phase 2

• Phase II reactions


  Usually known as conjugation reactions (e.g., with glucuronic acid, sulfonates (commonly known as sulfation), glutathione or amino acids)
• Phase II reactions


  • Usually detoxication in nature
  • Involve the interactions of the polar functional groups of phase I metabolites
• Sites on drugs where conjugation reactions occur

  • Carboxyl (-COOH)
  • Hydroxyl (-OH)
  • Amino (NH2)
  • Sulfhydryl (-SH) groups
• Products of conjugation reactions

  Have increased molecular weight and are usually inactive unlike Phase I reactions which often produce active metabolites
• Conjugating Enzymes


  Large number of phase 2 conjugating enzymes, all of which are considered to be synthetic in nature since they result in the formation of a metabolite with an increased molecular mass
• Phase 2 reactions


  • Terminate the biological activity of the drug
  • Two of the phase 2 reactions, glucuronidation and sulfation, result in the formation of metabolites with a significantly increased water-to-lipid partition coefficient, resulting in hydrophilicity and facilitating their transport into the aqueous compartments of the cell and the body
  • Glucuronidation also markedly increases the molecular weight of the compound, a modification that favors biliary excretion
  • Sulfation and acetylation terminate the biological activity of drugs, the solubility properties of these metabolites are altered through minor changes in the overall charge of the molecule
• Cofactors for phase 2 reactions

  UDP-glucuronic acid (UDP-GA) and 3'-phosphoadenosine-5'-phosphosulfate (PAPS), for UDP-glucuronosyltransferases (UGT) and sulfotransferases (SULT), respectively
• Phase 2 reactions


  • Carried out in the cytosol of the cell, with the exception of glucuronidation, which is localized to the luminal side of the endoplasmic reticulum
  • Catalytic rates are significantly faster than the rates of the CYPs
• If a drug is targeted for phase 1 oxidation through the CYPs, followed by a phase 2 conjugation reaction

  The rate of elimination will depend upon the initial (phase 1) oxidation reaction
• Glucuronidation


  Catalyzed by UDP-glucuronosyltransferases (UGTs), which catalyze the transfer of glucuronic acid from the cofactor to a substrate to form β-D-glucopyranosiduronic acids (glucuronides)
• Substrates for glucuronidation


  • Alcoholic and phenolic hydroxyl groups
  • Carboxyl
  • Sulfuryl
  • Carbonyl moieties
  • Primary, secondary, and tertiary amine linkages
• UGT proteins


  • 19 human genes that encode the UGT proteins, 9 encoded by the UGT1 locus and 10 encoded by the UGT2 family of genes
  • UGT2 family of proteins appears to have greater specificity for the glucuronidation of endogenous substances such as steroids
  • UGT1 locus spans nearly 200 kb, with over 150 kb encoding a tandem array of cassette exonic regions that encode approximately 280 amino acids of the amino terminal portion of the UGT1A proteins
• The expression of UGT1A1 assumes an important role in drug metabolism since the glucuronidation of bilirubin by UGT1A1 is the rate-limiting step in assuring efficient bilirubin clearance, and this rate can be affected by both genetic variation and competing substrates (drugs)
• Sulfonation


  A common 'Phase II' conjugation reaction that occurs across species, involves the conjugation of the substrate with a sulfonyl (SO3-) group, catalyzed by a SULT enzyme with the cosubstrate PAPS
• Substrates for sulfonation


  • –C–OH
  • –N–OH
  • –NH side chains
• Sulfonation reactions


  • Follow a random or ordered bi–bi mechanism depending on the substrate and specific SULT isozyme studied, via SN2 displacement
  • Substrate inhibition is commonly observed at high substrate concentrations, probably due to the formation of dead-end complexes
• Cellular Location and Tissue Expression of SULTs

  • Cytosolic SULTs exist as homodimers or heterodimers in solution, responsible for conjugation of endogenous substrates and xenobiotics
  • Membrane bound SULTs are present in the Golgi apparatus, catalyze sulfonation and posttranslational modification of peptides, proteins, lipids, and glycosaminoglycans
  • Sulfonation activity is highest in the liver and small intestine, endogenous steroids are sulfonated in most mammalian tissues, with high activity in the adrenal tissue, kidney, and brain
• Methyldopa


  • Centrally acting antihypertensive agent, a prodrug that exerts its antihypertensive action via an active metabolite
• Mechanism of action of methyldopa

  Methyldopa is metabolized to α-methyldopamine, which is then converted to α-methylnorepinephrine. α-Methylnorepinephrine acts in the CNS to inhibit adrenergic neuronal outflow from the brainstem, attenuating NE release and reducing the output of vasoconstrictor adrenergic signals to the peripheral sympathetic nervous system
• Pharmacological effects of methyldopa

  • Reduces vascular resistance without causing much change in cardiac output or heart rate in younger patients, may decrease cardiac output in older patients due to decreased heart rate and stroke volume
  • The fall in arterial pressure is maximal 6 to 8 hours after an oral or intravenous dose
  • Renal blood flow is maintained and renal function is unchanged during treatment
  • Plasma concentrations of norepinephrine fall in association with the reduction in arterial pressure
  • Renin secretion also is reduced, but this is not a major effect and is not necessary for its hypotensive effects
  • Salt and water often are gradually retained with prolonged use, causing "pseudotolerance" that can be overcome with concurrent use of a diuretic