|Year : 2022 | Volume
| Issue : 4 | Page : 221-228
Safety profile of Ayurveda Rasoushadhi: An appraisal of technical reports on quality and safety of selected Rasakalpa—Metal and mineral-based Ayurvedic formulations
Bidhan Mahajon, Sarada Ota, Shruti Khanduri, Bhagwan Sahai Sharma, Sanjaya Kumar, Narayanam Srikanth
Central Council for Research in Ayurvedic Sciences (CCRAS), Janakpuri, New Delhi, India
|Date of Submission||27-May-2022|
|Date of Acceptance||11-Jul-2022|
|Date of Web Publication||21-Nov-2022|
Dr. Bidhan Mahajon
Research Officer (Ayurveda), Central Council for Research in Ayurvedic Sciences (CCRAS), Janakpuri, New Delhi 110058
Source of Support: None, Conflict of Interest: None
To a large extent, the safety of Rasoushadhi (metal and mineral-based formulations) is evident by its long history of clinical use. However, certain published literature generated misconceptions regarding their quality and safety across the globe. Thus, to protect the massive trust in Ayurveda, a multidisciplinary study was conducted by Central Council for Research in Ayurvedic Sciences for the assessment of the quality and safety of eight important Rasoushadhi, viz., Arogyavardhini vati (Ayurvedic Formulary of India [AFI]-I, 20:4), Mahayogaraja guggulu (AFI-I, 5:6), Vasantakusumakar rasa (AFI-I, 20:42), Mahalaxmivilas rasa (AFI-I, 20:27), Rasamanikya (AFI-I, 20:33), Makaradhwaja (AFI-I, 15:2), Kajjaliyoga (AFI-III, 15:15), and Rasasindur (AFI-I, 15:6) under Golden Triangle Partnership (GTP) scheme. The present article is an appraisal of the published Technical Reports (Volumes 1 and 2) of GTP that highlights the collective safety outcome of these selected Rasoushadhi. Study investigators prepared these Rasoushadhi in a Good Manufacturing Practice (GMP)-certified pharmacy, and they performed repeated-dose oral toxicity studies per Organization for Economic Co-operation and Development-408 guidelines in Wistar albino rats. All the Rasoushadhi were orally administered at different dose levels for 90 days. Periodically, observations were done by the investigators for clinical signs of toxicity, mortality, morbidity, body weight changes, and feed consumption. After 90 days, they performed hematology, biochemistry, electrolytes, relative organ weight, and histological examinations. The study concluded that there were no significant differences in the observed parameters between the control and Rasoushadhi-treated rats. In histological examinations, also they found no toxicologically significant abnormalities related to any Rasoushadhi treatment. Based on the result, the investigators concluded that all the formulations were safe up to the tested high dose levels.
Keywords: Ayurveda, formulation, herbo-mineral, quality, Rasoushadhi, safety, toxicity
|How to cite this article:|
Mahajon B, Ota S, Khanduri S, Sharma BS, Kumar S, Srikanth N. Safety profile of Ayurveda Rasoushadhi: An appraisal of technical reports on quality and safety of selected Rasakalpa—Metal and mineral-based Ayurvedic formulations. J Drug Res Ayurvedic Sci 2022;7:221-8
|How to cite this URL:|
Mahajon B, Ota S, Khanduri S, Sharma BS, Kumar S, Srikanth N. Safety profile of Ayurveda Rasoushadhi: An appraisal of technical reports on quality and safety of selected Rasakalpa—Metal and mineral-based Ayurvedic formulations. J Drug Res Ayurvedic Sci [serial online] 2022 [cited 2022 Nov 30];7:221-8. Available from: http://www.jdrasccras.com/text.asp?2022/7/4/221/361579
| Introduction|| |
Rasoushadhi of Ayurveda are the metals or minerals containing medicines that have favorable effects on different biological systems. These metals or minerals are transformed into nontoxic forms through a meticulous process explained in the ancient texts.,,, Ayurveda has specified different methods of preparation and standard operative procedures, from collecting raw ingredients, their purification, and processing of intermediates alongside the method of uses/indication., In conjunction with incredible expansion in the worldwide use of Ayurveda, safety and efficacy and quality control have become essential concerns for the scientific presentation of facts. Adding to this, the information published through scientific and general print media also generated misconceptions regarding the quality and safety of metal and mineral-based formulations of Ayurveda across the globe. Thus, to protect the massive trust of the patient in Ayurveda, a prime initiative was taken by Central Council for Research in Ayurvedic Sciences (CCRAS) to assess the quality and safety of eight important metal and mineral-based formulations of Ayurveda. As these are the most popular and frequently used formulations, the documentation on chemical characterization and safety/toxicity studies is crucial to proving their safety profile., Therefore, pharmaceutical-analytical and toxicological studies were conducted to assess the quality and safety of eight important Rasoushadhi, viz., Arogyavardhini vati (AVV), Mahayogaraja guggulu (MYG), Vasantakusumakar rasa (VKR), Mahalaxmivilas rasa (MVR), Rasamanikya (RM), Makaradhwaja (MD), Kajjaliyoga (KY), and Rasasindur (RS) under Golden Triangle Partnership (GTP) scheme coordinated by CCRAS-AYUSH in collaboration with reputed organizations of Council of Scientific & Industrial Research (CSIR), viz., Indian Institute of Toxicology Research (CSIR-IITR), Lucknow; Indian Institute of Integrative Medicine (CSIR-IIIM), Jammu; CSIR-Central Drug Research Institute, Lucknow; CSIR-National Chemical Laboratory, Pune; and CSIR-Indian Institute of Chemical Technology, Hyderabad. Details of the quality and safety profile of these eight selected Rasoushadhi are already published as technical reports and available in the public domain.,,,,, The present article highlights the influential safety profile of these eight Rasoushadhi collectively.
| Materials and Methods|| |
Preparation of Rasa-aushadhi
The materials required for the preparation of the previously mentioned eight formulations were found to be procured from M/s. Maharishi Ayurveda Pharmacy, Noida, which is a GMP-certified pharmacy, thus ensuring quality raw materials. However, the investigators still subjected the raw materials for authenticity check through laboratory analysis and expert examination. It is noted that the eight drugs were prepared by using the pharmaceutical process mentioned in the Ayurvedic Formulary of India (AFI).,,,,,,, The compositions of the formulations are provided in [Table 1] and [Table 2].
In vivo toxicity studies
The investigators performed the repeated dose 90 days oral toxicity tests in male and female rats as per the standard guidelines of the Organization for Economic Co-operation and Development (OECD)-408.
The investigators obtained the healthy Wistar strain albino rats (Rattus norvegicus) from the Animal House of respective institutes of the CSIR. The weight and age of rats were between 120 and 160 g and 8–9 weeks, respectively. All the selected female rats were nulliparous and nonpregnant. They housed the rats in polycarbonate cages in an air-conditioned room maintained at a temperature of 22°C ± 3°C and a relative humidity of 30%–70%, with a 12:12 h light/dark illumination cycle. They submitted that CPCSEA (Committee for Control and Supervision of Experiments on Animals) guidelines for laboratory animal care were followed throughout the experiment. The protocol used in the study was approved by the Institutional Animals Ethics Committee vide protocol no. ITRC/IAEC/02/2007, IIIM/IAEC/12/2008 and 142/08/Toxicology/IAEC dated 05.11.08. They also acclimatized the animals for 7 days to the experimental animal room conditions and used the standard animal feed and water ad libitum for feeding the animals.
Preparation of dosage
The investigators administered all the drugs and vehicles orally via stainless steel gavages by calculating the dose based on the body weight of rats,,,, for 90 days. They fixed the duration of the toxicity study as 3 months (i.e., 90 days) based on its duration of the clinical use in the Ayurveda system. They explained that the weighed amount of dry powder of KY, RS, AVV, MYG, VKR, and MVR was suspended in 30%–33% (v/v) honey solution (30–33 mL honey plus 60–67 mL distilled water) and administered as per fixed dosage [Table 3], whereas the weighed amount of dry powder of RM and MD was suspended in 1% gum acacia, and a 1 mL/100 g rat dose was administered [Table 3].
|Table 3: Study design of repeated dose oral toxicity study of eight selected Rasoushadhi|
Click here to view
Details of the hematological study explained by the investigators state that they collected the blood in a 10% ethylenediamine tetraacetic acid (EDTA) solution. They analyzed red blood cells, white blood cells (WBC), hemoglobin, hematocrit (HCT), lymphocytes %, granulocytes %, and platelets using an automatic hematology analyzer (HUMA COUNT, Germany) and automatic hematology analyzer (MS9-3H, Kemia, Madrid, Spain) and also by using semi-automatic hematology analyzer (MS-9, Melet Schlosing).
For biochemical studies, blood samples were drawn by the investigators after 24 h of the last dose exposure. They analyzed blood samples for glucose, triglyceride, cholesterol, total protein, total bilirubin, alkaline phosphatase, aspartate aminotransferase (AST), alanine aminotransferase (ALT), blood urea nitrogen (BUN), uric acid, and creatinine using commercial diagnostic kits (Bayer’s Diagnostic, Pune, India) with semi-automatic biochemical analyzer (Erba, India) and clinical auto-analyzer (Chemwell 2910, USA). They also used fully automated Random Access Chemistry Analyser (Selectra Junior, Vital Scientific, Netherlands) using standard kits supplied by MERCK diagnostics reagents and systems.
The investigators performed the urine analysis for 24 h urine output. They followed the macroscopic examination, including color, pH, and specific gravity. They also performed a chemical examination to check the urine’s albumin, glucose, and ketone.
Necropsy, organ weight, and histopathology
At the end of the experiment, the investigators necropsied the animals. They immediately removed the organs such as the brain, liver, heart, lungs, kidneys, spleen, testes or ovary, intestine, stomach, and heart; weighed; and stored in 10% formalin for a histopathology examination.
| Results and Discussion|| |
Rasoushadhi (metal and mineral-based formulations) are widely used in the Ayurveda medicine system to treat the weakness of vital organs and systems. To take full advantage of the usage of potential sources of traditional health care, there is a need to draw attention to several issues related to policy on safety, efficacy, quality, accessibility, and rational use. Therefore, it is worthwhile to establish the safety of commonly used metal and mineral-based formulations of Ayurveda with widespread therapeutic applications. Several techniques have been adopted to assess the safety of novel drugs. Animal models help understand the safety risks of formulations with significance to comprehend the mechanism of meticulous toxicity., For the standardization of all the selected Rasoushadhi, the investigators followed the standard methods mentioned in Ayurvedic Pharmacopoeia of India (API) concerning different physicochemical analyses, viz., the estimation of loss on drying, ash content, acid insoluble ash, water/alcohol soluble extractive, pH, etc., qualitative/quantitative elemental testing, residual pesticide, microbiological examination, and tablet parameters, viz., hardness, friability, average weight, and dissolution time. They have quantitatively estimated the elemental assay by various techniques viz., atomic absorption spectrometer (Perkin Elmer [USA] Analyst 400), ICP-AES (Thermo Electron Corporations model IrisIntreprid II XDL), and also using by conventional methods as appropriate. They also carried out the powder x-ray diffraction analysis and X-ray photoelectron spectroscopy (XPS) measurements of the formulations using standard methods, and the outcome/findings of the chemical standardizations published separately.,,,,,,,
The present article appraises collective information on the safety profile of metal and mineral-based formulations of Ayurveda after repeated oral administration for 90 days in Wistar strain albino rats performed under the GTP scheme. The investigators of the study analyzed the findings. They reported that no mortality or significant signs of intoxication were observed in any control or drug-treated groups of animals throughout the study duration of 90 days. They found that Rasoushadhi-treated rats of both sexes exhibited the estimated pattern and similar body weight gain and feed consumption with those of control groups right through the dosing period, signifying normal growth, and development. Body weight gain and feed consumption are nonspecific and broad elements for assessing adverse systemic toxicity. The result of functional observation tests reported at different time points also did not reveal any abnormalities related to the administration of Rasoushadhi [Table 4].
|Table 4: General observation of subacute toxicity study of the selected Rasoushadhi|
Click here to view
Hematological and biochemical analyses conducted in the study revealed no dose-dependent treatment-related alterations, and all the values remained within the normal physiological range of the tested animals., The hemopoietic system is a significant target for several xenobiotics and a responsive indicator for various pathological states. The investigators opined that the treatment with all the Rasoushadhi did not produce any alterations in hematological parameters, which indicates the safety concern of the selected eight Rasoushadhi. They also observed that WBC and lymphocyte counts of the animals were mildly increased, whereas granulocyte (neutrophil) count was mildly decreased in the high dose–treated group of KY. The neutrophil count was mildly decreased in the high dose–treated group of KY. However, the findings were nonsignificant to indicate any sign of toxicity. While examining bone marrow smears, they found normal cellularity with fair numbers of functioning megakaryocytes in all the treated animals.
The liver plays a vital role in the metabolism and excretion of xenobiotics, making it highly predisposed to its adverse and toxic effects. The injury of hepatic cells can be diagnosed by specific biomarkers such as ALT, AST, ALP, and bilirubin. Augmented serum enzyme levels are taken as significant indicators of liver toxicity, whereas increases in total and conjugated bilirubin levels are indicators of overall liver function.,, Increase in the levels of ALT and AST in serum, combined with increased bilirubin levels, are essentially considered the most appropriate indicator of hepatic toxicity. The elevation of normal ALT and AST indicates their depletion into the bloodstream unsettled to damage parenchymal cells of the liver., The study revealed that the parameters of liver and renal functions were within the normal clinical range of animals in the Rasoushadhi or vehicle-treated groups, which indicates that all the Rasoushadhi did not affect the normal function or metabolism of the liver.
Early detection of drug-induced kidney injury is vital for the assessment of safety evaluation. The kidney is one of the major organs evoking drug-related toxic responses. It is an important target of toxicological studies as it plays a vital role in the excretion and detoxification of drugs or toxic metabolites. Serum creatinine and BUN are considered to be the fundamental parameters in preclinical or clinical trials for detecting and monitoring renal function. The values of serum creatinine or BUN levels reported in the study can be considered suggestive of the nontoxic nature of the Rasoushadhi to kidneys. Motor coordination is traditionally assessed in rodents by the rota rod test. Muscular strength or neuromuscular function in rodents may be influenced by sedatives, muscle relaxants, and toxic substances., In this study, the investigators found no significant changes in spontaneous motor activity and forced motor activity between the control and treatment groups. Assessing relative organ weights in toxicology studies is an integral module in assessing safety profiles. The Society of Toxicologic Pathology advocates the routine calculation and evaluation of organs or body weight ratios in toxicology studies.,,,,,,,, Organs or body weight ratios are considered more productive when body weights are affected., The mean data of relative organ weight of male and female rats euthanized at the end of 90 days of the treatment reported in the study were equivalent to that of their respective control groups. The gross pathological examination did not reveal any abnormal findings. The gold standard of pathology evaluation in toxicity studies has been the examination of paraffin-embedded, hematoxylin and eosin-stained tissue sections., The performed studies indicate a nonsignificant accumulation of metals in various tissues of animals that received the test drugs, and its proportion was within the permissible limits. The investigators explained that the levels of metals were found to decline in recovery groups.
| Conclusion|| |
In the said study, researchers observed no Rasoushadhi treatment-related abnormality concerning the gross or histological investigation of the organs examined. The hematology, clinical chemistry, organ weights, and gross and histopathological studies conducted as part of the study also demonstrated the nontoxicity of the test drugs in experimental animals. Thus, based on the study’s findings, the investigators affirmed the safety of the selected eight Rasoushadhi. Such studies may positively support the evidence-based practice of classical metal and mineral-based formulations of Ayurveda for centuries.
The authors express cordial gratitude to Dr. S. K. Sharma, former Advisor (Ayurveda), Dr. D. C. Katoch, Joint Advisor (Ay.), and Dr. G. P. Garg, Senior Chief Chemist, Ministry of Ayush, Government of India, Dr. G. S. Lavekar, Dr. K. S. Dhiman, former Director-General, and Dr. M. M. Padhi, former Deputy Director-General, CCRAS, for their precious support; Dr. Lalita Goyal and Dr. Naresh Kumar, CSIR, headquarters for extending her support in coordinating the related matters relating to CSIR; Dr. Poonam Kakkar and Dr. B. P. Chaturvedi of CSIR-IITR, Lucknow, Dr. Zabeer Ahmed and Dr. A. K. Saxena of CSIR-IIIM, Jammu, and Dr. J. Madhusudan Rao and Dr. Sreedhar Bojja, CSIR-IITC, Hyderabad, and the Directors, Former Directors of these institutes for the efforts in pharmacological and safety studies and chemical investigations; Dr. J Arunachalam, Former Head, National Centre for Compositional Characterisation of Material (BARC), Hyderabad, who has immensely contributed for interpretation of chemical studies. The authors are also thankful to Dr. V. K. Singh, M/s Maharishi Ayurveda Pharmacy for preparing of the formulations for the said study.
Financial support and sponsorship
Golden Triangle Project (GTP) Scheme under Ministry of Ayush, Government of India, New Delhi.
Conflicts of interests
There are no conflicts of interest.
| References|| |
Vagbhatta. Rasaratna Samuchchaya. Hindi commentary by Ambikadutt Shastri. 9th ed. Varanasi: Amarbharti Prakashan; 1995.
Savrikar SS, Ravishankar B Introduction to “rasashaastra” the iatrochemistry of Ayurveda. Afr J Tradit Complement Altern Med 2011;8:66-82.
Lavekar GS, Ravishankar B, Venugopal Rao S, Gaidhani S, Ashok BK, Shukla VJ Safety/toxicity studies of Ayurvedic formulation—Navratna Rasa. Toxicol Int 2009;16:37-42.
Pal S, Ramamurthy A, Mahajon B Arogyavardhini Vati: A theoretical analysis. J Sci Innov Res 2016;5:225-7.
Savrikar SS, Lagad CE “Preparation of Bhasma” in “Anonymous” Research Study Profile of Rasamanikya. New Delhi: Central Council for Research in Ayurveda and Siddha; 2009.
Srikanth N, Rao BCS, Sharma VK, Mahajon B, Khanduri S, Singh R, et al
. Safety and Prescription Trends of Rasaushadhis Critical Appraisal of Reported Medical Practices of Ayurveda Herbomineral Formulations from CCRAS Experience—Technical Report. New Delhi: Central Council for Research in Ayurvedic Sciences; 2021. p. 1-85.
Anonymous. Quality and Safety of Select Rasakalpa-Metal and Mineral Based Ayurvedic Formulations—Technical Report. 1st ed. Vol. I. New Delhi: Central Council for Research in Ayurvedic Sciences (CCRAS); 2019.
Anonymous. Quality and Safety of Select Rasakalpa-Metal and Mineral Based Ayurvedic Formulations—Technical Report. 1st ed. Vol. 2. New Delhi: Central Council for Research in Ayurvedic Sciences (CCRAS); 2019.
Dhiman KS, Srikanth N Quality and Safety of Select Rasakalpa-Metal & Mineral Based Ayurvedic Formulations—Arogyavardhini vati, Mahayogaraja guggulu, Vasantakusumakar rasa, Mahalaxmivilas rasa, Technical Report. Vol. 1. New Delhi: Central Council for Research in Ayurvedic Sciences (CCRAS); 2019. p. 1-256.
Dhiman KS, Srikanth N Quality and Safety of Select Rasakalpa-Metal & Mineral Based Ayurvedic Formulations-Rasamanikya, Makaradhwaja, Kajjaliyoga and Rasasindur, Technical Report. Vol. 2. New Delhi: Central Council for Research in Ayurvedic Sciences (CCRAS); 2019. p. 1-189.
Singh A, Ota S, Srikanth N, Sreedhar B, Dhiman KS Chemical characterization of an Ayurvedic herbo-mineral preparation—Arogyavardhani Vati: A potential tool for quality assurance. Indian J Traditional Knowledge 2018;17:176-83.
Singh A, Ota S, Srikanth N, Galib R, Bojja S, Dhiman KS Application of spectroscopic and chromatographic methods for chemical characterization of an Ayurvedic herbo-mineral preparation: Maha yograja guggulu. J Evid Based Integr Med 2018;23:2156587217751761.
Ota S, Singh A, Srikanth N, Sreedhar B, Ruknuddin G, Dhiman KS Chemical characterization of an Ayurvedic herbo-mineral formulation—Vasantakusumākara rasa: A potential tool for quality assurance. Anc Sci Life 2017;36:207-14.
Srikanth N, Singh A, Ota S, Sreedhar B, Galib , Dhiman KS Chemical characterization of an Ayurvedic herbo-mineral preparation—Mahalaxmivilas rasa. J Ayurveda Integr Med 2019;10:262-8.
Anonymous. The Ayurvedic Formulary of India (AFI) (Ministry of Health and Family Welfare, Govt. of India, New Delhi): Part I. 2nd ed. Delhi: The Controller of Publications; 2003. p. 68, 210, 212, 258, 267, 270, 273.
Anonymous. The Ayurvedic Formulary of India (AFI) (Ministry of Health and Family Welfare, Govt. of India, New Delhi): Part III. 1st ed. Delhi: The Controller of Publications; 2011. p. 306.
Anonymous. In: The Ayurvedic Pharmacopoeia of India, (Ministry of Health and Family Welfare, Govt. of India, New Delhi) Part I. 1st ed. vol. I. Delhi: The Controller of Publications; 1990.
Anonymous. In: The Ayurvedic Pharmacopoeia of India, (Ministry of Health and Family Welfare, Govt. of India, New Delhi) Part I. 1st ed. vol. III. Delhi: The Controller of Publications; 2001.
Anonymous. In: The Ayurvedic Pharmacopoeia of India, Minerals & Metals, (Ministry of Health and Family Welfare, Govt. of India, New Delhi) Part I. 1st ed. vol. VII. Delhi: The Controller of Publications; 2008.
Anonymous. In: The Ayurvedic Pharmacopoeia of India, (Ministry of Health and Family Welfare, Govt. of India, New Delhi) Part II. 1st ed. vol. I. Delhi: The Controller of Publications; 2008.
Anonymous. In: The Ayurvedic Pharmacopoeia of India, (Ministry of Health and Family Welfare, Govt. of India, New Delhi) Part II. 1st ed. vol. II. Delhi: The Controller of Publications; 2009.
Anonymous. Pharmacopoeial Standards for Ayurvedic formulations, (CCRAS, Ministry of Health and Family Welfare, Govt. of India, New Delhi), revised edition. Delhi: The Controller of Publications; 1987.
OECD. Test Guideline No. 408: Repeated Dose 90-Day Oral Toxicity Study in Rodents. Paris: OECD Publishing; 1998.
CPCSEA. Standard Operating Procedures (SOP) for IAEC. New Delhi: Committee for the Purpose of Control and Supervision of Experiments on Animals (CPCSEA); 2010. p. 41-2.
Reagan-Shaw S, Nihal M, Ahmad N Dose translation from animal to human studies revisited. Faseb J 2008;22:659-61.
Morgan SJ, Elangbam CS, Berens S, Janovitz E, Vitsky A, Zabka T, et al
. Use of animal models of human disease for nonclinical safety assessment of novel pharmaceuticals. Toxicol Pathol 2013;41:508-18.
Husain GM, Ahmed SS, Azhar M, Siddiqui JI, Waheed MA, Kazmi MH Comparative toxicity study on classical and modified version of jawarish jalinoos (a traditional unani formulation) in rats. Integr Med Res 2017;6:66-78.
Gad SC Repeat dose toxicity studies. In: Gad SC, editor. Preclinical Development Handbook: Toxicology. Hoboken, New Jersey: John Wiley & Sons Inc; 2008. p. 213-21.
Anonymus. Repeated Dose 28-day Oral Toxicity Study in Rodents, OECD Guidelines for the Testing of Chemicals, Section 4. Test No. 407; Paris: The Organisation of Economic Co-operation and Development (OECD) OECD Publishing; 2008. p. 1-13.
Kanu KC, Ijioma SN, Atiata O Haematological, biochemical and antioxidant changes in Wistar rats exposed to dichlorvos based insecticide formulation used in southeast Nigeria. Toxics 2016;4:28-9. doi: 10.3390/toxics4040028.
Devarbhavi H An update on drug-induced liver injury. J Clin Exp Hepatol 2012;2:247-59.
Wang J, Sun F, Tang S, Zhang S, Lv P, Li J, et al
. Safety assessment of vitacoxib: Acute and 90-day sub-chronic oral toxicity studies. Regul Toxicol Pharmacol 2017;86:49-58.
Arka G, Anindita K, Ankit S, Kumar SA, Kumar MS Preliminary evaluation of hepatoprotective potential of the polyherbal formulation. J Intercult Ethnopharmacol 2015;4:118-24.
Kang KS Abnormality on liver function test. Pediatr Gastroenterol Hepatol Nutr 2013;16:225-32.
Ozer J, Ratner M, Shaw M, Bailey W, Schomaker S The current state of serum biomarkers of hepatotoxicity. Toxicology 2008;245:194-205.
van Meer L, Moerland M, Cohen AF, Burggraaf J Urinary kidney biomarkers for early detection of nephrotoxicity in clinical drug development. Br J Clin Pharmacol 2014;77:947-57.
William MK Renal function test as indicator of kidney injury in subacute toxicity studies. Toxicol Appl Pharmacol 1981;57:414-24. doi:10.1016/0041-008x(81)90239-8.
Vogel HG, editor. Drug Discovery and Evaluation: Pharmacological Assays. 3rd ed. Germany: Springer-Verlag; 2008.
Campos FL, Carvalho MM, Cristovão AC, Je G, Baltazar G, Salgado AJ, et al
. Rodent models of parkinson’s disease: Beyond the motor symptomatology. Front Behav Neurosci 2013;7:175.
Sellers RS, Morton D, Michael B, Roome N, Johnson JK, Yano BL, et al
. Society of toxicologic pathology position paper: Organ weight recommendations for toxicology studies. Toxicol Pathol 2007;35:751-5.
Michael B, Yano B, Sellers RS, Perry R, Morton D, Roome N, et al
. Evaluation of organ weights for rodent and non-rodent toxicity studies: A review of regulatory guidelines and a survey of current practices. Toxicol Pathol 2007;35:742-50.
Thanabhorn S, Jaijoy K, Thamaree S, Ingkaninan K, Panthong A Acute and subacute toxicity study of the ethanol extract from lonicera japonica thunb. J Ethnopharmacol 2006;107:370-3.
Tucker DK, Foley JF, Hayes-Bouknight SA, Fenton SE Preparation of high-quality hematoxylin and eosin-stained sections from rodent mammary gland whole mounts for histopathologic review. Toxicol Pathol 2016;44:1059-64.
Shi SR, Liu C, Pootrakul L, Tang L, Young A, Chen R, et al
. Evaluation of the value of frozen tissue section used as “gold standard” for immunohistochemistry. Am J Clin Pathol 2008;129:358-66.
[Table 1], [Table 2], [Table 3], [Table 4]