Toxicology: Principles of Toxicology Department of Food Safety/Hygiene and Risk Management Rong-Jane Chen 2017 Sep 21

Introduction 聯絡小幫手 討論順序 主題: 13種列管化學品、基改食物、輻射污染食物、奈米保健食品或包材、油脂類、高溫烹調毒物、食品添加物 (醬油、零食)、食品中汙染物 (農藥殺蟲劑)等。 使用、ADME，毒理效應 (器官毒性、機制)，政府規範 (國內外)、毒理檢測 英文PPT檔案 (影片編輯)，列出參考資料 (包含網址) 不得抄襲 出席、討論、報告、考試

Ch. 2 Principles of Toxicology Casarett & Doull's Toxicology: The Basic Science of Poisons, 8th Edition Toxicology: the study of the adverse effects of xenobiotics and is a borrowing science that has evolved from ancient poisoners. 1900 1975 (safety evaluation/risk assessment) Albert Einstein (1879)，Maria Skłodowska-Curie (1867) Biomedical area, identification and quantification of hazards, occupational exposure, drug discovery, regulation design, mechanisms, clinical toxicologist (antidotes), food safety Obtained data about the toxicity of materials make reasonable predictions regarding the hazards of materials to people and the environment.

Ch. 2 Principles of Toxicology Antiquity The first toxicologist: Orfila (1787) Mustard gas (World War I) Wiley Bill: food and drugs Louis Casarett and John Doull: Toxicology text book (1975) 21th century: cellular and molecular mechanisms of toxicity, new animal models, epigenetic regulation (cancer, diabetes, neurodegenerative diseases and disorders), risk management. 神農 BC2000以前 Orfila 毒理作為獨立科學礦山從事繁重的手工業生產，勞動環境惡劣又缺乏防護，曾出現過多次嚴重急性中毒事件，如陶瓷工人發生鉛中毒，多數採礦工人發生汞中毒、磷中毒，礦工多數息有矽肺，甲醛中毒也屢見不鮮，為了適應當時的需要，許多學者圍繞各種職業接觸的化學物質開展大量實驗研究工作，使實驗毒理學獲得迅速發展其中Orfila做出巨大的貢獻，他曾在法國用幾千條狗做實驗，多次反覆地觀察到有毒物質與生物機體發生毒效應之間存在著「劑量-效應(反應)關係」，並建立了生物的組織和體液中鑒定毒物的化學分析方法，為法醫學和犯罪學開闢了用屍檢材料和化學分析方法作為中毒的法律證據，促進了法醫學和犯罪學的發展。1815 Orfila出版了第一本毒理學專著「Traite de loxicoloie（法語）」，並提出毒理學是一門獨立的學科。同期還有很多學者進行實驗研究，如 Wiley 檢驗酒精 色素 草藥

Ch. 2 Principles of Toxicology Toxicology: the study of the adverse effects of chemical or physical agents on living orgamisms. A toxicologist: examine and communicate the nature of these agents on human, animal, and environmental health. Examines the cellular, biochemical, molecular mechanisms of action and assesses the probability of their occurrence. Exposure Response Risk assessment: the quantitative estimate of the potential effects on human health and environmental significance of various types of chemical exposures.

Ch. 2 Principles of Toxicology 3 main categories: descriptive, mechanistic, regulation Mechanistic results: adverse response and mechanisms Ex: sweetener saccharin: bladder cancer in rat in human: crystalline precipitate (high conc.) Safer alternative chemicals, therapy of poisoning, ex thalidomide Genome-wide studies: toxicogenomics

Ch. 2 Principles of Toxicology Descriptive toxicology: provides information for safety evaluation and regulation requirements (appropriate toxicity tests). Cell culture systems and experimental animals. “omics”: genomics, transcriptomics, metabonomics/metabolomics… Regulatory toxicologist: has the responsibility for deciding, on the basis of data (descriptive and mechanistic toxicology), whether a drug or a chemical has a sufficiently low risk (or a benefit profile) to be marked. 台灣: 衛生福利部食品藥物管理暑，行政院環境保護署毒物及化學物質局 農藥管理: 行政院農委會 (地方，其他主管機關) US FDA: food additives, USEPA: other chemicals Forensic toxicology, clinical toxicology, environmental toxicology, ecotoxicology

Paracelsus (1493-1541): What is there that is not poison Paracelsus (1493-1541): What is there that is not poison? All things are poisons….Solely the dose determines that a thing is not a poison. Dose-response relationship Low dose effects

Toxic Agents/Undesired effects Classified in a variety of ways, depending on the needs and the interests of the classifiers. Target organ, use, source, effects, chemical structures… Toxin: biological systems (plants, animals, fungi, bacteria etc.) Toxicant: produced by or a by-product of human-made activities or natural processes. (TCDD, PAH) Undesired effects: allergic reactions, idiosynchratic reactions (abnormal reactivity to a chemical, low frequency), immediate and delayed toxicity, reversible and irreversible effects, local and systemic toxicity, interaction of chemicals, tolerance (biotransformation enzyme induction).

Characteristics of exposure 1.Routs and sites of exposure (Ch 5) 2.Duration and frequency of exposure: Acute: < 24 hrs, single dose Subacute: repeated,  1 month Subchronic: repeated, 1-3 month Chronic: repeated, > 3 month (1 yr) Effects of single or repeated dose are different: benzene (CNS depression or bone marrow toxicity)

2.Duration and frequency of exposure: Elimination rate is important, half life for elimination: 50% of the chemical to be removed from the blood stream Interval, repair of tissue damage Frequency is important than duration of exposure. 1 yr 1 day 5 hrs

Dose-Response Relationship The characteristics of exposure and the spectrum of toxic effects come together in a correlative relationship customarily referred to as the dose–response relationship. The most fundamental and pervasive concept in toxicology. 1. Individual, or Graded, Dose–Response: the response of an individual organism to varying doses of a chemical, often referred to as a “graded” response. because the measured effect is continuous over a range of doses. 2. Quantal Dose–Response Relationships: the distribution of individual responses to different doses in a population of individual organisms.

1. Individual, or Graded, Dose–Response brain 1. Individual, or Graded, Dose–Response: a dose related increase in the severity of the response results from an alteration of a specific biochemical process. Organophosphate insecticide chlorpyrifos:毒死蜱 Inhibits enzymes in the brain and liver: acetylcholinesterase and carboxylesterase. In the brain, cholinesterase is more easily inhibited than carboxylesterase.

2. Quantal Dose–Response Relationships hypersusceptible resistant 50% 2. Quantal Dose–Response Relationships: in a population are by definition quantal—or “all or none” At any given dose, an individual in the population is classified as either a “responder” or a “non-responder.” Estimating the response of a population to a toxic exposure is the “Effective Dose” or ED. “ED50” value. Where death is the measured end-point, the ED50 would be referred to as the Lethal Dose 50 (LD50). (new chemical) Differences in susceptibility to chemicals

2. Quantal Dose–Response Relationships The dose–response curves for the response of A and B A: a large change in dosage is required before a significant change in response. B: “steep” dose–response curve, where a relatively small change in dosage will cause a large change in response. ED50 the same (8 mg/kg)

Shape of the Dose–Response Curve The shape of the dose–response relationship has many important implications in toxicity assessment. (Vitamin A) U-shaped. high doses of vitamin A can cause liver toxicity and birth defects. Hormetic effects: when relatively low doses result in the stimulation of a beneficial or protective response.

Shape of the Dose–Response Curve Nonnutritional toxic substances may also impart beneficial or stimulatory effects at low doses but at higher doses, they produce adverse effects: hormesis. Ability of substances to stimulate biological systems at low doses but to inhibit them at high doses. chronic alcohol consumption

Threshold Another important aspect of the dose–response relationship at low doses is the concept of the threshold. Acute toxicologic responses are associated with thresholds. Some dose below which the probability of an individual responding is zero. Acute toxicity, less in chronic toxicity, especially in the area of chemical carcinogenesis. The apparent threshold, or “no observable adverse effect level” (NOAEL) threshold

Nonmonotonic Dose-Response Curves Fig. 2-10 NMDR: upregulation of some receptors at low conc, down regulation of receptors at high conc. Interconnected mol pathways. Threshhold non-threshhold NMDR terminal end bud

Evaluating the Dose–Response Relationship effective dose (ED) toxic dose (TD): liver injury lethal dose (LD) therapeutic index (TI): comparison of the therapeutically effective dose to the toxic dose of a chemical. Defined as the ratio of the dose required to produce a toxic effect and the dose needed to elicit the desired therapeutic response. Therapeutic Index = TD50/ED50 The larger the ratio, the greater the relative safety. The use of the median effective and median toxic doses is not without disadvantages, because median doses tell nothing about the slopes of the dose–response curves for therapeutic and toxic effects. Fig 2-11

Potency versus Efficacy Fig. 2-12 Thus, A is more potent than B and C is more potent than D A and B have equal maximal efficacy, whereas the maximal efficacy of C is less than that of D.

DESCRIPTIVE ANIMAL TOXICITY TESTS Fig. 2-13 Two main principles: 1. the effects produced by a compound in laboratory animals, when properly qualified, are applicable to humans. 2. exposure of experimental animals to chemicals in high doses is a necessary and valid method of discovering possible hazards in humans. Saccharin Toxicity tests are not designed to demonstrate that a chemical is safe but to characterize the toxic effects a chemical can produce.

Toxicity Testing 1. Acute Toxicity Testing: single high dose, objectives: (1) provide an estimate of the intrinsic toxicity of the substance, lethal dose (e.g., LD50), 14 days. (2) provide information on target organs and other clinical manifestations of toxicity (3) identify species differences and susceptible species (4) establish the reversibility of the toxic response (5) provide information that will assist in the design and dose selection for longer term (sub-chronic, chronic) studies. If no toxicity is evident at 2 g/kg, further acute dermal toxicity testing is usually not performed. inhalation studies: the length of exposure is 4 hours

Toxicity Testing 2. Sub-acute Toxicity Testing: obtain information on the toxicity of a chemical after repeated administration and as an aid to establish doses for sub-chronic studies. To give three to four different dosages of the chemicals to the animals by mixing it in their feed. Clinical chemistry and histopathology are performed after 14 days of exposure. (<1 month)

Toxicity Testing 3. Sub-chronic Toxicity Testing: objectives: 1-3 month Establish a NOAEL Identify and characterize the specific organ or organs affected by the test compound after repeated administration. EPA utilizes the NOAEL to calculate the reference dose (RfD), which may be used to establish regulatory values for “acceptable” pollutant levels. rat and dog for FDA; and mouse for EPA, oral 3 doses, (a high dose that produces toxicity but does not cause more than 10% fatalities, a low dose that produces no apparent toxic effects, and an intermediate dose) Observed once or twice daily for signs of toxicity Drug: acute and subchronic studies must be completed before the company can file an Investigational New Drug (IND) application with the FDA

Toxicity Testing 4. Chronic Toxicity Testing: 6 months to 2 years. If the chemical is a food additive with the potential for lifetime exposure in humans, a chronic study up to 2 years in duration is likely to be required. Most regulatory guidelines require that the highest dose administered be the estimated maximum tolerable dose (MTD) one or two additional doses, usually fractions of the MTD (1/2 of MTD and ¼ of MTD), and a control group are tested Include a consideration of the carcinogenic potential of chemicals so that a separate lifetime feeding study that addresses carcinogenicity does not have to be performed.

Toxicity Testing 1 D: acute toxicity: single dose, high dose, LD50 1 wk (2 wk): sub-acute: repeated dose 1 month (1-3): sub-chronic: repeated dose, NOAEL 1 yr (0.5-2): chronic: repeated dose, MTD

Toxicity Testing NeurotoxicityAssessment The developing nervous system is sensitive (cell lines, food additives) Eyes toxicity Inhalation toxicity acute toxicity Skin toxicity Mutagenicity screen for potential carcinogens Developmental and Reproductive Toxicity Oncogenicity Bioassays suspect that a chemical may be carcinogenic, or when there may be wide spread, long term exposures to humans (food additives), life time exposure Immunotoxicity Assessment -Immunosuppression - Hypersensitivity - Autoimmunity

Toxicity Testing Fug. 2-14, the large number of animals that would be required to test the potential carcinogenicity of a chemical at the doses usually encountered by people.

Toxicity Testing Fig. 2-15, tumors, both benign and malignant, are not uncommon events in animals even in the absence of exposure to any known carcinogen. different tumor types that develop “spontaneously” in both sexes of both rats and mice, but at different rates. Background tumors that are common in one species may be uncommon in another.

TOXICOGENOMICS Fig 2-17, genomics, transcriptomics, proteomics metabonomics (characterization of most or all of the small molecules in a cell or tissue, including substrates, products, and co-factors of enzyme reaction Systems toxicology:combines transcript, protein and metabolite profiling, with conventional toxicology to investigate the interaction between, genes and environmental stress in disease causation Nuclear Magnetic Resonance (NMR), and mass spectrometry

TOXICOGENOMICS Epigenetics/epigenomics: something acting above or in addition to genes DNA methylation, histone acetylatin Tobacco smoke, BPA, alcohol, nutrition, Vit.

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Threshold DBC: dibenzo[a,l]pyrene, AFB1: alpha toxin

TOXICOGENOMICS Fig. 2-18

2. Quantal Dose–Response Relationships 校正之後

Toxicity Testing Fig. 2-16