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Genetic polymorphism in dextromethorphan metabolism by CYP2D6 and CYP3A4 enzyme isoforms / Mthokozisi Muziwandile Nkosingiphile MgwabiMgwabi, Mthokozisi Muziwandile Nkosingiphile January 2003 (has links)
Most administered drugs are metabolised in the liver by Phase I enzymes and more
importantly by the cytochrome P450 (CYP) system. The extent of first-pass metabolism is
important in determining whether the drug will have therapeutic or adverse effects after being
administered to a patient. To date the CYP family has been shown to consist of 74 families
denoted as CYPl to CYP118, and only a few families are significantly involved in drug
metabolism. CYP3A4 is the most important isoenzyme followed by CYP2D6, CYP2C9, and
CYP2C19 with a small contribution by CYP2E1, CYP2A6, and CYPlA4. CYP2D6 and
CYP3A4 enzyme isoforms have been well established to exhibit interethnic and interindividual
variability with regard to drug metabolising capacity.
Mutation on the gene coding for a metabolising enzyme is a major cause of variation in drug
metabolism. This mutation gives rise to allelic variants producing enzymes with altered
metabolising activity. The presence of an allele with decreased metabolic activity in an
individual gives rise to the poor metabolising (PM) phenotype. When the PM phenotype
occurs at a frequency of more than 1% within a given population, then the term genetic
polymorphism applies. The aberrant metabolic capacity translates into variable drug
responses of more than 20-fold, leading to different susceptibility to sub-therapeutic effects or
adverse drug reactions. A significant number of drugs, such as the B-adrenergic blockers,
antidepressants, antipsychotic and antiarrhythmic agents, are entirely or partly metabolised by
CYP2D6 and CYP3A4. Genetic polymorphism is especially important for drugs with a narrow
therapeutic/toxicity window.
Phenotyping involves the use of a probe drug that is administered to the subject, followed by
determination of the parent drug and its metabolites in the urine. The aim of this study was to
develop and validate an HPLC method for phenotypic determination of the CYP3A4 and
CYP2D6 enzymes, followed by the application of the assay in a random heterogeneous
population of males.
Dextromethorphan (DXM) was used as an in vivo probe for simultaneous determination of the
phenotypic expression of CYP2D6 and CYP3A4. An HPLC method coupled with a
fluorescence detector was developed for the phenotypic determination of CYP2D6 and
CYP3A4 iso-enzymes as determined by the concentration of dextromethorphan/dextrophan
(DXM/DX) and dextromethorphan/3methoxy-morphinan (DXM/3MM) metabolic ratios
respectively. The compounds were separated on a phenyl column (150 x 4,6 mm, 5-um
particle size) serially connected to nitrile column (250 x 4,6 mm, 5-um particle size) using
mobile phase of 80% (1.5% glacial acetic acid and 0.1% triethyl amine in distilled water) and
20% acetonitrile. Solid phase extraction was used to extract the analytes from urine samples
using silica cartridges. The suitability of the method was demonstrated in a preliminary study
with sixteen healthy Caucasian males. After a single oral 30 mg DXM dose, the volunteers
were required to collect all urine samples voided 8 hours post oral dose. DXM/3HM and
DXM/DX metabolic ratios were determined from collected urine samples.
The method was validated for DXM and DX at a concentration range of 0.25 - 30 ug/ml, and at
0.025 - 3 ug/ml for 3MM. Calibration curves were linear with R2 values of at-least 0.999 for all
compounds of interest. Recoveries were 97%, 93%, and 65% for DX, DXM and 3MM,
respectively. The method was reproducible with intra-day precision having coefficients of
variation percentage (CV%) of less than 17% for all analytes. Inter-day precision had a CV%
of less than 14% for all analytes. The limit of detection was 30 ug/ml for all compounds. All
volunteers were classified with an extensive metaboliser (EM) phenotype. In conclusion the
method described is suitable for polymorphic determination of CYP2D6 and CYP3A4 in a
population study, and may have value in further studies planned at investigating the critical
issue of racial genetic polymorphism in ethnic groups in South Africa. / Thesis (M.Sc. (Pharm.))--North-West University, Potchefstroom Campus, 2004.
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Genetic polymorphism in dextromethorphan metabolism by CYP2D6 and CYP3A4 enzyme isoforms / Mthokozisi Muziwandile Nkosingiphile MgwabiMgwabi, Mthokozisi Muziwandile Nkosingiphile January 2003 (has links)
Most administered drugs are metabolised in the liver by Phase I enzymes and more
importantly by the cytochrome P450 (CYP) system. The extent of first-pass metabolism is
important in determining whether the drug will have therapeutic or adverse effects after being
administered to a patient. To date the CYP family has been shown to consist of 74 families
denoted as CYPl to CYP118, and only a few families are significantly involved in drug
metabolism. CYP3A4 is the most important isoenzyme followed by CYP2D6, CYP2C9, and
CYP2C19 with a small contribution by CYP2E1, CYP2A6, and CYPlA4. CYP2D6 and
CYP3A4 enzyme isoforms have been well established to exhibit interethnic and interindividual
variability with regard to drug metabolising capacity.
Mutation on the gene coding for a metabolising enzyme is a major cause of variation in drug
metabolism. This mutation gives rise to allelic variants producing enzymes with altered
metabolising activity. The presence of an allele with decreased metabolic activity in an
individual gives rise to the poor metabolising (PM) phenotype. When the PM phenotype
occurs at a frequency of more than 1% within a given population, then the term genetic
polymorphism applies. The aberrant metabolic capacity translates into variable drug
responses of more than 20-fold, leading to different susceptibility to sub-therapeutic effects or
adverse drug reactions. A significant number of drugs, such as the B-adrenergic blockers,
antidepressants, antipsychotic and antiarrhythmic agents, are entirely or partly metabolised by
CYP2D6 and CYP3A4. Genetic polymorphism is especially important for drugs with a narrow
therapeutic/toxicity window.
Phenotyping involves the use of a probe drug that is administered to the subject, followed by
determination of the parent drug and its metabolites in the urine. The aim of this study was to
develop and validate an HPLC method for phenotypic determination of the CYP3A4 and
CYP2D6 enzymes, followed by the application of the assay in a random heterogeneous
population of males.
Dextromethorphan (DXM) was used as an in vivo probe for simultaneous determination of the
phenotypic expression of CYP2D6 and CYP3A4. An HPLC method coupled with a
fluorescence detector was developed for the phenotypic determination of CYP2D6 and
CYP3A4 iso-enzymes as determined by the concentration of dextromethorphan/dextrophan
(DXM/DX) and dextromethorphan/3methoxy-morphinan (DXM/3MM) metabolic ratios
respectively. The compounds were separated on a phenyl column (150 x 4,6 mm, 5-um
particle size) serially connected to nitrile column (250 x 4,6 mm, 5-um particle size) using
mobile phase of 80% (1.5% glacial acetic acid and 0.1% triethyl amine in distilled water) and
20% acetonitrile. Solid phase extraction was used to extract the analytes from urine samples
using silica cartridges. The suitability of the method was demonstrated in a preliminary study
with sixteen healthy Caucasian males. After a single oral 30 mg DXM dose, the volunteers
were required to collect all urine samples voided 8 hours post oral dose. DXM/3HM and
DXM/DX metabolic ratios were determined from collected urine samples.
The method was validated for DXM and DX at a concentration range of 0.25 - 30 ug/ml, and at
0.025 - 3 ug/ml for 3MM. Calibration curves were linear with R2 values of at-least 0.999 for all
compounds of interest. Recoveries were 97%, 93%, and 65% for DX, DXM and 3MM,
respectively. The method was reproducible with intra-day precision having coefficients of
variation percentage (CV%) of less than 17% for all analytes. Inter-day precision had a CV%
of less than 14% for all analytes. The limit of detection was 30 ug/ml for all compounds. All
volunteers were classified with an extensive metaboliser (EM) phenotype. In conclusion the
method described is suitable for polymorphic determination of CYP2D6 and CYP3A4 in a
population study, and may have value in further studies planned at investigating the critical
issue of racial genetic polymorphism in ethnic groups in South Africa. / Thesis (M.Sc. (Pharm.))--North-West University, Potchefstroom Campus, 2004.
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Palladium telluride quantum dots biosensor for the determination of indinavir drugFeleni, Usisipho January 2013 (has links)
Magister Scientiae - MSc / Indinavir is a potent and well tolerated protease inhibitor drug used as a component of the highly active antiretroviral therapy (HAART) of HIV/AIDS, which results in pharmacokinetics that may be favourable or adverse. These drugs work by maintaining a plasma concentration that is sufficient to inhibit viral replication and thereby suppressing a patient’s viral load. A number of antiretroviral drugs, including indinavir, undergo metabolism that is catalysed by cytochrome P450-3A4 enzyme found in the human liver microsomes. The rate of drug metabolism influences a patient’s response to treatment as well as drug interactions that may lead to life-threatening toxic conditions, such as haemolytic anaemia, kidney failure and liver problems. Therapeutic drug monitoring (TDM) during HIV/AIDS treatment has been suggested to have a potential to reduce drug toxicity and optimise individual therapy. A fast and reliable detection technique, such as biosensing, is
therefore necessary for the determination of a patient’s metabolic profile for indinavir and for appropriate dosing of the drugs. In this study biosensors developed for the determination of ARV drugs comprised of cysteamine self-assembled on a gold electrode, on which was attached 3-mercaptopropionic acid-capped palladium telluride (3-MPA-PdTe) or thioglycolic acid-capped palladium telluride (TGA-PdTe) quantum dots that are cross-linked to cytochrome P450-3A4 (CYP3A4) in the presence of 1-ethyl-3(3-dimethylaminopropyl) carbodiimide hydrochloride and N-hydroxysuccinimide. The quantum dots were synthesized
in the presence of capping agents (3-MPA or TGA) to improve their stability, solubility and biocompatibility. The capping of PdTe quantum dots with TGA or 3-MPA was confirmed by FTIR, where the SH group absorption band disappeared from the spectra of 3-MPA-PdTe and TGA-PdTe. The particle size of the quantum dots (< 5 nm) was estimated from high resolution transmission electron microscopy (HRTEM) measurements. Optical properties of the materials were confirmed by UV-Vis spectrophotometry which produced absorption iii bands at ~320 nm that corresponded to energy band gap values of 3 eV (3.87 eV) for TGAPdTe (3-MPA-PdTe) quantum dots. The electrocatalytic properties of the quantum dots biosensor systems were studied by cyclic voltammetry (CV) for which the characteristic reduction peak at 0.75 V was used to detect the response of the biosensor to indinavir. Results for indinavir biosensor constructed with 3-MPA-SnSe quantum dots are also reported in this thesis. The three biosensors systems were very sensitive towards indinavir; and gave low limits of detection (LOD) values of 3.22, 4.3 and 6.2 ng/mL for 3-MPA-SnSe, 3-MPA-PdTe and TGA-PdTe quantum dots biosensors, respectively. The LOD values are within the
‘maximum plasma concentration’ (Cmax) value of indinavir (5 - 15 ng/mL) normally observed 8 h after drug intake.
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Development of electrochemical ZnSe Quantam dots biosensors for low-level detection of 17β-Estradiol estrogenic endocrine disrupting compoundJijana, Abongile Nwabisa January 2010 (has links)
<p>The main thesis hub was on development of two electrochemical biosensors for the determination of 17&beta / -estradiol: an estrogenic endocrine disrupting compound. Endocronology have significantly shown that the endocrine disruptors contribute tremendously to health problems encountered by living species today, problems such as breast cancer, reproductive abnormalities, a decline in male population most significant to aquatic vertebrates, reduced fertility and other infinite abnormalities recurring in the reproductive system of mostly male species. The first biosensor developed for the detection of 17&beta / -estradiol endocrine disrupting compound / consisted of an electro-active polymeric 3-mercaptoprorionic acid capped zinc selenide quantum dots cross linked to horseradish peroxidase (HRP) enzyme as a bio-recognition element. The second biosensor developed was comprised of cysteamine self assembled to gold electrode, with 3-mercaptopropionic acid capped zinc selenide quantum dots cross linked to cytochrome P450-3A4 (CYP3A4) enzyme in the presence of 1-ethyl-3-(3- dimethylaminopropyl)carbodiimide hydrochloride and succinimide.</p>
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Development of electrochemical ZnSe Quantam dots biosensors for low-level detection of 17β-Estradiol estrogenic endocrine disrupting compoundJijana, Abongile Nwabisa January 2010 (has links)
<p>The main thesis hub was on development of two electrochemical biosensors for the determination of 17&beta / -estradiol: an estrogenic endocrine disrupting compound. Endocronology have significantly shown that the endocrine disruptors contribute tremendously to health problems encountered by living species today, problems such as breast cancer, reproductive abnormalities, a decline in male population most significant to aquatic vertebrates, reduced fertility and other infinite abnormalities recurring in the reproductive system of mostly male species. The first biosensor developed for the detection of 17&beta / -estradiol endocrine disrupting compound / consisted of an electro-active polymeric 3-mercaptoprorionic acid capped zinc selenide quantum dots cross linked to horseradish peroxidase (HRP) enzyme as a bio-recognition element. The second biosensor developed was comprised of cysteamine self assembled to gold electrode, with 3-mercaptopropionic acid capped zinc selenide quantum dots cross linked to cytochrome P450-3A4 (CYP3A4) enzyme in the presence of 1-ethyl-3-(3- dimethylaminopropyl)carbodiimide hydrochloride and succinimide.</p>
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Development of electrochemical ZnSe Quantam dots biosensors for low-level detection of 17β-Estradiol estrogenic endocrine disrupting compoundJijana, Abongile Nwabisa January 2010 (has links)
Magister Scientiae - MSc / The main thesis hub was on development of two electrochemical biosensors for the determination of 17β-estradiol-estradiol: an estrogenic endocrine disrupting compound. Endocronology have significantly shown that the endocrine disruptors contribute tremendously to health problems encountered by living species today, problems such as breast cancer, reproductive abnormalities, a decline in male population most significant to aquatic vertebrates, reduced fertility and other infinite abnormalities recurring in the reproductive system of mostly male species. The first biosensor developed for the detection of 17β-estradiol-estradiol endocrine disrupting compound; consisted of an electro-active polymeric 3-mercaptoprorionic acid capped zinc selenide quantum dots cross linked to horseradish peroxidase (HRP) enzyme as a bio-recognition element. The second biosensor developed was comprised of cysteamine self assembled to gold electrode, with 3-mercaptopropionic acid capped zinc selenide quantum dots cross linked to cytochrome P450-3A4 (CYP3A4) enzyme in the presence of 1-ethyl-3-(3- dimethylaminopropyl)carbodiimide hydrochloride and succinimide. / South Africa
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