Toxicity studiesDr. S. Parasuraman, M.Pharm., Ph.D.,Senior Lecturer, Faculty of PharmacyAIMST University
Toxicity studies- Introduction• Toxicology is a branch of science that deals with toxins and poisonsand their effects and treatment.• Toxicological screening is very important for the development ofnew drugs and for the extension of the therapeutic potential ofexisting molecules.• The US-FDA states that it is essential to screen new molecules forpharmacological activity and toxicity potential in animals (21CFRPart 314).• Toxicity tests are mostly used to examine specific adverse events orspecific end points such as cancer, cardiotoxicity, and skin/eyeirritation.• Toxicity testing also helps calculate the No Observed Adverse EffectLevel (NOAEL) dose and is helpful for clinical trails.
History of Toxicity Studies• Paracelsus (Father of Toxicology): determinedspecific chemicals responsible for the toxicity ofplants and animals (dose-response relationship).• "All substances are poisons; there is none which isnot a poison. The right dose differentiates a poisonand a remedy”– Paracelsus• Mathieu Orfila, determined the relationshipbetween poisons and their biological He is referredto as the father of modern toxicology.Paracelsus(1493-1541)Recent developments: after 1920(introduced determine LD50 byUSFDA )
Sources of toxic substances• Classified based on their– chemical nature– mode of action– class (exposure class and use class)• Exposure class: Food, air, water or soil.• Use class: drugs as drug of abuse, therapeutic drugs, agriculturechemicals, food additives, pesticides, plant toxins and cosmetics.
Biomedical ethics• Before conducting any toxicological testing in animals or collectingtissue/cell lines from animals, the study should be approved by theInstitute Animal Ethics Committee (IAEC) or the protocol shouldsatisfy the guidelines of the local governing body.• Malaysia:– http://www.animalethics.org.au/animal-ethics-committees
Acute toxicity studies• Acute toxicity testing- study the effect of a single dose on aparticular animal species.• Acute toxicity testing be carried out with two different animalspecies (one rodent and one non-rodent).• In acute toxicological testing, the investigational product isadministered at different dose levels, and the effect is observed for14 days. All mortalities caused by the investigational product duringthe experimental period are recorded andmorphological, biochemical, pathological, and histological changesin the dead animals are investigated.
Acute toxicity studies• The LD50 was used as an indicator of acute toxicity previously. Thedetermination of the LD50 involves large numbers of animals, andthe mortality ratio is high. (24 h testing)– Graphical method– Arithmetical method (karbers’s) method. when number of animal issmall)• Because of theselimitations, modified methodswere developed:1. The fixed dose procedure2. The acute toxic category method3. The up-and-down methodCont.,
Acute toxicity studies• Signs recorded during acute toxicity studiesCont.,• Increases motor activity• Decreased motor activity• Tremors and ataxia• Tonic convulsion• Catatonia• loss of righting reflex• Sedation• muscle relaxation• Piloerection/ straub tail phenomenon• Cannibalism• Straub reaction• Hypnosis• Analgesia• Lacrimation• Diarrahoea• Salivation: viscid or watery• Writhing• Respiration:depression, stimulation orfailure.• Skin colorPiloerection PiloerectionNormal
Sub-acute toxicity studies• Rodents and non-rodents are used to study the subchronic toxicityof a substance.• Dose: Expected therapeutic level (daily) or expected therapeuticlevel to increasing dose phase-wise manner.• The test substance is administered orally for =/>90 days, andregular body weight variations, biochemical and cardiovascularparameters changes, and behavioral changes are observed.• At the end of the study, the experimental animals are sacrificed.Gross pathological changes are observed, and all the tissues aresubjected to histopathological analyses.• There should be little individual variation between the animals, andthe allowed weight variation range is ±20%.• Used to determine the maximum tolerable dose and nature oftoxicity.
Chronic toxicity studies• Chronic toxicity studies are conducted with a minimum of onerodent and one nonrodent species.• The test compound is administered over more than 90 days, andthe animals are observed periodically.• Dose:• A chronic toxicology study provides inferences about the long-termeffect of a test substance in animals, and it may be extrapolated tothe human safety of the test substance.• The report on chronic oral toxicity is essential for new drug entities.There should be little individual variation between the animals, andthe allowable weight variation range is ±20%.
• High dose: Produce significant retardation of growth or somepathological changes (10 times the expected maximum clinicaldose).• The low dose is about twice the expected maximum clinicaldose• Third dose is (medium dose) fixed midway between the highand low doseChronic toxicity studies Cont.,
Chronic toxicity studies• During the study period, the animals are observed for normalphysiological functions, behavioral variations and alterationsin biochemical parameters at regular intervals (atleast every14 days). At the end of the study, tissues are collected from allparts of the animal and subjected to histological analyses.