normal distribution law (the Shapiro-Wilk test). Univariate analysis of data variance (ANOVA) did not show the effect of the group factor on SBP in male rats on day 2 (p>0.05).

Despite the absence of statistically significant differences, it should be noted that a slight increase in heart rate was present in the maximum dose group, but the increase was no more than 7% of the control values and did not differ statistically significantly from the control values. As well, in the group that received the test object at the maximum dose (150 mg/kg), there was a slight increase in systolic blood pressure, the increase did not exceed 7% relative to the control and was not statistically significant.

The absence of statistically significant changes in the studied parameters suggested that lithium ascorbate did not have a significant effect on the functional activity of the cardiovascular system.

6. Evaluation of the pharmacological safety of the test object in relation to the central nervous system

Irwin’s Test

The results of the evaluation of the pharmacological safety of the test object for the central nervous system are given in Table 2.6.1. The table shows only deviations from the normal for the number of animals in which these deviations were found. These data are presented as numerical values for the evaluation of a particular parameter. The data of animals that did not differ from the normal in one or another parameter were not included in the table.

As a result of the evaluation of the pharmacological safety of the test object on the central nervous system of experimental animals on day 2 of the administration, insignificant differences from the norm in the parameter “Movement along the corridor” were found both in the groups that received the test object and in the group that received the control substance. There was no dependence on the received dose of the test object, changes even within the same group were multidirectional. All the identified changes can be considered as individual characteristics of the animals.

Open Field Test

The orientation and exploratory activity of male rats was studied on day 2 of the experiment in the “open field” test to record the effect of the studied objects on individual behavior. The results are shown in Tables 2.6.2 and 2.6.3.

The data corresponded to the normal distribution law (number of wall racks and number of squares visited, Shapiro-Wilk test). Univariate analysis of variance did not show the effect of the “group” factor on locomotor activity after administration of the studied objects (Table 2.6.2).

The data did not correspond to the normal distribution law (number of central visits, number of free racks, number of autogrooming, number of urinations, number of bowel movements). There were no statistically significant differences in the values of the parameters of male activity in the “Open Field” setting on the next day after the administration of the studied objects (Kruskel-Wallis test, p>0.05), (Table 2.6.3).

Thus, according to the results of the evaluation of the pharmacological safety of lithium ascorbate in relation to the central nervous system, no clinically significant effect was found.

FINDINGS

The study of the pharmacological safety of substance Lithium Ascorbate (Normopharm LLC, Russia) following a single intragastric administration to rats concluded the following:

1. No animal deaths were observed. Throughout the experiment, the animals felt satisfactorily. The effect of the test object on the dynamics of body weight and on body temperature was not found.

2. In the study of pharmacological safety in relation to the respiratory system, the effect of the test object under the conditions of the experiment was not found.

3. In the study of pharmacological safety in relation to the cardiovascular system, the effect of the test object under the conditions of the experiment was not found.

4. In the study of pharmacological safety in relation to the central nervous system under the conditions of the experiment, the effect of the test object was not found.
   

CONCLUSION

According to the results of the evaluation of pharmacological safety of substance Lithium Ascorbate (Normopharm LLC, Russia), it was established that the test object had no effect on the respiratory, cardiovascular and central nervous systems following a single administration in doses of 10 mg/kg (1HTD), 50 mg/kg (5HTD) and 150 mg/kg (15HTD).