PRINCIPLES OF MEDICAL TOXICOLOGYchanif.lecture.ub.ac.id/files/2019/09/farmakologi-pertemuan-6-toksikologi-1a.pdfRute Pemaparan Toksikan (Exposure) •Rute dan Titik tangkap Pemaparan

PRINCIPLES OF MEDICAL

TOXICOLOGY

Prof.Dr. H.Achmad Basori, MS

Departement of Pharmacology

Medical Faculty

Airlangga University

Basic Science

Biology, Biochemistry,Pathology, Physiology, Genetic, Pharmacology

TOXICOLOGY

Medical Toxicology :

- Biochemical Toxicology

- Analytical Toxicology

- Cellular Toxicology

- Molecular Toxicology

-- Clinical Toxicology

-- Forensic Toxicology

Food Toxicology

Ecotoxicology

Industrial Toxicology

Enviromental Toxicology

Occupational Toxicology

Developmental

and reproductive Toxicology

Regulatory Toxicology

Mechanistic Toxicology

Descriptive Toxicology

Ukrainian president Viktor Yushchenko, after alleged

poisoning with dioxin, and, possibly endotoxin, prior to

the 2004 elections.

•

MOLECULES OF DEATH

1.

1.Aflatoxin

2. Botulinus Toxin

3. Carbon Monoxide – Ther Silent Killer

4. Domoic Acid

5. Ecstacy

6. Heroin

7.Hydrofluoric Acid

8.Hydrogen Sulphide

9.Lead : An old and Modern Poison

10.Mercury

11.Mushroom Toxin

12.Nerve Gases

13.Nicotine and Tobacco Alkaloid

14.Paracetamol (Acetominophen)

15.Paraquat and Diquat

16.Phosphorus

17.Radon

18.Ricin

19.Snake Toxin

20.Spider Toxin

21.Strychnine

22.Tetrodotoxin

23.Thallium

24.Arsen

25.Cyanide

Keracunan bahan kimia di IRD RSUD Dr. Soetomo Surabaya dalam 5 tahun terakhir (Hernomo, 2001)

Nama Bahan 1996 1997 1998 1999 2000

1. Pestis. 128 (32.82%) 150 (29.30%) 84 (22.11%) 75 (22.52%) 78 (31.84%)

2. Ob. Farm. 120 (30.77%) 227 (44.34%) 159 (41.84%) 137 (41.14%) 81 (33.06%)

3. Minyak 60 (15.38%) 45 (8.79%) 29 (7.63%) 38 (11.41%) 32 (13.06%)

4. Makanan 13 (3.33%) 35 (6.84%) 39 (10.26%) 23 (6.91%) 8 (3.27%)

5. Alkohol 24 (6.15%) 14 (2.73%) 22 (5.79%) 30 (9.01%) 20 (8.16%)

6. Rmh tng 8 (2.05%) 11 (2.15%) 7 (1.84%) 5 (1.50%) 3 (1.22%)

7. Gas 2 (0.51%) 4 (0.78%) 2 (0.53%) 0 (0%) 0 (0%)

8. Ob. Trad. 11 (2.82%) 3 (0.59%) 6 (1.58%) 12 (3.60%) 2 (0.82%)

9. Korosif 18 (4.62%) 14 (2.73%) 10 (2.63%) 11 (3.30%) 5 (2.04%)

10. Lain-lain 2 (0.60%) 0 (0%) 0 (0%) 0 (0%) 3 (1.22%)

11. Tak diket. 6 (1.54%) 16 (4.21%) 0 (0%) 0 (0%) 13 (5.31%)

Total 390 (100%) 512 (100%) 380 (100%) 333 (100%) 245 (100%)

Target Organ Toxicity

Central Nervous System – lead

Immune System - isocyanates

Liver - ethanol, acetaminophen

Respiratory Tract - tobacco smoke, asbestos, ozone

Eye - UV light (sunlight)

Kidney - metals

Skin - UV light, gold, nickel

Reproductive System –

dibromochloropropane

Ecotoxicology toksikologi ekosistem

Bioaccumulation = the accumulation of a contaminant

or toxin in or on an organism

from all sources (e.g., food, water, air).

Biomagnification = the increase in concentration of

toxin as it passes through successive levels of the

food web

Bioaccumulation

• Assimilation Efficiency (= Lindeman’s Efficiency

Lindeman 1942. Ecology 23: 399-418)

• AE increases with trophic level

• When a chemical is assimilated more efficiently than

organic energy -> bioaccumulation

AE

Biomagnification

Scenario 1: Alewife (2o predator) eats Cercopagis 1o predator

1 10 100 1 100

cals.

ppm toxin 10,000

1 100

1 1000

cals.

ppm toxin

Scenario 2

Food Web Bioaccumulation

The Mercury Cycle

Toxicokinetis and Toxicodynamics

Karakteristik

Rute Pemaparan Toksikan

(Exposure)

• Rute dan Titik tangkap Pemaparan

– Ingestion (Gastrointestinal Tract)

– Inhalation (Lungs)

– Dermal/Topical (Skin)

– Injection

• intravenous, intramuscular, intraperitoneal

• Effectiveness pemaparan:

iv > inhale > ip > im > ingest > topical

Dosis Jumlah bahan kimia / Toxicant yang memasuki

tubuh Umumnya dalam satuan mg /kg BW

Dosis Toxicant tergantung pada bbp faktor :

* concentration di lingkungan sekitarnya

* Karakteristik exposure

* Lama exposure

Frekwensi exposure

* Sifat toxicant

TOXICOKINETICS:

Study of the time-course of toxins (study of what the body does to the

toxin).

TOXICODYNAMICS:

Study of biochemical and physiological effects of toxicants (study of

what the toxicant does to the body).

TOXICOKINETIC:

Absorption, Distribution, Metabolism, and

Excretion

• Toxicant tubuh manusia target site

adverse effect.

• Tubuh mempunyai pertahanan :

– Membrane barriers

• Passive dan facilitated diffusion, active

transport

– Enzim Biotransformasi , antioxidants

– Mekanisme Eliminasi

Non-ionised drug

More lipid soluble drug

Diffuse across cell

membranes more easily

Ionised drug

Less lipid soluble drug

TOXICANT

v Asam lemah : HA H+ + A-

Persamaan Henderson-Hasselbach :

pH = pK’ + Log10[A-]/[HA]

v Basa lemah :BH+ B + H+

Persamaan Henderson-Hasselbach :

pH = pK’ + Log10[B]/[BH+]

TOXICANT : asam lemah / basa lemah , Aspirin, Barbiturates (acid), Propranolol,

Opioids (base)

Asam lemah Basa lemah

H+

HA A-

HA

H+

A-

B BH+

H+

H+

B BH+

ionized = polar = water soluble

non-ionized = nonpolar = more lipid-soluble

Ion Trapping HA <==> H+ + A- B + HCl <==> BH+ + Cl-

[ UI ] [ I ] [ UI ] [ I ]

pKa=pH+log(HA/A-) pKa=pH+log(BH+/B)

pKa = 4.5 (Toxicant : a weak acid)

100 = [ UI ] [ UI ] = 100

pH = 2

pH = 7.4

0.1 = [ I ] [ I ] = 9990

ATP

ADP-Pi

Passive diffusion

Carrier-mediated transport

Active Facilitated

Transporter Molecule

MEKANISME TRANSPORT

DARI TOXICANT

• Memerlukan carrier

• Transport menjadi jenuh (saturated) pada

konsentrasi tinggi

• Proses bersifat selective

• Dua obat yang ditranspor oleh mekanisme

yg sama akan menghambat satu sama lain

• Melawan concentration gradient ( active

transport)

Tdk melawan cocentration gradient (

facilitated

transport)

• Memerlukan energy

• Mekanisme transport dapat dihambat oleh

obat obat yang mempengaruhi cellular

metabolism

Karakteristik facilitated diffusion dan active transport

Un-ionized Ionized

Pharmacologic effect Active Inactive

Solubility Lipids Water

Cross lipid barriers Yes No (gastrointestinal tract,

blood-brain barrier, placenta)

Hepatic metabolism Yes No

Renal excretion No Yes

Karakteristik dari molekul Un-ionized

Dan Ionized Toxicant

Absorption: Kemampuan bhn kimia memasuki darah

(darah berkesimbangan dgn jaringan) • Inhalasi--gas menuju darah melalui alveoli. (luas

permukaan alveolar, aliran darah banyak, lapisan antara darah menuju alveolar air)

• Ingestion--absorpsi melalui GI tract : stomach (asam), small intestine (contact time panjang, luas permukaan luas--villi; bases dan transporter bahan bahan tertentu)

– 1st Pass Effect (liver metabolism)

• Dermal—absorpsi melalui epidermis (stratum corneum), dermis; titik tangkap dan keadaan kulit

• Surface area

approximately 50 to

100 m2 Nasopharynx

Oropharynx

Right main bronchus

Pharynx

Thyroid cartilage

Cricoid cartilage

Epiglottis

Lungs

Larynx

Bronchiole

Diaphragm

Trachea

Left main bronchus

Bronchiole

Alveolus

Alveolar sac

Respiratory System

Respiratory Physiology

Aveolus

.

Blood from right side of heart

Reoxygenated blood

Blood to left side of heart

Red blood cells

Capillary

(low in O,

high in CO) 2

2

(high in O, low in CO) 2 2

O 2

CO 2 CO 2

CO 2

CO 2

O 2

O 2 O 2

Absorpsi Pulmonary • Systemic (e.g. insulin, anesthetics)

dan local delivery

• Area absorpsi sangat luas

• Suplai darah sangat baik

• Tidak mengalami first pass effect

• Bentuk sediaan mahal

• Ukuran partikel : 2-5 m

Absorption of Aerosols and

Particles:

1- Particle Size

2- Water solubility of the

chemical present in the

aerosol or particle

REMOVAL OF

PARTICLES

Physical

Phagocytosis

Lymph

Absorption from the Lungs

Pemberian per inhalasi

• Patikel > 10 um : diendapkan, dihembuskan dan

berbangkis

• Partikel < 0.01 um : terbuang pada saat inspirasi dan

ekspirasi

• Partikel 0.01 – 10 um :diendapkan pada alveoli,

nasopharyng sampai bronchioli

• 25% dikeluarkan bersama udara nafas

• 50% diendapkan disalurannafas bagian atas

• 25% diendapkan disaluran nafas bagian bawah

Absorpsi dari Paru

• Gas, vapors,volatile liquids, aerosols and

particles

• Large surface area, thin barrier, high blood

flow rapid absorption

• Blood:air partition coefficient –

dipengaruhi respiratory rate dan blood flow

• Blood:tissue partition coefficient

Nasopharyngeal

Region

5-30 µm

Trachea

Bronchi

Bronchioles

1-5 µm

Alveolar Region

1 µm

DEPOSISI PARTIKEL TOKSIKAN

DI DLM SALURAN RESPIRASI

Absorpsi dari kulit • Melewati bbg lapisan sel (stratum

corneum, epidermis, dermis) menuju

pembuluh darah .

• Faktor yang mempengaruhi :

lipid solubility, hydrasi kulit

(sole of feet vs. scrotum)

Absorption by the Skin

Absorpsi melalui kulit • Permeability depends on the diffusivity and thickness

(depends on the area of the body) of the stratum

corneum

• Polar outer surface of protein filaments of the

hydrated stratum corneum

• Nonpolar lipid matrix between protein filaments

• Percutaneous absorption lower layers of the

epidermis and dermis

• Below the s.corneum porous, nonselective aqueous

medium

• Compromised stratum corneum integrity

• Increased stratum corneum hydration

• Increased temperature increased blood flow

• Low solubility of toxicant in the vehicle • Small size Increased Absorption

Distribution:

proses translokasi dari Toxicant menuju

seluruh bagian tubuh

• Darah membawa Toxicant menuju site of

action, storage depots, organ transformasi, dan

organ eliminasi

• Kecepatan distribusi Toxicant tergantung :

-- aliran darah

– karakteristik toxicant (afinitas thd jaringan dan

partition coefficient)

• Distribusi mungkin berubah setiap waktu

Distribusi:

Storage / Binding • Storage di dlm Adipose tissue sangat

lipophylic (DDT). Cepat dimobilisasi dari fat

(starvation) , cepat meningkat dalam darah

cepat meningkat dalam darah

• Storage dalam tulang (Bone) Chemicals

analog dgn Calcium--Fluoride, Lead,

Strontium

• Ikatan dgn Plasma proteins mendesak

senyawa endogenous . Hanya fraksi bebas

adverse effects dan excretion

Metabolism:

• Toxicant lebih water soluble (Polar)

ekskresi

– Menurunkan lipid solubility

menurunkan jumlah toxicant pada

target

– Meningkatkan ionisasi

meningkatkan excretion rate -->

menurunkan toxicity

• Bioactivasi Biotransformasi

pembentukan reactive metabolites

Biotransformation (Metabolism)

• Meningkatkan

kec clearance

dari toxicant

• Dapat terjadi

mulai absorpsi

ekskreri

Toxicant Tanpa

Metabolisme

Dengan

Metabolisme

Ethanol 4 minggu 10mL/hr

Phenobarbital 5 bulan 8hrs

DDT infinity Bbp hari

bbp minggu

Biotransformation

• Key organs in biotransformation

– LIVER (high)

– Lung, Kidney, Intestine (medium)

– Others (low)

• Biotransformation Pathways

* Phase I--make the toxicant more water

soluble

* Phase II--Links with a soluble endogenous

agent (conjugation)

Individual Susceptibility

--there can be 10-30 fold difference in response to a

toxicant in a population

• Genetics-species, strain variation, interindividual variations (yet still can extrapolate between mammals--similar biological mechanisms)

• Gender (gasoline nephrotox in male mice only)

• Age--young (old too)

– underdeveloped excretory mechanisms

– underdeveloped biotransformation enzymes

– underdeveloped blood-brain barrier • Age--old

– changes in excretion and metabolism rates, body fat

• Nutritional status

• Health conditions

• Previous or Concurrent Exposures

– additive --antagonistic

– synergistic

Distribution:

the process in which a chemical agent translocates

throughout the body

• Blood carries the agent to and from its site of action, storage

depots, organs of transformation, and organs of elimination

• Rate of distribution (rapid) dependent upon

– blood flow

– characteristics of toxicant (affinity for the tissue, and the

partition coefficient)

• Distribution may change over time

Distribution:

Storage and Binding

• Storage in Adipose tissue--Very lipophylic

compounds (DDT) will store in fat. Rapid

mobilization of the fat (starvation) can rapidly

increase blood concentration

• Storage in Bone--Chemicals analogous to Calcium--

Fluoride, Lead, Strontium

• Binding to Plasma proteins--can displace

endogenous compounds. Only free is available for

adverse effects or excretion

Target Organs: adverse effect is dependent upon the

concentration of active compound at the target site for

enough time

• Not all organs are affected equally

– greater susceptibility of the target organ

– higher concentration of active compound

• Liver--high blood flow, oxidative reactions

• Kidney--high blood flow, concentrates chemicals

• Lung--high blood flow, site of exposure

• Neurons--oxygen dependent, irreversible damage

• Myocardium--oxygen dependent

• Bone marrow, intestinal mucosa--rapid divide

Target organ

• Carbon tetrachloride – liver

• Mercury & lead – CNS, kidneys &

hematopoietic system

• Benzene – hematopoietic system

Storage sites

• Dichlorodiphenyltrichloroethane (DDT) –

fat depots no toxic effect

• Nose is a ―scrubber‖ for water-soluble and highly reactive gases

• Solubility ratio (blood-to-gas partition coefficient) – conc. in blood/conc. in gas phase before or at saturation

• Low solubility ratio – blood flow through the lungs (perfusion-limited)

• Highs solubility ratio – rate and depth of respiration (ventilation-limited)

• Lungs are capable of biotransformation & elimination

• Steady state concentration can be reached

• Aerosols dependent on aerosol size & water solubility

• 5um or more – lodged in nasopharyngeal region

• 2.5 um – tracheobronchial region

• 1 um or less – alveolar sacs of blood

ABSORPSI

DISTRIBUSI

METABOLISME

ELIMINASI

INTERAKSI TOKSIKAN – RESEPTOR

TOXIC EFFECTS

TOXICOKINETICS

TOXICODYNAMICS

PEMAPARAN (EXPOSURE ) TOXICANT

Molecular effects Cellular effects Tissues effects Organ effects

Interaksi Toxicant – reseptor

Perubahan fungsi biokimia sel

Perubahan fisiologik jaringan

Perubahan fisiologik organ

Sign and symptom Efek Toksik

Toxic effects

Toxicant

Allergic (hypersensitivity,Antigen)

Idiosyncratic (e.g. G6PD def., Drugs)

Local vs. Systemic (Corrosive agent)

Reversible vs. Irreversible

Necrosis /organ damage (Ozone, Lead, Cd, Sr)

Carcinogenecity (Benzene, Rokok, Asbestos, Coloring Agent)

Mutagenicity (uv light, Coloring Agent)

Teratogenicity (Drugs:Thalidomide, Valproic acid, Herbal)

Death (Arsen, Cyanide)

Efek Toxic Berdasarkan Mekanisme

Efek Toksik Berdasarkan

Lama Pemaparan (Exposure)

Acute toxicity < 24hr umumnya 1 x paparan

Subacute toxicity 1 bulan dosis berulang

Subchronic toxicity 1-3 bulan dosis berulang

Chronic toxicity > 3 bulan dosis berulang

Pada pemakaian berulang akumulasi Toxicant didalam tubuh

Target Organs: adverse effect tergantung pada

kadar senyawa aktif dlm target site untuk waktu

yang cukup

• Tidak semua organ dipengaruhi sama ,tetapi

tergantung

– Kepekaan target organ

– Kadar toxicant yg tinggi dalam target organ

• Liver—aliran drh sangat tinggi,oxidative reactions

• Kidney—aliran drh sangat tinggi, bhn kimia

terkonsentrat

• Lung--high blood flow, tempat pemaparan

• Neurons--oxygen dependent, kerusakan irreversible

• Myocardium--oxygen dependent

• Bone marrow, intestinal mucosa -- rapid divide cell

Target Sites:

Mechanisms of Action

• Adverse effects can occur at the level of

the molecule, cell, organ, or organism

• Molecularly, chemical can interact with

Proteins Lipids DNA

• Cellularly, chemical can

– interfere with receptor-ligand binding

– interfere with membrane function

– interfere with cellular energy production

– bind to biomolecules

– perturb homeostasis (Ca)

Excretion: Toxicants are eliminated from the

body by several routes

• Urinary excretion

– water soluble products are filtered out of

the blood by the kidney and excreted into

the urine

• Exhalation

– Volatile compounds are exhaled by

breathing

• Biliary Excretion via Fecal Excretion

– Compounds can be extracted by the liver

and excreted into the bile. The bile drains

into the small intestine and is eliminated in

the feces.

• Milk Sweat Saliva

Mekanisme kerusakan sel (cellular injury)

1. Perubahan permeabilitas cell membrane

2. Perubahan enzymes activity.

3. Modifikasi carriers.

4. Reaksi yg menyebabkan deplesi GSH.

5. Interaksi dgn co-enzyme.

6. Interaksi dgn nucleic acid.

7. Pembentukan reactive metabolite.

8. Perubahan protein synthesis.

9. Immunotoxicity.

10. Perubahan Lysosomal

11. Inhibisi cellular respiration.

Acute Toxicity LD 50,Max Tolerated Dose,2 species,2

route, single dose

Subacute Toxicity 3 doses,2 doses, 4 weeks-3 months,

Chronic Toxicity Rodent,non-rodent, 6 months and more

Effect on reproductive performance Effects on animal mating

behavior,reproduction,parturition,prog

eny,birth defects,postnatal

development

Carcinogenic potential 2 years, 2 species

Mutagenic potential Effects on genetic stability and

mutations in bacteria (Ames test) or

mammalian cells in culture, dominant

lethal test and clastogenicity in mice

Investigative Toxicology Determine sequence and mechanisms

of toxic action, etc

Toxicity Studies

Qualitative Observation

• Body Weight and Food Consumption

• Ophthalmology interval

• Hematology parameters

• Clinical Chemistry Parameters

• Urinalysis Parameters

• Organ Weight

• Microscopic Pathology

• Animal Responses

Clinical Signs of Toxicity

Autonomic Signs

• Etc

CRC Handbook of Toxicology,2005

Quantitative Observation

• Acute Toxicity

ED-50, LD-50, TI

• Sub Chronic and Chronic Toxicity

ADI, NOEL, NOAEL

CRC Handbook of Toxicology,2005

Acute Toxicity

- Acute toxicity dilakukan pertama kalinya

(biasanya oral dan IV)

- Menentuklan harga LD-50

- Binatang coba mati dlm waktu 7-14 hari

period after a single dose is tabulated.

- Tanda tanda intoksikasi, lethargy,

perubahan perilaku, studi biokimia

harus dilakukan

LD50

• Quantal responses dihitung bila data dari

populasi.

• Bila mortality berupa response, maka

dosis pada 50% dari populasi LD50

• LD 50 paling kecil paling toxic

• Therapeutic Index (TI) is the ratio of the

dose required to produce a toxic effect

to that required to produce a desired

therapeutic response(LD50/ED50)

LD50 berbagai bahan kimia Toxicant LD50 (mg/kg)

Ethyl alcohol 10,000

Salt (sodium chloride) 4,000

Iron (Ferrous sulfate) 1,500

Morphine 900

Mothballs (paradichlorobenzene) 500

Aspirin 250

DDT 250

Cyanide 10

Nicotine 1

Black Widow Spider venom 0.55

Rattle Snake venom 0.24

Tetrodotoxin (from fish) 0.01

Dioxin (TCDD) 0.001

Botulinum Toxin 0.00001

Subchronic toxicity tests

• Uji toksisitas selama 90 hari

• Dua species (rats dan dogs)

• 3 dosis level

• Tiap dosis minimum 15 binatang (jantan/betina)

• Pengamatan : Mortality, body weight, diet

consumption, hematology dan clinical chemistry.

• Pemeriksaan Gross dan microscopic dari tiap

organs dan jaringan.

Long term / chronic exposure

studies

• Dilakukan mirip dengan pengamatan pada studi

sub chronic, kecuali dengan periode lebih lama .

Mis, uji toksisitas Antimicrobial agents dan food

additives.

• Terutama penentuan carcinogenic potential

• Dilakukan pada tikus, mice, spesies lainnya

selama life spent (masa hidup) dari tiap spesies

Chronic Toxicity:

(repeated exposures)

x

Threshold concentrat ion

SYMPTOMS

TIME: Weeks, months, years

Blo

od

or

Tis

su

e

Co

nc

en

tra

tio

n

x

x

x

x

x

x

x

70

Dose levels (animal studies)

– NOEL no-observed effect level

– NOAEL no-observed-adverse effect level

– LOAEL lowest-observed-adverse effect level

– MTD maximum tolerated dose

– LD50 dose which kills 50% of population

– LC50 concentration which kills 50% of

population; must include time frame

Inc

rea

sin

g d

ose