2 IntroductionXenobiotics: is a compound that is foreign to the body ; is a chemical which is found in an organism but which is not normally produced or expected to be present in body.Endogenous: Pigments , hormonenonendogenous : Such as drugs , food additives, pollutants, toxin, etcMost of these compounds are subject to metabolism (biotransformation) in human body.
3 Definition of the biotransformation Conversion of lipophilic xenobiotics to water-soluble chemicals by a process catalyzed by enzymes in the liver and other tissues.In most cases, biotransformation lessens the toxicity of xenobiotics, but many must undergo the process to exert their toxic effects.卫生毒理学3
4 Purpose of biotransformation 1. facilitates excretion: Converts lipophilic to hydrophilic compounds2. Detoxification/inactivation: converts chemicals to less toxic forms3. Metabolic activation: converts chemicals to more toxic active forms
6 Sites of biotransformation LiverPrimary site! Rich in enzymesActs on endogenous and exogenous compoundsExtrahepatic metabolism sitesIntestinal wallSulfate conjugationEsterase and lipases - important in prodrug metabolismLungs, kidney, placenta, brain, skin, adrenal glands
7 Knowledge of the metabolism of xenobiotics: rational understanding of pharmacology and therapeutics , pharmacy, toxicology, cancer research , and drug addiction .
8 General Metabolic Pathways Approximately 30 different enzymes catalyze reactions involved in xenobiotic metabolism; however, this note will only cover a selected group of them.It is convenient to consider the metabolism of xenobiotics in two phasesphase Ⅰand phase Ⅱ
9 Phase I reactions ♣♣ Functionalization Purpose Oxidation Reduction Hydrolytic reactionsPurposeIntroduction of polar functional groups in a molecules♣ Increase a molecule’s polarity♣ Does provide a site for phase II metabolism
10 Phase II reactions ♣♣ Conjugation ★ Purpose Introduce highly polar conjugates:☻☻Glucuronic acid ☻☻ SulfateDetoxificationGlycine or other Amino Acids (some solubility), Acetyl , Methylations , Glutathione★ Site of attachment often introduced in Phase IHydroxyl , Carboxylate , Amino
12 RH + O2 + NADPH + H+ R-OH + H2O + NADP+ Phase Ⅰ: Oxidation1. HydroxylationRH + O2 + NADPH + H+ R-OH + H2O + NADP+Addition of an oxygen atom or bondRequire NADH or NADPH and O2 as cofactorsRH: drugs, cacinogens, pesticides, petroleum products, pollutants, steroids, eicosanoids, fatty acids, retinoids, etc.Enzyme:Cytochrome P450s-dependent monooxygenase
13 Hydroxylation：O2 * It has been shown by the use of O2 that one atom of oxygen enters R-OH and one atom enters H2O.* This dual fate of the oxygen accounts for the former naming of monooxygenases as “mixed-function oxidases”.RH + O2 + NADPH + H+ R-OH + H2O + NADP+
14 Cytochrome P450s-dependent monooxygenase -----the most versatile biocatalyst----Works on a large number of diverse compounds★ Microsomal drug oxidations requirecytochrome P450,cytochrome P450 reductase,NADPH , & O2The actual reaction mechanism is as follows:
16 Cytochrome P450s-dependent monooxygenase CYP or Cytochrome P-450★ Heme proteins★ Iron containing porphyrin - binds O2★ The name cytochrome P450 is derived from the spectral properties of this hemoprotein in its reduced (ferrous, Fe2+) form, it binds CO to give a complex that absorbs light maximally at 450 nm
17 Cytochrome P-450 Enzymes isolated by disruption of the liver cells Endoplasmic reticulum - microsomes when disruptedEnzymes are membrane boundExplains why lipophilic drugs are processedCatalytic process heme binds O2
18 Cytochrome P450: Isozymes ★ Isozymes - multiple forms of an enzyme★ Supergene family- More than 8,000 P450 genes as of November/2007- More than 368 gene families, 814 subfamilies- Human: 18 families, 43 subfamilies, 57 sequencedgenes★ NomenclatureCYP1A2familysubfamilyindividual member of that subfamily
19 Cytochrome P450This enzyme is very important. approximately 50% of the drugs that humans ingest are metabolized by isoforms of cytochrome P450.These enzymes also act on various carcinogens and pollutants.
21 2. Monoamine oxidase, MAO RCH2NH2+O2+H2O2 RCHO+NH3+H2O ★ MAO catalyze the oxidative deamination of monoamines.★ Oxygen is used to remove an amine group from a molecule, resulting in the corresponding aldehyde and ammonia.★ MAO are found bound to the outer membrane of mitochondria in most cell types in the body. They belong to protein family of flavin containing amine oxidoreductases.
22 3. ADH and ALDHADH alcohol dehydrogenaseALDH aldehyde dehydrogenaseAlcohol Dehydrogenase belongs to the oxidoreductase family of enzymes.high concentrations within the liver and kidney.
23 FunctionThe primary and most common role of ADH in humans is to detoxify incoming ethanol by converting it into aldehyde.The resulting aldehyde, a more toxic molecule than ethanol, is quickly converted into acetate by aldehyde dehydrogenase (ALDH) and other molecules easily utilized by the cell.ALDHADH
24 ADHALDHDuring this reaction, hydrogen is removed from the alcohol and transferred to a molecule called nicotinamide adenine dinucleotide (NAD), converting it to reduced NAD (NADH).NADH participates in numerous other metabolic reactions, passing on the hydrogen to other compounds or electron transfer chain.
26 Absorption Soluble in water stomach20%Soluble in waterSmall size - penetrates everywhere, easily crosses all bio membranesRapidly absorbed from GIsmallintestine80%Blooduntil metabolized
27 In people who consume alcohol at moderate levels and/or only occasionally, most of the alcohol is broken down by ADH and ALDH.after higher alcohol consumption, The MEOS plays a role in alcohol metabolism.CH3CH2OH + NADPH + O2 + H CH3CHO + NADP+ + 2H2OMEOSCH3CHOALDHCH3COOHMEOS: Microsomal Ethanol-Oxidizing System , is also called Cytochrome P450-dependent Microsomal Ethanol Oxidizing System. converts alcohol to acetaldehyde
28 CH3CH2OH + NADPH + O2 + H+ CH3CHO + NADP+ + 2H2O MEOSThis reaction also relies on oxygen and NADPH, and results in the formation of NADP and water.◆consume oxygens of liver and NADPH◆ As byproducts of these reactions, oxygen radicals or reactive oxygen species (ROS) are generated. These ROS can contribute to liver damage through a variety of mechanisms.
29 ADH and MEOS consume energy ADH MEOS In liver cytoplasma microsome substratealcohol、NAD+alcohol 、NADPH、O2Km to alcohol2mmol/L8.6mmol/LInduced by alcoholnoyesenergyRelease energyconsume energy
30 Although the rate at which ADH breaks down alcohol generally stays the same, the activity of the MEOS can be increased (induced) by alcohol consumption.
31 Because the MEOS metabolizes not only alcohol but also other compounds (certain medications), enhanced MEOS activity resulting from high alcohol consumption also can alter the metabolism of those medications.This may contribute to harmful interactions between alcohol and those medications or otherwise influence the activity of those medications.
32 Alcoholism leads to fat accumulation in the liver, hyperlipidemia, and ultimately cirrhosis.
33 4. Nitro and Azo Reduction Phase Ⅰ: Reduction4. Nitro and Azo ReductionNADPH dependent microsomal and nitro-reductase enzymes.Bacterial reductases play a role in enterohepatic recirculation of nitro or azo containing drugs.
36 5. Hydrolysis Substrates: esters , amide , glycoside, etc. Phase Ⅰ: Reduction5. HydrolysisSubstrates: esters , amide , glycoside, etc.Catalyzed by widely distributed hydrolytic enzymesHydrolysis of esters major metabolic pathway for ester drugs☻Non-specific esterases (liver, kidney, and intestine)☻Plasma pseudocholinesterases also participateesteraseAcetylsalicylic Acid, ASAsalicylic acid
37 Phase II: ConjugationIn phase Ⅰ reactions, xenobiotics are generally converted to more polar, hydroxylated derivatives.In phase Ⅱ reactions, these derivatives are conjugated with molecules such as glucuronic acid, sulfate, or glutathione.This renders them even more water-soluble, and they are eventually excreted in the urine or bile.
38 xenobiotic excrection nontoxic metabolite cancer Phase I Phase II ProtectionEliminationReactivemetaboliteCell injuryAntibody productmutationcancer
39 Five types of phase II reactions A. GlucuronidationB. SulfationC. Conjugation with glutathioneD. AcetylationE. Methylation
40 1. Glucuronidation the most frequent conjugation reaction. UDP-glucuronic acid (UDPGA) is the glucuronyl donorUDP-glucuronyl transferases (UGT), present in both the endoplasmic reticulum(ER) and cytosol, are the catalysts.Liver, lung, kidney, skin, brain and intestineAttachment sites are hydroxylsAlcohols, phenols, enols, N-hydroxyls, acidsOxygen site often from Phase I
43 2. Sulfate Conjugation Catalyzed by sulfotransferases Some alcohols, arylamines, and phenols are sulfated.Catalyzed by sulfotransferasesliver, kidney and intestineSulfate donor: adenosine 3’-phosphate-5’-phosphosulfate (PAPS); this compound is called “active sulfate.”Leads to inactive water-soluble metabolitesGlucuronate conjugation often more competitive processsulfotransferasePAPS
49 4. Acetylation X + Acetyl-CoA - - - - >Acetyl-X + CoS where X represents a xenobiotics.(for: aromatic amines)Enzyme: acetyltransferasespresent in the cytosol of various tissues, particularly in liver.isoniazid
50 Important for drugs with primary amino groups sulfanilamideImportant for drugs with primary amino groupsGenerally, metabolites are nontoxic and inactiveAcetylation does NOT increase water solubilityDetoxification or termination of drug activity
51 Methylation A few xenobiotics are subject to methylation by methyltransferase,emplyoing S-adenosylmethione(SAM) as the methyl donor.SAMcatechol
52 Metabolism via Methylation Key for biosynthesis of many compoundsImportant in the inactivation of physiologically active biogenic amines neurotransmittersnorepinephrine, dopamine, serotonin, histamineMinor pathway in the metabolism of drugsMethylation does NOT increase water solubilityMost methylated products are inactive
53 Factors that influence metabolism Ageolder people less efficient at metabolismSexLinked to hormonal differencesHeredityGenetic differences can influence amounts and efficiency of metabolic enzymesDisease statesLiver, cardiac, kidney disease
54 Summary Phase I reactions: Phase II: Conjugation Rx Xenobiotic, BiotransformationPhase I reactions:Purpose:Enhances eliminationConverts chemical to less toxic forms (detoxification)Converts chemicals to more toxic active forms (activation)Functionalization:Oxidation: monooxygenase, CYP450Reduction: ADH, ALDHHydrolytic reactions: esterasePhase II: Conjugation RxPurpose: more water-soluble, excreted in the urine or bileGlucuronidation, Sulfation, Conjugation with glutathione, Acetylation, Methylation
55 ALCOHOL Mechanism of Fatty Liver The likelihood of hypoglycemia is also increased in alcoholics when they fast, as they often have low hepatic stores of glycogen because of poor nutrition.The shift in the NADH/NAD+ ratio also inhibits β-oxidation of fatty acids and promotes triglyceride synthesis; this increases hepatic synthesis of VLDL, and the remaining excess triglyceride is deposited in the liver.