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١٨ ١٨ Extent of Absorption Extent of Absorption The extent of absorption is important in determining the total The extent of absorption is important in determining the total body exposure or internal dose, and therefore is an important body exposure or internal dose, and therefore is an important variable during chronic toxicity studies and/or chronic human variable during chronic toxicity studies and/or chronic human exposure. The extent of absorption depends on the extent to exposure. The extent of absorption depends on the extent to which the chemical is transferred from the site of which the chemical is transferred from the site of administration into the local tissue, and the extent to which it administration into the local tissue, and the extent to which it is metabolized or broken down by local tissues prior to is metabolized or broken down by local tissues prior to reaching the general circulation. An additional variable reaching the general circulation. An additional variable affecting the extent of absorption is the rate of removal from affecting the extent of absorption is the rate of removal from the site of administration by other processes compared with the the site of administration by other processes compared with the rate of absorption rate of absorption

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Extent of AbsorptionExtent of AbsorptionThe extent of absorption is important in determining the total The extent of absorption is important in determining the total

body exposure or internal dose, and therefore is an important body exposure or internal dose, and therefore is an important variable during chronic toxicity studies and/or chronic human variable during chronic toxicity studies and/or chronic human exposure. The extent of absorption depends on the extent to exposure. The extent of absorption depends on the extent to which the chemical is transferred from the site of which the chemical is transferred from the site of administration into the local tissue, and the extent to which itadministration into the local tissue, and the extent to which it is metabolized or broken down by local tissues prior to is metabolized or broken down by local tissues prior to reaching the general circulation. An additional variable reaching the general circulation. An additional variable affecting the extent of absorption is the rate of removal from affecting the extent of absorption is the rate of removal from the site of administration by other processes compared with the the site of administration by other processes compared with the rate of absorptionrate of absorption

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Chemicals given Chemicals given via via the gastrointestinal tract may be the gastrointestinal tract may be subject to a wide range of pH values and subject to a wide range of pH values and metabolizing enzymes in the gut lumen, gut wall, and metabolizing enzymes in the gut lumen, gut wall, and liver before they reach the general circulation. The liver before they reach the general circulation. The initial loss of chemical prior to it ever entering the initial loss of chemical prior to it ever entering the blood is termed blood is termed firstfirst--pass metabolism or prepass metabolism or pre-- systemic metabolismsystemic metabolism; it may in some cases remove ; it may in some cases remove up to 100% of the administered dose so that none of up to 100% of the administered dose so that none of the parent chemical reaches the general circulation. the parent chemical reaches the general circulation. The intestinal lumen contains a range of hydrolytic The intestinal lumen contains a range of hydrolytic enzymes involved in the digestion of nutrients. The enzymes involved in the digestion of nutrients. The gut wall can perform similar hydrolytic reactions and gut wall can perform similar hydrolytic reactions and contains enzymes that can oxidize many drugscontains enzymes that can oxidize many drugs

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Absorption and BioavailabilityAbsorption and BioavailabilityIrrespective of the reason that is responsible for the incompletIrrespective of the reason that is responsible for the incomplete e

absorption of the chemical as the parent compound, it is absorption of the chemical as the parent compound, it is essential that there is a parameter which defines the extent of essential that there is a parameter which defines the extent of transfer of the intact chemical from the site of administration transfer of the intact chemical from the site of administration into the general circulation. This parameter is the into the general circulation. This parameter is the bioavailabilitybioavailability, which is simply the fraction of the dose , which is simply the fraction of the dose administered that reaches the general circulation as the parent administered that reaches the general circulation as the parent compound. (The term bioavailability is perhaps the most compound. (The term bioavailability is perhaps the most misused of all kinetic parameters and is sometimes used misused of all kinetic parameters and is sometimes used incorrectly in a general sense as the amount of drug available incorrectly in a general sense as the amount of drug available specifically to the site of toxicity.)specifically to the site of toxicity.)

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Calculation of BioavailabilityCalculation of BioavailabilityThe fraction absorbed as the intact compound or bioavailability The fraction absorbed as the intact compound or bioavailability ((FF) is ) is

determined by comparison with intravenous (determined by comparison with intravenous (i.vi.v.) dosing (where .) dosing (where F =F = 1 by 1 by definition). The bioavailability can be determined from the areadefinition). The bioavailability can be determined from the area under the under the plasma concentrationplasma concentration––time curve (AUC) of the parent compound , or the time curve (AUC) of the parent compound , or the percentage dose excreted in urine as the parent compound, percentage dose excreted in urine as the parent compound, i.e. i.e. for an oral for an oral dose:dose:

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2. Distribution2. Distribution

Distribution is the Distribution is the reversible transferreversible transfer of the of the chemical between the general circulation and chemical between the general circulation and the tissues. Irreversible processes such as the tissues. Irreversible processes such as excretion, metabolism, or covalent binding are excretion, metabolism, or covalent binding are part of elimination and do not contribute to part of elimination and do not contribute to distribution parameters. The important distribution parameters. The important distribution parameters relate to the rate and distribution parameters relate to the rate and extent of distribution.extent of distribution.

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Rate of DistributionRate of DistributionThe rate at which a chemical may enter or leave a tissue may be The rate at which a chemical may enter or leave a tissue may be

limited by two factors:limited by two factors:

(i) the ability of the compound to cross cell membranes and(i) the ability of the compound to cross cell membranes and(ii) the blood flow to the tissues in which the chemical (ii) the blood flow to the tissues in which the chemical

accumulates.accumulates.

The rate of distribution of highly waterThe rate of distribution of highly water--soluble compounds may soluble compounds may be slow due to their slow transfer from plasma into body be slow due to their slow transfer from plasma into body tissues such as liver and muscle; watertissues such as liver and muscle; water--soluble compounds do soluble compounds do not accumulate in adipose tissue. In contrast, very lipidnot accumulate in adipose tissue. In contrast, very lipid--soluble soluble chemicals may rapidly cross cell membranes but the rate of chemicals may rapidly cross cell membranes but the rate of distribution may be slow because they accumulate in adipose distribution may be slow because they accumulate in adipose tissue, and their overall distribution rate may be limited by tissue, and their overall distribution rate may be limited by blood flow to adipose tissueblood flow to adipose tissue

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The rate of distribution is indicated by the distribution The rate of distribution is indicated by the distribution rate constant, which is determined from the decrease rate constant, which is determined from the decrease in plasma concentrations in early time points after an in plasma concentrations in early time points after an intravenous dose. The rate constants refer to a mean intravenous dose. The rate constants refer to a mean rate of removal from the circulation and may not rate of removal from the circulation and may not correlate with uptake into a specific tissue. Once an correlate with uptake into a specific tissue. Once an equilibrium has been reached between the general equilibrium has been reached between the general circulation and a tissue, any process which lowers the circulation and a tissue, any process which lowers the blood (plasma) concentration will cause a parallel blood (plasma) concentration will cause a parallel decrease in the tissue concentration.decrease in the tissue concentration.

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Factors affecting distributionFactors affecting distribution1.1. Blood flowBlood flowDrugs are readily distributed to highly Drugs are readily distributed to highly perfusedperfused tissue tissue

like brain, liver, and kidneyslike brain, liver, and kidneys2. Permeability limitations2. Permeability limitationsMany drugs do not readily enter the brain due to the Many drugs do not readily enter the brain due to the

blood brain barrierblood brain barrier3. Protein binding3. Protein bindingAcidic drugs are bound to the most abundant plasma Acidic drugs are bound to the most abundant plasma

protein (albumin); while basic drugs bind to protein (albumin); while basic drugs bind to --11-- acid glycoprotein.acid glycoprotein.

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4. Effect of pH4. Effect of pHThe pH of the blood or tissue affect the The pH of the blood or tissue affect the

ionization of the drug and thus its distributionionization of the drug and thus its distribution5. Age5. AgeIn old people, Protein binding and body water In old people, Protein binding and body water

will decrease, thus increasing the concentration will decrease, thus increasing the concentration of the drug per unit timeof the drug per unit time

6. Existence of storage sites:6. Existence of storage sites:These include: Adipose tissue, plasma proteins, These include: Adipose tissue, plasma proteins,

liver and kidneys, and boneliver and kidneys, and bone

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Extent of DistributionExtent of Distribution

The extent of tissue distribution of a chemical depends The extent of tissue distribution of a chemical depends on the relative affinity of the blood or plasma on the relative affinity of the blood or plasma compared with the tissues. Highly watercompared with the tissues. Highly water--soluble soluble compounds that are unable to cross cell membranes compounds that are unable to cross cell membranes readily are largely restricted to readily are largely restricted to extracellularextracellular fluid fluid (about 13 L per 70 kg body weight). Water(about 13 L per 70 kg body weight). Water--soluble soluble compounds capable of crossing cell membranes (compounds capable of crossing cell membranes (e.g. e.g. caffeine, ethanol) are largely present in total body caffeine, ethanol) are largely present in total body water (about 41 L per 70 kg body weight).water (about 41 L per 70 kg body weight).

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LipidLipid--soluble compounds frequently show extensive soluble compounds frequently show extensive uptake into tissues and may be present in the lipids of uptake into tissues and may be present in the lipids of cell membranes, adipose tissue. cell membranes, adipose tissue.

The partitioning between circulating lipoproteins and The partitioning between circulating lipoproteins and tissue constituents is complex and may result in tissue constituents is complex and may result in extremely low plasma concentrations. A factor which extremely low plasma concentrations. A factor which may further complicate the plasma/tissue partitioning may further complicate the plasma/tissue partitioning is that some chemicals bind reversibly to circulating is that some chemicals bind reversibly to circulating proteins such as albumin (for acid molecules) and proteins such as albumin (for acid molecules) and acid glycoprotein (for basic molecules).acid glycoprotein (for basic molecules).

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The extent and pattern of tissue distribution can The extent and pattern of tissue distribution can be investigated by direct measurement of be investigated by direct measurement of tissue concentrations in animals. Tissue tissue concentrations in animals. Tissue concentrations cannot be measured in human concentrations cannot be measured in human studies and, therefore, the extent of distribution studies and, therefore, the extent of distribution in humans has to be determined based solely in humans has to be determined based solely on the concentrations remaining in plasma or on the concentrations remaining in plasma or blood after distribution is complete.blood after distribution is complete.

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The parameter used to reflect the extent of distribution is the The parameter used to reflect the extent of distribution is the apparent volume of distribution (apparent volume of distribution (VV), which relates the total ), which relates the total amount of the chemical in the body (amount of the chemical in the body (AbAb) to the circulating ) to the circulating concentration (concentration (CC) at any time after distribution is complete:) at any time after distribution is complete:

The volumes of distribution of tubocurarine

and caffeine are about 13 and 41 L per 70 kg because of their restricted distribution

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CaffeineCaffeine tubocurarine

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when a chemical shows a more extensive reversible when a chemical shows a more extensive reversible uptake into one or more tissues the plasma uptake into one or more tissues the plasma concentration will be lowered and the value of concentration will be lowered and the value of V V will will increase. For highly lipidincrease. For highly lipid--soluble chemicals, such as soluble chemicals, such as organochlorineorganochlorine pesticides, which accumulate in pesticides, which accumulate in adipose tissue, the plasma concentration may be so adipose tissue, the plasma concentration may be so low that the value of low that the value of V V may be many may be many litreslitres for each for each kilogram of body weight. This is not a real volume of kilogram of body weight. This is not a real volume of plasma and therefore plasma and therefore V V is called the apparent volume is called the apparent volume of distribution.of distribution.

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It is an important parameter because extensive It is an important parameter because extensive reversible distribution into tissues, which will reversible distribution into tissues, which will give a high value of give a high value of VV, is associated with a low , is associated with a low elimination rate and a long halfelimination rate and a long half--life . It must be life . It must be emphasized that the apparent volume of emphasized that the apparent volume of distribution simply reflects the extent to which distribution simply reflects the extent to which the chemical has moved out of the site of the chemical has moved out of the site of measurement (the general circulation) into measurement (the general circulation) into tissues, and it does not reflect uptake into any tissues, and it does not reflect uptake into any specific specific tissue(stissue(s).).

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EliminationElimination

The parameter most commonly used to describe The parameter most commonly used to describe the rate of elimination of a chemical is the the rate of elimination of a chemical is the halfhalf--life . Most life . Most toxicokinetictoxicokinetic processes are processes are firstfirst--order reactions, order reactions, i.e. i.e. the rate at which the the rate at which the process occurs is proportional to the amount of process occurs is proportional to the amount of chemical present. High rates (expressed as chemical present. High rates (expressed as mass/time) occur at high concentrations and mass/time) occur at high concentrations and the rate decreases as the concentration the rate decreases as the concentration decreases; in consequence the decrease is an decreases; in consequence the decrease is an exponential curve.exponential curve.

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The usual way to analyze exponential changes is to use The usual way to analyze exponential changes is to use logarithmically transformed data which converts an logarithmically transformed data which converts an exponential into a straight line. The slope of the line exponential into a straight line. The slope of the line is the rate constant (is the rate constant (kk) for the process and the half) for the process and the half--life life for the process is calculated as 0.693/for the process is calculated as 0.693/kk. Rate . Rate constants and halfconstants and half--lives can be determined for lives can be determined for absorption, distribution, and elimination processes.absorption, distribution, and elimination processes.

The The clearanceclearance of a chemical is determined by the of a chemical is determined by the ability of the organs of elimination (ability of the organs of elimination (e.g. e.g. the liver, the liver, kidney, or lungs) to extract the chemical from the kidney, or lungs) to extract the chemical from the plasma or blood and permanently remove it by plasma or blood and permanently remove it by metabolism or excretion.metabolism or excretion.

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The mechanisms of elimination depend on the The mechanisms of elimination depend on the chemical characteristics of the compound:chemical characteristics of the compound:

volatile chemicals are exhaled,volatile chemicals are exhaled,

waterwater--soluble chemicals are eliminated in the soluble chemicals are eliminated in the urine and/or bile andurine and/or bile and

lipidlipid--soluble chemicals are eliminated by soluble chemicals are eliminated by metabolism to more watermetabolism to more water--soluble molecules, soluble molecules, which are then eliminated in the urine and/or which are then eliminated in the urine and/or bile.bile.

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If a chemical undergoes If a chemical undergoes metabolic activationmetabolic activation then then toxicokinetictoxicokinetic studies should measure both the parent studies should measure both the parent chemical and the active metabolite. If the metabolite chemical and the active metabolite. If the metabolite is so reactive that it does not leave the tissue in which is so reactive that it does not leave the tissue in which it is produced (it is produced (e.g. e.g. alkylatingalkylating metabolites of chemical metabolites of chemical carcinogens), then carcinogens), then toxicokinetictoxicokinetic studies should define studies should define the delivery of the parent chemical to the tissues, and the delivery of the parent chemical to the tissues, and the process of local activation should be regarded as the process of local activation should be regarded as part of tissue sensitivity (part of tissue sensitivity (toxicodynamicstoxicodynamics) because it ) because it is not part of is not part of toxicokineticstoxicokinetics, , i.e. i.e. the movement of the the movement of the chemical and/or metabolites around the body.chemical and/or metabolites around the body.

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Clearance = a ratio relating the Clearance = a ratio relating the raterate of of elimination of a chemical from an appropriate elimination of a chemical from an appropriate reference fluid (usually plasma) reference fluid (usually plasma) to its to its concentrationconcentration in the same reference fluid. in the same reference fluid. Clearance has the units of flow rate in Clearance has the units of flow rate in milliliters per minutes (milliliters per minutes (mLmL/min)./min).

A clearance of 100 A clearance of 100 mLmL/minute of a chemical /minute of a chemical means that 100 means that 100 mLmL of blood/plasma is of blood/plasma is completely cleared of the compound in each completely cleared of the compound in each minute.minute.

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The best measure of the ability of the organs of The best measure of the ability of the organs of elimination to remove the compound from the elimination to remove the compound from the body is the clearance (body is the clearance (CLCL):):

Because the rate of elimination is proportional to the concentration, clearance is a constant for first-order processes and is independent of dose. It can be regarded as the volume of plasma (or blood) cleared of compound within a unit of time (e.g. mL/ min).

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Renal clearance depends on the extent of protein Renal clearance depends on the extent of protein binding, tubular secretion and passive binding, tubular secretion and passive reabsorptionreabsorption in in the renal tubule; it can be measured directly from the the renal tubule; it can be measured directly from the concentrations present in plasma and urine:concentrations present in plasma and urine:

The total clearance or plasma clearance (which is the The total clearance or plasma clearance (which is the sum of all elimination processes, sum of all elimination processes, i.e. i.e. renal metabolic, renal metabolic, etc.etc.) is possibly the most important ) is possibly the most important toxicokinetictoxicokinetic parameter.parameter.

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It is measured from the total amount of compound available for It is measured from the total amount of compound available for removal (removal (i.e. i.e. an intravenous dose) and the total area under the an intravenous dose) and the total area under the plasma concentrationplasma concentration––time curve (AUC) extrapolated to time curve (AUC) extrapolated to infinity.infinity.

Plasma clearance reflects the overall ability of the body to remove permanently the chemical from the plasma. Plasma clearance is the parameter that is altered by factors such as enzyme induction, liver disease, kidney disease, inter-individual or inter-species differences in hepatic enzymes or in some cases organ blood flow.

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Once the chemical is in the general circulation, the same volumeOnce the chemical is in the general circulation, the same volume of plasma will be cleared of chemical per minute (of plasma will be cleared of chemical per minute (i.e. i.e. the the clearance value) applies irrespective of the route of delivery oclearance value) applies irrespective of the route of delivery of f chemical into the circulation. However, the bioavailability (chemical into the circulation. However, the bioavailability (FF) ) will determine the proportion of the dose reaching the general will determine the proportion of the dose reaching the general circulation. Therefore, bioavailability has to be taken into circulation. Therefore, bioavailability has to be taken into account if clearance is calculated from data from a nonaccount if clearance is calculated from data from a non-- intravenous route (intravenous route (e.g. e.g. oral):oral):

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Measurement of dose/AUC for an oral dose determines Measurement of dose/AUC for an oral dose determines CL/FCL/F, which contains , which contains two potentially independent variables two potentially independent variables ––

the amount of chemical delivered the amount of chemical delivered

to the blood from the site of administration and the clearance oto the blood from the site of administration and the clearance of chemical f chemical present in the blood. The overall rate of elimination, as indicapresent in the blood. The overall rate of elimination, as indicated by the ted by the terminal halfterminal half--life (life (t t ), is dependent on two physiologically related and ), is dependent on two physiologically related and independent variables:independent variables:

where where CL CL is the ability to extract and remove irreversibly the compound fis the ability to extract and remove irreversibly the compound from rom the general circulation, and the general circulation, and V V the extent to which the compound has left the the extent to which the compound has left the general circulation in a reversible equilibrium with tissues.general circulation in a reversible equilibrium with tissues.

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Chemicals that are extremely lipidChemicals that are extremely lipid--soluble and soluble and are sequestered in adipose tissue are are sequestered in adipose tissue are eliminated slowly. Lipid soluble eliminated slowly. Lipid soluble organochlorineorganochlorine compounds, which are not compounds, which are not substrates for P450 oxidation, due to the substrates for P450 oxidation, due to the blocking of possible sites of oxidation by blocking of possible sites of oxidation by chlorochloro--substituentssubstituents, are eliminated extremely , are eliminated extremely slowly: for example, the halfslowly: for example, the half--life of 2,3,7,8life of 2,3,7,8-- tetrachlorodibenzodioxin (TCDD) is about 8 tetrachlorodibenzodioxin (TCDD) is about 8 years in humans.years in humans.

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Summary of Summary of ToxicokineticToxicokinetic ParametersParameters

Apparent volume of distribution (Apparent volume of distribution (VVdd ))

A way to express the apparent space in the body that a chemical A way to express the apparent space in the body that a chemical occupiesoccupies

Expressed as a proportionality constant (in units of volume [e.gExpressed as a proportionality constant (in units of volume [e.g., ., liters of blood] or volume/body weight)liters of blood] or volume/body weight)

Relates the total amount of chemical in the body to the Relates the total amount of chemical in the body to the concentration in plasmaconcentration in plasma

For a 70For a 70--kg human,kg human,

Plasma volume Plasma volume ≈≈

3 L (~0.045 L/kg body weight)3 L (~0.045 L/kg body weight)

Total blood volume Total blood volume ≈≈

5 L (~0.07 L/kg body weight)5 L (~0.07 L/kg body weight)

Total Total extracellularextracellular fluid volume fluid volume ≈≈

12 L (~0.2 L/kg body weight)12 L (~0.2 L/kg body weight)

Total body water Total body water ≈≈

42 L (~0.6L/kg body weight)42 L (~0.6L/kg body weight)

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Example Example ––

If you know that 3 mg of chemical has been If you know that 3 mg of chemical has been injected into the body, and the concentration of the injected into the body, and the concentration of the chemical in the plasma is 1 mg/L, then you can calculate chemical in the plasma is 1 mg/L, then you can calculate that the apparent that the apparent VVdd to be 3 L. This suggests that the to be 3 L. This suggests that the chemical is confined to the plasma space, and is not chemical is confined to the plasma space, and is not significantly absorbed into the tissues or into the significantly absorbed into the tissues or into the particulate components (i.e., blood cells) in blood.particulate components (i.e., blood cells) in blood.

3 mg 3 mg ÷÷

1.0 mg/L = 3.0 L1.0 mg/L = 3.0 L

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Example Example ––

If you know that 3 mg of chemical has been If you know that 3 mg of chemical has been injected and the concentration of the chemical in the injected and the concentration of the chemical in the plasma is 0.55 mg/L, then the apparent plasma is 0.55 mg/L, then the apparent VVdd = 5.5 L. This = 5.5 L. This suggests that the chemical may be distributed evenly suggests that the chemical may be distributed evenly between the plasma and the blood cells. More likely, between the plasma and the blood cells. More likely, the reality is that the chemical is somewhat absorbed the reality is that the chemical is somewhat absorbed into the tissues.into the tissues.

3 mg 3 mg ÷÷

0.55 mg/L = 5.5 L0.55 mg/L = 5.5 L

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Example Example ––

If you know that 3 mg of chemical has been If you know that 3 mg of chemical has been injected and the concentration of the chemical in the injected and the concentration of the chemical in the plasma is 0.25 mg/L, then the apparent plasma is 0.25 mg/L, then the apparent VVdd = 12 L. This = 12 L. This suggests that the chemical may be distributed evenly suggests that the chemical may be distributed evenly within the total within the total extracellularextracellular water. More likely, the water. More likely, the reality is that the chemical is somewhat absorbed into reality is that the chemical is somewhat absorbed into the tissues, and perhaps somewhat metabolized.the tissues, and perhaps somewhat metabolized.

3 mg 3 mg ÷÷

0.25 mg/L = 12 L0.25 mg/L = 12 L

It is possible to have a It is possible to have a VVdd greater than the total body greater than the total body waterwater

A high A high VVdd is an indication that one or both of two things is an indication that one or both of two things has occurred,has occurred,

The chemical has been absorbed from the blood and The chemical has been absorbed from the blood and concentrated in the tissues.concentrated in the tissues.

The chemical has been metabolized.The chemical has been metabolized.

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Clearance (Clearance (ClCl))

A ratio relating the A ratio relating the raterate of elimination of a chemical of elimination of a chemical from an appropriate reference fluid (usually plasma) from an appropriate reference fluid (usually plasma) to to its concentrationits concentration in the same reference fluid. in the same reference fluid.

Has the units of flow rate in volume cleared per unit Has the units of flow rate in volume cleared per unit time (e.g., time (e.g., mLmL/min)/min)

A clearance of 100 A clearance of 100 mLmL/minute of a chemical means that /minute of a chemical means that 100 100 mLmL of blood/plasma is completely cleared of the of blood/plasma is completely cleared of the compound in each minute .compound in each minute .

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ZeroZero-- versus firstversus first--order kineticsorder kinetics

For zero order kinetics, the rate of elimination of a compound iFor zero order kinetics, the rate of elimination of a compound is a s a constant, and is independent of the concentration of the chemicaconstant, and is independent of the concentration of the chemical in l in the blood. For example, the average human body is able to the blood. For example, the average human body is able to eliminate ~10eliminate ~10--15 15 mLmL of ethanol per hour, regardless of the amount of ethanol per hour, regardless of the amount of ethanol consumed. This may be higher or lower depending on of ethanol consumed. This may be higher or lower depending on the factors previously discussed (e.g., induction of alcohol the factors previously discussed (e.g., induction of alcohol dehydrogenasedehydrogenase by repeated alcohol exposure).by repeated alcohol exposure).

For firstFor first--order kinetics, the rate of elimination of a compound is order kinetics, the rate of elimination of a compound is dependent on the concentration of the chemical in the blood. Thdependent on the concentration of the chemical in the blood. The e higher the concentration, the more rapidly the chemical is higher the concentration, the more rapidly the chemical is eliminated, unless the elimination mechanisms have been saturateeliminated, unless the elimination mechanisms have been saturated. d. At that point, the kinetics become zeroAt that point, the kinetics become zero--order. This is known as order. This is known as saturationsaturation kinetics.kinetics.

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HalfHalf--life (tlife (t1/21/2 ))

The time required for the concentration of a chemical in The time required for the concentration of a chemical in the plasma to decrease by 50%.the plasma to decrease by 50%.

This is a constant for all but zeroThis is a constant for all but zero--order kinetics. For order kinetics. For example, for a compound eliminated by firstexample, for a compound eliminated by first--order order kinetics, if the concentration at time 0 is 4 mg/L, and the kinetics, if the concentration at time 0 is 4 mg/L, and the tt1/21/2 is 6 hours, then at the end of 6 hours, the plasma is 6 hours, then at the end of 6 hours, the plasma concentration will be (0.5 x 4 mg/concentration will be (0.5 x 4 mg/mLmL =) 2 mg/L, and at =) 2 mg/L, and at the end of the next 6 hours, the concentration will be the end of the next 6 hours, the concentration will be (0.5 x 2 mg/(0.5 x 2 mg/mLmL =) 1 mg/L, and so on.=) 1 mg/L, and so on.

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Area under the Curve (AUC)Area under the Curve (AUC)

A measure of the total amount of chemical present in A measure of the total amount of chemical present in the body over a defined period. This is defined as the the body over a defined period. This is defined as the area under the concentration area under the concentration vsvs time curve for the time curve for the defined period.defined period.

Example: The shaded area in the figure below Example: The shaded area in the figure below represents the AUC(0represents the AUC(0--6hr). You can express the AUC 6hr). You can express the AUC for any given time period. It is often expressed as the for any given time period. It is often expressed as the AUC(0AUC(0--∞∞).).

Concentrationin

plasma

Time (hrs) 60

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Classical Classical toxicokineticstoxicokinetics

One compartment modelOne compartment model

The elimination of a chemical is said to follow a oneThe elimination of a chemical is said to follow a one-- compartment model when compartment model when the elimination phase of the the elimination phase of the log concentration log concentration vsvs time curve is a straight linetime curve is a straight line. . Conceptually, this is as though the chemical were Conceptually, this is as though the chemical were evenly distributed throughout a single body evenly distributed throughout a single body compartment (e.g., total body water), and is eliminated compartment (e.g., total body water), and is eliminated at a constant rate over time. That is, a constant at a constant rate over time. That is, a constant percentagepercentage of the chemical present is eliminated over of the chemical present is eliminated over any given time period.any given time period.

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The rate at which a chemical is eliminated at any time is directThe rate at which a chemical is eliminated at any time is directly ly proportional to the amount of that chemical in the body at that proportional to the amount of that chemical in the body at that time.time.

A oneA one--compartment model indicates that no one tissue has a high affinicompartment model indicates that no one tissue has a high affinity ty for the chemical. Changes in the plasma reflect changes in the for the chemical. Changes in the plasma reflect changes in the tissuetissue..

Plasma Concentratio

n

Time

Log of Plasma

Concentratio n

Time

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Two compartment modelTwo compartment model

The elimination of a chemical is said to follow a twoThe elimination of a chemical is said to follow a two-- compartment model when compartment model when the elimination phase of the the elimination phase of the log concentration log concentration vsvs time curve is not a straight linetime curve is not a straight line. . Conceptually, this is as though the chemical were Conceptually, this is as though the chemical were unevenly distributed throughout two body unevenly distributed throughout two body compartments (e.g., highly concentrated in tissues and a compartments (e.g., highly concentrated in tissues and a lower concentration in the plasma). The net elimination lower concentration in the plasma). The net elimination is a sum of the rates of elimination from the various is a sum of the rates of elimination from the various compartments. compartments.

The appearance of a twoThe appearance of a two--compartment curve suggests compartment curve suggests that the chemical is distributed out of the blood into that the chemical is distributed out of the blood into tissues.tissues.

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Time

Log of Plasma Concentration

Alpha distribution

Beta distribution

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Chronic AdministrationChronic Administration

The kinetic concepts and parameters of a single dose (as The kinetic concepts and parameters of a single dose (as discussed above) apply to chronic administration, but discussed above) apply to chronic administration, but the exposure has to allow for the fact that not all of the exposure has to allow for the fact that not all of the previous the previous dose(sdose(s) may have been eliminated when ) may have been eliminated when the subsequent dose is given. Therefore, there may be the subsequent dose is given. Therefore, there may be an increase in plasma concentration (and body load) an increase in plasma concentration (and body load) until an equilibrium is reached in which the rate of until an equilibrium is reached in which the rate of elimination balances out the rate of input, in other elimination balances out the rate of input, in other words, the daily dose is eliminated each day. words, the daily dose is eliminated each day.

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The equations above assume that The equations above assume that CL CL is not is not altered by repeated exposure; the assumption is altered by repeated exposure; the assumption is not correct if the chemical induces or inhibits not correct if the chemical induces or inhibits its own elimination because clearance would its own elimination because clearance would be increased or decreased, respectively, after be increased or decreased, respectively, after the period of chronic intake.the period of chronic intake.

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Because the plasma and therefore tissue concentrations Because the plasma and therefore tissue concentrations increase during chronic intake until an equilibrium is increase during chronic intake until an equilibrium is reached , the amount in the body (reached , the amount in the body (AbAb) will also ) will also increase to reach a steady state. The time taken to increase to reach a steady state. The time taken to reach steady state is 4reach steady state is 4––5 times the elimination half5 times the elimination half-- life and, therefore, the life and, therefore, the true duration oftrue duration of steadysteady--state state exposureexposure in a toxicity study is the study duration in a toxicity study is the study duration minus 4minus 4––5 half5 half--lives of the chemical (needed to reach lives of the chemical (needed to reach the steady state). This is particularly important for the steady state). This is particularly important for chemicals that have a very long halfchemicals that have a very long half--life; for example life; for example in rodents the steadyin rodents the steady--state body load of TCDD, which state body load of TCDD, which has a halfhas a half--life in rats of about 1 month, will not be life in rats of about 1 month, will not be reached until after about 4reached until after about 4––5 months of continuous 5 months of continuous treatment.treatment.

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Saturation KineticsSaturation Kinetics

All the parameters described above relate to firstAll the parameters described above relate to first--order order processes and, therefore, are independent of dose at processes and, therefore, are independent of dose at low doses. However, at high doses and/or during low doses. However, at high doses and/or during chronic studies it is possible to overload or saturate chronic studies it is possible to overload or saturate compoundcompound––protein interactions. Under such protein interactions. Under such circumstances any increase in the concentration of the circumstances any increase in the concentration of the compound cannot give a proportional (firstcompound cannot give a proportional (first--order) order) increase in the rate of the process. When a process is increase in the rate of the process. When a process is saturated the rate is at the maximum possible and is saturated the rate is at the maximum possible and is essentially independent of concentration.essentially independent of concentration.

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In simple mathematical terms this means that the reaction In simple mathematical terms this means that the reaction changes from first to zero order. This is best described by changes from first to zero order. This is best described by MichaelisMichaelis––MentenMenten kinetics, kinetics, i.e.i.e.

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3. Metabolism3. Metabolism

Many Many xenobioticsxenobiotics undergo chemical transformation undergo chemical transformation (biotransformation; metabolism) when introduced (biotransformation; metabolism) when introduced into biologic systems like the human body.into biologic systems like the human body.

Biotransformation is often mediated by enzymesBiotransformation is often mediated by enzymes

End result of biotransformation is either alteration of End result of biotransformation is either alteration of the parent molecule, or conjugation of the parent the parent molecule, or conjugation of the parent molecule (or its metabolites) with endogenous molecule (or its metabolites) with endogenous substances in the body. substances in the body.

Enzymes involved in biotransformation can act on Enzymes involved in biotransformation can act on either either endogenousendogenous or or xenobioticxenobiotic compounds, compounds, especially if the especially if the xenobioticsxenobiotics are structurally similar to are structurally similar to endogenous compoundsendogenous compounds

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Example: Example: CholinesteraseCholinesterase is an enzyme that normally metabolizes is an enzyme that normally metabolizes acetylcholine, a neurotransmitter found in the synapse. acetylcholine, a neurotransmitter found in the synapse. Cholinesterase can also metabolize the local anesthetic agent Cholinesterase can also metabolize the local anesthetic agent procaine and the muscleprocaine and the muscle--paralyzing agent paralyzing agent succinylcholinesuccinylcholine. If a . If a person has high levels of cholinesterase activity, the effectiveperson has high levels of cholinesterase activity, the effectiveness ness of these drugs can be compromised. If a person is exposed to of these drugs can be compromised. If a person is exposed to anything that inhibits cholinesterase activity, the pharmacologianything that inhibits cholinesterase activity, the pharmacologic c effects of the drugs can be enhanced, resulting in toxicity.effects of the drugs can be enhanced, resulting in toxicity.

Example: Example: Monoamine Monoamine oxidaseoxidase (MAO) is an enzyme that normally (MAO) is an enzyme that normally metabolizes biologic amines like epinephrine. MAO can also metabolizes biologic amines like epinephrine. MAO can also oxidize a variety of drugs. If a person is taking a drug that ioxidize a variety of drugs. If a person is taking a drug that inhibits nhibits MAO activity (like many blood pressure medications), it can be MAO activity (like many blood pressure medications), it can be dangerous for that person to take other drugs that can be dangerous for that person to take other drugs that can be metabolized by MAO.metabolized by MAO.

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The products of biotransformation can be either less toxic The products of biotransformation can be either less toxic than, more toxic than, or about as toxic as the parent than, more toxic than, or about as toxic as the parent molecules. molecules.

Enzymes involved in biotransformation are sometimes called Enzymes involved in biotransformation are sometimes called ““drug metabolizing enzymesdrug metabolizing enzymes””. Although strictly speaking this . Although strictly speaking this is a misnomer because many of the substrates are not drugs, is a misnomer because many of the substrates are not drugs, the term is still commonly used.the term is still commonly used.

Location of metabolic enzymesLocation of metabolic enzymes

Species Species ––

found in virtually every species, although the type and found in virtually every species, although the type and amount vary tremendously.amount vary tremendously.

Organs Organs ––

present in many tissues. Many enzymes are particularly present in many tissues. Many enzymes are particularly abundant in the liver. abundant in the liver.

SubcellularSubcellular ––

many of the drugmany of the drug--metabolizing enzymes are located in metabolizing enzymes are located in the smooth endoplasmic reticulum (SER). These are called the smooth endoplasmic reticulum (SER). These are called ““microsomalmicrosomal””

enzymes.enzymes.

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Types of Biotransformation ReactionsTypes of Biotransformation Reactions

Basically, two types of reactions, Basically, two types of reactions, nonsyntheticnonsynthetic (Phase I) and (Phase I) and synthetic (Phase II)synthetic (Phase II)

Phase I reactionsPhase I reactions

Involve modification of the basic structure of the substrateInvolve modification of the basic structure of the substrate

Do not involve covalent binding of the substrate to an endogenouDo not involve covalent binding of the substrate to an endogenous s compoundcompound

Examples include hydrolysis, oxidation, and reduction reactionsExamples include hydrolysis, oxidation, and reduction reactions

Phase I enzymes are often membranePhase I enzymes are often membrane--bound (e.g., bound (e.g., microsomalmicrosomal). This is because they generally act on more lipid). This is because they generally act on more lipid-- soluble (soluble (nonpolarnonpolar) substrates, and their purpose is to make the ) substrates, and their purpose is to make the compounds MORE POLAR and therefore, MORE EASILY compounds MORE POLAR and therefore, MORE EASILY EXCRETABLE by the kidney and EXCRETABLE by the kidney and biliarybiliary tract. Think of the tract. Think of the active transport mechanisms in the kidney. active transport mechanisms in the kidney.

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OxidationOxidation

Uses molecular oxygen (O2). One atom of oxygen is Uses molecular oxygen (O2). One atom of oxygen is combined with hydrogen to form water, and the other combined with hydrogen to form water, and the other atom of oxygen is introduced into the substrate atom of oxygen is introduced into the substrate molecule.molecule.

Involves several enzymatic steps.Involves several enzymatic steps.

The oxidative system is often known as the The oxidative system is often known as the ““mixed mixed function function oxidaseoxidase””

systemsystem””. These enzymes are some of . These enzymes are some of

the most thoroughly researched enzymes in biological the most thoroughly researched enzymes in biological systems.systems.

One subfamily of the mixed function One subfamily of the mixed function oxidaseoxidase system is the system is the group of enzymes known as group of enzymes known as CytochromeCytochrome PP--450450 enzymes. enzymes. They are so called because of their absorbance characteristics They are so called because of their absorbance characteristics at wavelengths of 448at wavelengths of 448--450 nm. 450 nm.

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Anything that affects the activity of any one of the steps can aAnything that affects the activity of any one of the steps can affect ffect the way the body reacts to a given drug or other the way the body reacts to a given drug or other xenobioticxenobiotic. .

Examples of the various types of oxidation reactions are in the Examples of the various types of oxidation reactions are in the textbook,textbook,

DeaminationDeamination ––

replacement of an amine group (NHreplacement of an amine group (NH22 ) with an oxygen ) with an oxygen (O) atom(O) atom

NN--, O, O--, or S, or S--DealkylationDealkylation ––

replacement of an alkyl group (e.g., CHreplacement of an alkyl group (e.g., CH33 ) ) with a hydrogen atom. Typically, the alkyl group in the parent with a hydrogen atom. Typically, the alkyl group in the parent molecule is bonded to a N, O, or S atom.molecule is bonded to a N, O, or S atom.

Aliphatic or aromatic hydroxylation Aliphatic or aromatic hydroxylation ––

addition of a hydroxyl group addition of a hydroxyl group (OH) to a molecule(OH) to a molecule

NN--oxidation oxidation ––

replacement of a hydrogen atom on an amine with an replacement of a hydrogen atom on an amine with an oxygenoxygen

SS--oxidation oxidation ––

addition of an oxygen atom to a sulfur atomaddition of an oxygen atom to a sulfur atom

Conversion of a hydroxyl group (alcohol) to a carboxyl group (acConversion of a hydroxyl group (alcohol) to a carboxyl group (acid)id)

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ReductionReduction

AzoAzo reduction reduction ––

reductionreduction of an of an azoazo bond (N=N) to two bond (N=N) to two amines (NHamines (NH22 ))

Nitro reduction Nitro reduction ––

reductionreduction of a nitro group (NOof a nitro group (NO22 ) to an ) to an amineamine

HydrolysisHydrolysis

Addition of water (HAddition of water (H22 O) to an ester bond (CO) to an ester bond (C--OO--OO--C) to C) to form an alcohol (Cform an alcohol (C--OH) and a carboxylic acid (COOH)OH) and a carboxylic acid (COOH)

RR--CC--OO--OO--CC--R + HR + H--OO--H H == RR--CC--OH + ROH + R--COOHCOOH

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Phase II reactionsPhase II reactions

Involve addition of a cofactor to a substrate to form a new Involve addition of a cofactor to a substrate to form a new product. Therefore, the rate of these reactions can be product. Therefore, the rate of these reactions can be limited by the availability of the cofactor.limited by the availability of the cofactor.

Phase II enzymes may be either Phase II enzymes may be either microsomalmicrosomal or or cytosoliccytosolic. . This is because the primary purpose of the Phase II This is because the primary purpose of the Phase II reactions is not so much to increase the polarity of the reactions is not so much to increase the polarity of the parent compound (although that is part of what they parent compound (although that is part of what they accomplish). The primary purpose is to increase the accomplish). The primary purpose is to increase the molecular weight of the parent compound to make it a molecular weight of the parent compound to make it a better substrate for active transport mechanisms in the better substrate for active transport mechanisms in the biliarybiliary tract.tract.

Various factors can affect the availability of cofactors. For Various factors can affect the availability of cofactors. For example, fasting markedly reduces the amount of example, fasting markedly reduces the amount of glutathione available in the liver.glutathione available in the liver.

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SulfationSulfation

Replacement of a hydrogen atom (H) with a Replacement of a hydrogen atom (H) with a sulfonatesulfonate (SO3)(SO3)

Uses the enzyme Uses the enzyme sulfotransferasesulfotransferase

Uses the cofactor called PAPS (Uses the cofactor called PAPS (phosphoadenosinephosphoadenosine phosphosulfatephosphosulfate))

Produces a highly waterProduces a highly water--soluble sulfuric acid estersoluble sulfuric acid ester

GlucuronidationGlucuronidation

Replacement of a hydrogen atom with a Replacement of a hydrogen atom with a glucuronicglucuronic acidacid

Uses the enzyme UDPUses the enzyme UDP--glucuronosylglucuronosyl transferasetransferase (UDP(UDP--GT)GT)

Uses the cofactor called UDPGA (Uses the cofactor called UDPGA (uridineuridine diphosphatediphosphate glucuronicglucuronic acid)acid)

One of the major Phase II enzymatic pathwaysOne of the major Phase II enzymatic pathways

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AcetylationAcetylation

Replacement of a hydrogen atom with an acetyl groupReplacement of a hydrogen atom with an acetyl group

Uses the enzyme Uses the enzyme acetyltransferaseacetyltransferase

Uses the cofactor called acetyl Uses the cofactor called acetyl CoACoA (acetyl coenzyme A)(acetyl coenzyme A)

Sometimes results in a less waterSometimes results in a less water--soluble productsoluble product

Remember the discussion of slow versus fast Remember the discussion of slow versus fast acetylatorsacetylators..

MethylationMethylation

Replacement of a hydrogen atom with a methyl groupReplacement of a hydrogen atom with a methyl group

Uses the enzyme Uses the enzyme methyltransferasemethyltransferase

Uses the cofactor called SAM (SUses the cofactor called SAM (S--adenosyladenosyl methioninemethionine))

Common but relatively minor pathwayCommon but relatively minor pathway

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Glutathione conjugationGlutathione conjugation

Adds a glutathione molecule to the parent compound, either by Adds a glutathione molecule to the parent compound, either by direct addition or by replacement of an direct addition or by replacement of an electrophilicelectrophilic substituentsubstituent (e.g., a halogen atom)(e.g., a halogen atom)

Uses the enzyme glutathione Uses the enzyme glutathione transferasetransferase (GST)(GST)

Uses the cofactor called glutathione (a Uses the cofactor called glutathione (a tripeptidetripeptide made up of made up of glycineglycine, , cysteinecysteine, and , and glutamicglutamic acidacid

One of the major Phase II enzymatic pathwaysOne of the major Phase II enzymatic pathways

Amino acid conjugationAmino acid conjugation

Adds an amino acid to the parent compound.Adds an amino acid to the parent compound.

MercapturicMercapturic acid formationacid formation

Formed by cleavage of the Formed by cleavage of the glycineglycine and and glutamicglutamic acid acid substituentssubstituents from a glutathione conjugate, followed by Nfrom a glutathione conjugate, followed by N--acetylationacetylation of the of the resulting productresulting product

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Significance of Biotransformation Reactions Significance of Biotransformation Reactions in Toxicologyin Toxicology

Biotransformation is a major part of the Biotransformation is a major part of the pathway for elimination of many pathway for elimination of many xenobioticxenobiotic compounds.compounds.

Biotransformation can result in either a Biotransformation can result in either a decrease or an increase (or no change) in decrease or an increase (or no change) in toxicity.toxicity.

Biotransformation can result in the formation Biotransformation can result in the formation of of reactive metabolitesreactive metabolites..

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Good example Good example ––

metabolism of acetaminophenmetabolism of acetaminophen

Acetaminophen is ordinarily metabolized in the liver by Acetaminophen is ordinarily metabolized in the liver by sulfationsulfation and and glucuronidationglucuronidation to form nonto form non--toxic conjugatestoxic conjugates

These are low capacity pathways, in that the cofactors are availThese are low capacity pathways, in that the cofactors are available in able in only limited concentrations, so these are rateonly limited concentrations, so these are rate--limiting.limiting.

As long as the amount of acetaminophen in the liver is relativelAs long as the amount of acetaminophen in the liver is relatively low, y low, the Phase II pathways can handle the compound, and there is no the Phase II pathways can handle the compound, and there is no toxicity.toxicity.

If the concentration of acetaminophen becomes high enough to If the concentration of acetaminophen becomes high enough to overwhelm the capacity of the Phase II pathways, an alternate overwhelm the capacity of the Phase II pathways, an alternate metabolic pathway, involving Phase I enzymes, becomes active.metabolic pathway, involving Phase I enzymes, becomes active.

The product of the Phase I reaction is a highly reactive The product of the Phase I reaction is a highly reactive quinoneiminequinoneimine, , which can form adducts with (bind covalently to) cellular which can form adducts with (bind covalently to) cellular macromolecules, especially proteins.macromolecules, especially proteins.

The binding of the reactive intermediate to cellular macromolecuThe binding of the reactive intermediate to cellular macromolecules les destroys the activity of those molecules, and can lead to comprodestroys the activity of those molecules, and can lead to compromised mised cell function and, ultimately, cell death.cell function and, ultimately, cell death.

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Another good example Another good example ––

metabolism of carbon metabolism of carbon

tetrachloridetetrachloride

Carbon tetrachloride is metabolized by the Carbon tetrachloride is metabolized by the cytochromecytochrome PP--450 system in the liver by 450 system in the liver by abstraction of one of the four chlorine atoms.abstraction of one of the four chlorine atoms.

This results in formation of a highly reactive This results in formation of a highly reactive trichloromethanetrichloromethane radical, which initiates a cascade radical, which initiates a cascade of lipid of lipid peroxidationperoxidation by removing a hydrogen atom by removing a hydrogen atom from membrane phospholipids. from membrane phospholipids.

Damage to the cell membrane causes loss of Damage to the cell membrane causes loss of osmotic integrity, cell swelling and death.osmotic integrity, cell swelling and death.

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The activity of drug metabolizing enzymes is The activity of drug metabolizing enzymes is dependent on numerous factorsdependent on numerous factors

SpeciesSpecies

Age (activity is generally lower in very young and aged Age (activity is generally lower in very young and aged animals)animals)

Sex (activity is generally higher in males than in females)Sex (activity is generally higher in males than in females)

Genetics (remember slow versus fast Genetics (remember slow versus fast acetylatorsacetylators))

Organ (activity of many enzymes is highest in the liver)Organ (activity of many enzymes is highest in the liver)

General health status (e.g., hepatic injury decreases General health status (e.g., hepatic injury decreases metabolic activity in the liver)metabolic activity in the liver)

Diet (remember how fasting decreases the amount of Diet (remember how fasting decreases the amount of glutathione available for GST)glutathione available for GST)

Previous exposure to other compounds Previous exposure to other compounds

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InductionInduction ––

an increase in the activity of one or more an increase in the activity of one or more

enzymes as a result of previous exposure of the enzymes as a result of previous exposure of the organism to compounds that serve as substrates for organism to compounds that serve as substrates for the the enzyme(senzyme(s))

Classic example of an inducer is Classic example of an inducer is phenobarbitalphenobarbital, which , which induces the activity of induces the activity of cytochromecytochrome PP--450 enzymes450 enzymes

Induction may involve either increases in the synthesis of Induction may involve either increases in the synthesis of enzymatic protein, or increases in activation of enzymatic protein, or increases in activation of proenzymesproenzymes..

Induction may result in increases in the amount of cellular Induction may result in increases in the amount of cellular protein, hypertrophy (increases in size) of the cells, and protein, hypertrophy (increases in size) of the cells, and increases in weight of the affected organs (especially liver).increases in weight of the affected organs (especially liver).

One effect of induction of One effect of induction of microsomalmicrosomal enzymes is an enzymes is an increase in the amount of smooth endoplasmic reticulum in increase in the amount of smooth endoplasmic reticulum in a cell. This can be seen microscopically.a cell. This can be seen microscopically.

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Induction is usually temporary, and enzyme activity levels Induction is usually temporary, and enzyme activity levels return to normal after several weeks.return to normal after several weeks.

Induction can result in tolerance to drugs, if the metabolism Induction can result in tolerance to drugs, if the metabolism of the drugs results in a product with lower (or no) of the drugs results in a product with lower (or no) pharmacologic activity. This is why, for example, patients pharmacologic activity. This is why, for example, patients can develop tolerance to Phenobarbital anesthesia after can develop tolerance to Phenobarbital anesthesia after repeated administration.repeated administration.

Induction may result in increases or decreases in toxicity, Induction may result in increases or decreases in toxicity, depending on whether the metabolite is more or less toxic depending on whether the metabolite is more or less toxic than the parent compound. This is why, for example, than the parent compound. This is why, for example, alcoholics are more susceptible to acetaminophen toxicity, alcoholics are more susceptible to acetaminophen toxicity, since alcohol induces the enzyme that is responsible for since alcohol induces the enzyme that is responsible for production of the reactive metabolite from acetaminophen.production of the reactive metabolite from acetaminophen.

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InhibitionInhibition ––

a decrease in the activity of one or more a decrease in the activity of one or more

enzymesenzymes

Classic example of an inhibitor is SKFClassic example of an inhibitor is SKF--525A, which 525A, which inhibits inhibits microsomalmicrosomal enzymesenzymes

Inhibition may be either competitive or nonInhibition may be either competitive or non--competitive.competitive.

Competitive inhibitionCompetitive inhibition

Occurs when an inhibitor binds to the same active site on the Occurs when an inhibitor binds to the same active site on the enzyme as the substrate. The higher the concentration of the enzyme as the substrate. The higher the concentration of the inhibitor, the less likely it is that the substrate molecule wilinhibitor, the less likely it is that the substrate molecule will be able l be able to find and bind to an available enzyme molecule.to find and bind to an available enzyme molecule.

Reversible, since the binding of the inhibitor to the active sitReversible, since the binding of the inhibitor to the active site is not e is not covalentcovalent

Example: Example: OmeprazoleOmeprazole and diazepam are both metabolized by and diazepam are both metabolized by cytochromecytochrome PP--450 2C19 (CYP2C19). Co450 2C19 (CYP2C19). Co--administration of these administration of these two drugs results in prolonged plasma halftwo drugs results in prolonged plasma half--life for diazepam.life for diazepam.

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NonNon--competitive inhibitioncompetitive inhibition

May occur when an inhibitor binds to the same active site on theMay occur when an inhibitor binds to the same active site on the enzyme enzyme as the substrate, but binds so tightly that it is effectively noas the substrate, but binds so tightly that it is effectively not released. Thus, t released. Thus, the binding site is permanently blocked.the binding site is permanently blocked.

May also occur when an inhibitor binds tightly (sometimes covaleMay also occur when an inhibitor binds tightly (sometimes covalently) to a ntly) to a different site on the enzyme than the active site. This can resdifferent site on the enzyme than the active site. This can result in ult in conformational or affinity changes that effectively inactive theconformational or affinity changes that effectively inactive the enzyme.enzyme.

NonNon--competitive inhibition is generally not reversible. Therefore, competitive inhibition is generally not reversible. Therefore, recovery takes much longer because it requires the synthesis of recovery takes much longer because it requires the synthesis of new new enzyme.enzyme.

A special subset of nonA special subset of non--competitive inhibition is competitive inhibition is ““suicide inhibitionsuicide inhibition””, in , in which a compound binds to the active site of an enzyme and is which a compound binds to the active site of an enzyme and is metabolized, but the product then binds irreversibly to the actimetabolized, but the product then binds irreversibly to the active site. An ve site. An example of this is the binding of organophosphate insecticides texample of this is the binding of organophosphate insecticides to the o the enzyme enzyme acetylcholinesteraseacetylcholinesterase ((AchEAchE). This results in a prolonged inhibition ). This results in a prolonged inhibition of of AchEAchE activity.activity.

Inhibition may result in increases or decreases in toxicity, depInhibition may result in increases or decreases in toxicity, depending on ending on whether the metabolite is more or less toxic than the parent comwhether the metabolite is more or less toxic than the parent compound.pound.

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Factors affecting metabolismFactors affecting metabolism1.1. AgeAgeThe metabolizing enzymes in neonates are not fully The metabolizing enzymes in neonates are not fully

developed, therefore those cannot efficiently developed, therefore those cannot efficiently metabolize drugs. Also in the elderly, enzymatic metabolize drugs. Also in the elderly, enzymatic systems may not function well leading to same systems may not function well leading to same conclusion.conclusion.

2. Sex2. SexMales who are deficient in glucose Males who are deficient in glucose --66--phosphate phosphate

dehydrogenasedehydrogenase are more prone to are more prone to hemolysishemolysis when when subjected to some drugs like sulfonamidessubjected to some drugs like sulfonamides

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3. 3. PharmacogeneticPharmacogenetic factorsfactorsSome individuals may be deficient in some enzymes, regardless Some individuals may be deficient in some enzymes, regardless

of sexof sex4. Pregnancy4. PregnancyHepatic metabolism of drugs is decreased in pregnancyHepatic metabolism of drugs is decreased in pregnancy5. Nutritional status/ liver dysfunction5. Nutritional status/ liver dysfunctionMalnutrition can cause a decreased level of some enzyme system Malnutrition can cause a decreased level of some enzyme system

and liver dysfunction can lead to decreased metabolismand liver dysfunction can lead to decreased metabolism6. 6. BioactivationBioactivationSome drugs may be transformed to more toxic metabolitesSome drugs may be transformed to more toxic metabolites7. Enzyme induction / inhibition7. Enzyme induction / inhibitionA result of this is either an increase in the metabolism or a A result of this is either an increase in the metabolism or a

decrease in the drug metabolismdecrease in the drug metabolism8. Changes in the kinetic mechanism: depending on whether the 8. Changes in the kinetic mechanism: depending on whether the

concentration of drug is in the therapeutic or overdose rangeconcentration of drug is in the therapeutic or overdose range

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4. Excretion4. Excretion

Excretion Excretion

The removal of materials from the bodyThe removal of materials from the body

Routes of excretionRoutes of excretion

Bile Bile ––

through the liverthrough the liver

Urine Urine ––

through the kidneysthrough the kidneys

Feces Feces ––

through the intestinesthrough the intestines

Expired air Expired air ––

through the lungsthrough the lungs

Sweat Sweat ––

through the skinthrough the skin

Saliva Saliva ––

through the mouththrough the mouth

MilkMilk

HairHair

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Factors affecting excretionFactors affecting excretion1.1. AgeAgeMany of the functions of the kidney are not well developed in Many of the functions of the kidney are not well developed in

neonates, toxic drugs will be slowly excreted. In the elderly, neonates, toxic drugs will be slowly excreted. In the elderly, lower renal plasma flow will also decrease excretionlower renal plasma flow will also decrease excretion

2. Disease states2. Disease statesRenal and gastrointestinal diseases can considerably slow the Renal and gastrointestinal diseases can considerably slow the

excretionexcretion3. Recirculation 3. Recirculation Some drugs that are excreted as their more water soluble Some drugs that are excreted as their more water soluble

metabolites may be back transformed to the parent metabolites may be back transformed to the parent compound by some bacteriacompound by some bacteria

4. Ion trapping4. Ion trappingWeakly acidic drugs trapped in the tubules are excreted at higheWeakly acidic drugs trapped in the tubules are excreted at higher r

urinary pH as the ion form, thus preventing urinary pH as the ion form, thus preventing reabsorptionreabsorption..