Leah Falade - Academia.edu (original) (raw)
Papers by Leah Falade
Bioanalysis, Feb 1, 2014
The US FDA published A Guidance for Industry: Bioanalytical Method Validation in May 2001. Despit... more The US FDA published A Guidance for Industry: Bioanalytical Method Validation in May 2001. Despite the publication of the guidance, companies continue to submit bioequivalence studies with bioanalytical deficiencies that preclude Abbreviated New Drug Application approval. The Divisions of Bioequivalence in the FDA's Office of Generic Drugs conducted a survey of the bioequivalence submissions over a 10-year period (2001-2011) to identify the most commonly occurring bioanalytical deficiencies. Data from a total of 4028 Abbreviated New Drug Application submissions were collected to identify bioanalytical deficiencies. Of the three categories of bioanalytical deficiencies (method, validation and report), the majority of the deficiencies were from the bioanalytical method validation section. Globally, the percentage of bioanalytical method validation deficiencies was 62%. The approval of generic drugs would be accelerated if these deficiencies were avoided by generic companies by adhering to the guidance and therefore submitting a more complete application.
Journal of Pharmacology and Experimental Therapeutics, Jun 4, 2007
Persistent behavioral abnormalities have been commonly associated with acute organophosphate (OP)... more Persistent behavioral abnormalities have been commonly associated with acute organophosphate (OP) pesticide poisoning; however, relatively little is known about the consequences of chronic OP exposures that are not associated with acute cholinergic symptoms. In this study, the behavioral and neurochemical effects of chronic, intermittent, and subthreshold exposures to the OP pesticide, chlorpyrifos (CPF), were investigated. Rats were injected with CPF s.c. (dose range, 2.5-18.0 mg/kg) every other day over the course of 30 days and then were given a 2-week CPF-free washout period. In behavioral experiments conducted during the washout period, dosedependent decrements in a water-maze hidden platform task and a prepulse inhibition procedure were observed, without significant effects on open-field activity, Rotorod performance, grip strength, or a spontaneous novel object recognition task. After washout, levels of CPF and its metabolite 3,5,6-trichloro-2-pyridinol were minimal in plasma and brain; however, cholinesterase inhibition was still detectable. Furthermore, the 18.0 mg/kg dose of CPF was associated with (brain region-dependent) decreases in nerve growth factor receptors and cholinergic proteins including the vesicular acetylcholine transporter, the high-affinity choline transporter, and the ␣ 7-nicotinic acetylcholine receptor. These deficits were accompanied by decreases in anterograde and retrograde axonal transport measured in sciatic nerves ex vivo. Thus, low-level (intermittent) exposure to CPF has persistent effects on neurotrophin receptors and cholinergic proteins, possibly through inhibition of fast axonal transport. Such neurochemical changes may lead to deficits in information processing and cognitive function. This work was supported by the National Institute of Environmental Health Sciences (ES012241 to A.V.T.). Salary support (to J.J.B.) also was contributed through a Merit Review Award from the Veterans Administration. Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
Rapid Communications in Mass Spectrometry, 2006
A method has been developed to quantify chlorpyrifos (O,O‐diethyl‐O‐[3,5,6,‐trichloro‐2‐pyridyl] ... more A method has been developed to quantify chlorpyrifos (O,O‐diethyl‐O‐[3,5,6,‐trichloro‐2‐pyridyl] phosphorothionate) and its metabolites chlorpyrifos‐oxon (O,O‐diethyl‐O‐[3,5,6,trichloro‐2‐pyridinyl] phosphate) and TCP (3,5,6,‐trichloro‐2‐pyridinol) in rat brain tissue by coupled‐column liquid chromatography/electrospray ionization tandem mass spectrometry (LC/LC/ESI‐MS/MS). Rat brains were homogenized and treated by protein precipitation using ice‐cold acetonitrile. The supernatant was directly injected onto the coupled‐column system. Sample clean‐up was achieved on a Zorbax Extend‐C18 column (2.1 × 50 mm, 5 µm) using a mobile phase of acetonitrile/water with 0.0025% formic acid (40:60, v/v). The compounds were separated isocratically on a Zorbax Eclipse XDB C8 column (2.0 × 150 mm, 5 µm) using a mobile phase of acetonitrile/water with 0.0025% formic acid (75:25, v/v). Chlorpyrifos and chlorpyrifos‐oxon were detected in positive ion mode using multiple reaction monitoring (MRM). TCP was detected in negative ion mode using precursor‐to‐precursor transition monitoring. The method was validated and the specificity, linearity, limit of quantitation (LOQ), precision, accuracy, stability, and recoveries were determined. Calibration curves for all three analytes yielded correlation coefficients of 0.993 or greater. The LOQs were 25.3 ng/g for chlorpyrifos and 6.3 ng/g for chlorpyrifos‐oxon and TCP. All precision relative standard deviations (RSDs) were less than 16% for the LOQ and less than 11% for the other QC samples. This method was successfully applied to six rats that were injected subcutaneously with chlorpyrifos. Copyright © 2006 John Wiley & Sons, Ltd.
Journal of Liquid Chromatography & Related Technologies, 2007
A method has been developed to quantify chlorpyrifos (O,O‐diethyl O‐[3,5,6,‐trichloro‐2‐pyridyl] ... more A method has been developed to quantify chlorpyrifos (O,O‐diethyl O‐[3,5,6,‐trichloro‐2‐pyridyl] phosphorothionate) and its metabolites chlorpyrifos‐oxon (O,O‐diethyl‐O[3,5,6,trichloro‐2‐pyridyl] phosphate) and TCP (3,5,6,‐trichloro‐2‐pyridinol) in rat blood by liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI‐MS/MS). Rat blood was treated by liquid‐liquid extraction and the analytes were separated by gradient elution on an Agilent Zorbax Extended‐C8 column (2.0×150 mm, 5 µm). Chlorpyrifos and chlorpyrifos‐oxon were detected in
Biomedical Chromatography, 2007
Time-of-flight (TOF) instruments have recently gained popularity in quantitative analyses. Normal... more Time-of-flight (TOF) instruments have recently gained popularity in quantitative analyses. Normally, TOF mass spectrometers are used for accurate mass measurements for empirical formula verification. However, over the past decade, they have been used quantitatively as well. Because of the fast separations and narrow peaks that result from gas chromatography separations, scanning mass spectrometers are not ideal detectors. TOF mass spectrometers, however, have the ability to collect spectra at a faster rate. Two-dimensional gas chromatography has also been introduced to further resolve peaks from complex matrices. Two-dimensional gas chromatography results in a faster separation as well as narrower peaks. This paper reviews the methods currently in the literature for the quantitation of compounds using one- and two-dimensional gas chromatography and TOF mass spectrometry detection.
Journal of Chromatography B, Nov 1, 2004
2 ,3-Dideoxycytidine (DDC) is a nucleoside reverse transcriptase inhibitor that has been shown to... more 2 ,3-Dideoxycytidine (DDC) is a nucleoside reverse transcriptase inhibitor that has been shown to inhibit the human immunodeficiency virus (HIV). DDC is a candidate for treatment of pregnant women to prevent prenatal transmission of HIV/AIDS to their unborn children. A quick and simple high-performance liquid chromatography (HPLC) method has been developed and validated for the determination of DDC concentrations in samples collected from a pregnant rat model (maternal plasma, amniotic fluid, placental and fetal tissues). Extraction of DDC and its internal standard 2 ,3-dideoxy-3-thiacytidine (3TC) in plasma and amniotic fluid was carried out by protein precipitation. Extraction from placental and fetal homogenates was achieved by solid phase extraction using Waters Oasis HLB solid phase extraction cartridges. Chromatographic separation was achieved on a Waters Spherisorb S3W silica column (4.6 mm × 100 mm) equipped with a Phenomenex guard column. The mobile phase used was 10% methanol in water with 22 mM formic acid. The flow rate was 0.5 ml/min, and the detection wavelength was optimized at 275 nm. Under these chromatographic conditions, DDC eluted around 12 min, and 3TC eluted around 10 min. The calibration curves for each day of validation and analysis showed good linear response through the range of 0.15-75.0 g/ml in each of the four matrices. The relative recovery for DDC in each of the matrices ranged from 87.8% to 103.0%. Acceptable intra-and inter-day assay precision (<15% R.S.D.) and accuracy (<15% error) were observed over 0.15-75.0 g/ml for all four matrices.
Biomedical Chromatography, 2007
Over the last decade, time‐of‐flight (TOF) instruments have increasingly been used as quantitatio... more Over the last decade, time‐of‐flight (TOF) instruments have increasingly been used as quantitation tools. In addition, because of their high resolving power, they can be used for verification of empirical formulas. Historically, TOF instruments have had limited quantitation capabilities because of their narrow dynamic range. However, recent advances have improved these limitations. This review covers the rationale for using TOF for LC detection, and describes the many methods currently in the literature for the quantitation of pharmaceuticals, environmental pollutants, explosives and many phytochemicals. Copyright © 2007 John Wiley & Sons, Ltd.
Humans can be exposed to central nervous system agents environmentally and/or therapeutically. Ch... more Humans can be exposed to central nervous system agents environmentally and/or therapeutically. Chronic exposure to these agents can cause damage to the central nervous system. In order to study the neurological effects of these agents, sensitive and selective analytical methods must be developed and validated in biological matrices, such as brain tissue, plasma, and whole blood. Chlorpyrifos (CPF) is an organophosphate pesticide that inhibits acetylcholinesterase, which is an enzyme that is necessary for normal function of the nervous system. CPF is metabolized by cytochrome P450 into chlorpyrifos-oxon (CPF-O) and TCP. CPF-O is about 3,000 times more potent than CPF in its inhibition of acetylcholinesterase activity, while TCP is non-toxic and is eliminated by the kidneys. Therapeutically, chronic exposure to antipsychotic drugs can result in abnormalities in motor function. Additionally, it is important to determine the extent to which a correlation exists between plasma and brain levels of these agents and cognitive function. Chapter 1 is the introduction and literature review that describes the layout of the dissertation and reviews the literature for analytical methods using liquid chromatography with time-of-flight mass spectrometry (TOF-MS) for quantitation. Chapter 2 reviews the literature for analytical methods using gas chromatography and TOF-MS v ACKNOWLEDGEMENTS I would first like to thank God for bringing me this far and for blessing me with an abundance of loving and supportive family members and friends. I thank my parents, James and Starletta Williamson, my brother James, and everyone else in my family that has continued to support me during my academic career. I love you all and am so grateful to have you in my life. I would like to thank my graduate advisor, Dr. Michael Bartlett. It has been an excellent experience learning and working under your direction. I feel extremely prepared for a career in analytical chemistry; whether it be in academia or industry. I would also like to thank my committee members, Drs. Beach, Capomacchia, Orlando and White. I owe a special thanks to Dr. Capomacchia for providing me with the opportunity to study here at UGA with the Bridges to Doctorate grant. I thank my friends from home and my new friends I have made during my time here at UGA. Thank you for being there for me if I needed to vent, needed to pull an allnighter studying, wanted you to try a new recipe I found, or if I just wanted to chat. I would like to thank my past and present labmates for their training and incite; Amy, Shonetta, Guodong, Yongzhen, and Meng. I could not have done this alone, thank you all for your support. vi
Journal of Liquid Chromatography & Related Technologies, Sep 30, 2008
Abstract Tandem mass spectrometry (MS/MS) is frequently used for the detection of small molecules... more Abstract Tandem mass spectrometry (MS/MS) is frequently used for the detection of small molecules in biological samples because of the high specificity and sensitivity associated with these instruments. Time-of-flight mass spectrometers (TOF-MS) are typically used ...
Journal of Pharmacology and Experimental Therapeutics, 2007
Persistent behavioral abnormalities have been commonly associated with acute organophosphate (OP)... more Persistent behavioral abnormalities have been commonly associated with acute organophosphate (OP) pesticide poisoning; however, relatively little is known about the consequences of chronic OP exposures that are not associated with acute cholinergic symptoms. In this study, the behavioral and neurochemical effects of chronic, intermittent, and subthreshold exposures to the OP pesticide, chlorpyrifos (CPF), were investigated. Rats were injected with CPF s.c. (dose range, 2.5-18.0 mg/kg) every other day over the course of 30 days and then were given a 2-week CPF-free washout period. In behavioral experiments conducted during the washout period, dosedependent decrements in a water-maze hidden platform task and a prepulse inhibition procedure were observed, without significant effects on open-field activity, Rotorod performance, grip strength, or a spontaneous novel object recognition task. After washout, levels of CPF and its metabolite 3,5,6-trichloro-2-pyridinol were minimal in plasma and brain; however, cholinesterase inhibition was still detectable. Furthermore, the 18.0 mg/kg dose of CPF was associated with (brain region-dependent) decreases in nerve growth factor receptors and cholinergic proteins including the vesicular acetylcholine transporter, the high-affinity choline transporter, and the ␣ 7-nicotinic acetylcholine receptor. These deficits were accompanied by decreases in anterograde and retrograde axonal transport measured in sciatic nerves ex vivo. Thus, low-level (intermittent) exposure to CPF has persistent effects on neurotrophin receptors and cholinergic proteins, possibly through inhibition of fast axonal transport. Such neurochemical changes may lead to deficits in information processing and cognitive function. This work was supported by the National Institute of Environmental Health Sciences (ES012241 to A.V.T.). Salary support (to J.J.B.) also was contributed through a Merit Review Award from the Veterans Administration. Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
Experiments using inductively coupled plasma-mass spectrometry (ICP-MS) as a detection system for... more Experiments using inductively coupled plasma-mass spectrometry (ICP-MS) as a detection system for reversed-phase chromatographic methods have been conducted for the analyses of various pharmaceutical compounds. Interfacing the ICP-MS instrument to the high performance liquid chromatography (HPLC) system was straightforward. The eluent from the HPLC column was pumped through a photodiode array (PDA) and then directly into the ICP-MS nebulizer using standard sample introduction components at flow rates ranging from 0.5 to 1.5 mL/min. Two separate sets of experiments were carried out. Initial speciation experiments were conducted on the B supplements cyanocobalamin (vitamin B 12 ), biotin, and thiamine (vitamin B 1 ) using isocratic separations. ICP-MS chromatograms were generated by monitoring cobalt at m/z 59, phosphorusatm/z31,andsulfuratm/z34simultaneously. Linearregression data were obtained for cyanocobalamin, thiamine hydrochloride, and biotin. Elemental detection was used to re...
Bioanalysis, 2014
The US FDA published A Guidance for Industry: Bioanalytical Method Validation in May 2001. Despit... more The US FDA published A Guidance for Industry: Bioanalytical Method Validation in May 2001. Despite the publication of the guidance, companies continue to submit bioequivalence studies with bioanalytical deficiencies that preclude Abbreviated New Drug Application approval. The Divisions of Bioequivalence in the FDA's Office of Generic Drugs conducted a survey of the bioequivalence submissions over a 10-year period (2001-2011) to identify the most commonly occurring bioanalytical deficiencies. Data from a total of 4028 Abbreviated New Drug Application submissions were collected to identify bioanalytical deficiencies. Of the three categories of bioanalytical deficiencies (method, validation and report), the majority of the deficiencies were from the bioanalytical method validation section. Globally, the percentage of bioanalytical method validation deficiencies was 62%. The approval of generic drugs would be accelerated if these deficiencies were avoided by generic companies by adh...
Journal of Liquid Chromatography & Related Technologies, 2007
A method has been developed to quantify chlorpyrifos (O,O‐diethyl O‐[3,5,6,‐trichloro‐2‐pyridyl] ... more A method has been developed to quantify chlorpyrifos (O,O‐diethyl O‐[3,5,6,‐trichloro‐2‐pyridyl] phosphorothionate) and its metabolites chlorpyrifos‐oxon (O,O‐diethyl‐O[3,5,6,trichloro‐2‐pyridyl] phosphate) and TCP (3,5,6,‐trichloro‐2‐pyridinol) in rat blood by liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI‐MS/MS). Rat blood was treated by liquid‐liquid extraction and the analytes were separated by gradient elution on an Agilent Zorbax Extended‐C8 column (2.0×150 mm, 5 µm). Chlorpyrifos and chlorpyrifos‐oxon were detected in
Journal of Chromatography B, 2004
2 ,3-Dideoxycytidine (DDC) is a nucleoside reverse transcriptase inhibitor that has been shown to... more 2 ,3-Dideoxycytidine (DDC) is a nucleoside reverse transcriptase inhibitor that has been shown to inhibit the human immunodeficiency virus (HIV). DDC is a candidate for treatment of pregnant women to prevent prenatal transmission of HIV/AIDS to their unborn children. A quick and simple high-performance liquid chromatography (HPLC) method has been developed and validated for the determination of DDC concentrations in samples collected from a pregnant rat model (maternal plasma, amniotic fluid, placental and fetal tissues). Extraction of DDC and its internal standard 2 ,3-dideoxy-3-thiacytidine (3TC) in plasma and amniotic fluid was carried out by protein precipitation. Extraction from placental and fetal homogenates was achieved by solid phase extraction using Waters Oasis HLB solid phase extraction cartridges. Chromatographic separation was achieved on a Waters Spherisorb S3W silica column (4.6 mm × 100 mm) equipped with a Phenomenex guard column. The mobile phase used was 10% methanol in water with 22 mM formic acid. The flow rate was 0.5 ml/min, and the detection wavelength was optimized at 275 nm. Under these chromatographic conditions, DDC eluted around 12 min, and 3TC eluted around 10 min. The calibration curves for each day of validation and analysis showed good linear response through the range of 0.15-75.0 g/ml in each of the four matrices. The relative recovery for DDC in each of the matrices ranged from 87.8% to 103.0%. Acceptable intra-and inter-day assay precision (<15% R.S.D.) and accuracy (<15% error) were observed over 0.15-75.0 g/ml for all four matrices.
Humans can be exposed to central nervous system agents environmentally and/or therapeutically. Ch... more Humans can be exposed to central nervous system agents environmentally and/or therapeutically. Chronic exposure to these agents can cause damage to the central nervous system. In order to study the neurological effects of these agents, sensitive and selective ...
Biomedical Chromatography, 2007
Time-of-flight (TOF) instruments have recently gained popularity in quantitative analyses. Normal... more Time-of-flight (TOF) instruments have recently gained popularity in quantitative analyses. Normally, TOF mass spectrometers are used for accurate mass measurements for empirical formula verification. However, over the past decade, they have been used quantitatively as well. Because of the fast separations and narrow peaks that result from gas chromatography separations, scanning mass spectrometers are not ideal detectors. TOF mass spectrometers, however, have the ability to collect spectra at a faster rate. Two-dimensional gas chromatography has also been introduced to further resolve peaks from complex matrices. Two-dimensional gas chromatography results in a faster separation as well as narrower peaks. This paper reviews the methods currently in the literature for the quantitation of compounds using one- and two-dimensional gas chromatography and TOF mass spectrometry detection.
Biomedical Chromatography, 2007
Over the last decade, time-of-flight (TOF) instruments have increasingly been used as quantitatio... more Over the last decade, time-of-flight (TOF) instruments have increasingly been used as quantitation tools. In addition, because of their high resolving power, they can be used for verification of empirical formulas. Historically, TOF instruments have had limited quantitation capabilities because of their narrow dynamic range. However, recent advances have improved these limitations. This review covers the rationale for using TOF for LC detection, and describes the many methods currently in the literature for the quantitation of pharmaceuticals, environmental pollutants, explosives and many phytochemicals.
Journal of Liquid Chromatography & Related Technologies, 2008
Abstract Tandem mass spectrometry (MS/MS) is frequently used for the detection of small molecules... more Abstract Tandem mass spectrometry (MS/MS) is frequently used for the detection of small molecules in biological samples because of the high specificity and sensitivity associated with these instruments. Time-of-flight mass spectrometers (TOF-MS) are typically used ...
Bioanalysis, Feb 1, 2014
The US FDA published A Guidance for Industry: Bioanalytical Method Validation in May 2001. Despit... more The US FDA published A Guidance for Industry: Bioanalytical Method Validation in May 2001. Despite the publication of the guidance, companies continue to submit bioequivalence studies with bioanalytical deficiencies that preclude Abbreviated New Drug Application approval. The Divisions of Bioequivalence in the FDA&amp;amp;amp;amp;amp;amp;amp;#39;s Office of Generic Drugs conducted a survey of the bioequivalence submissions over a 10-year period (2001-2011) to identify the most commonly occurring bioanalytical deficiencies. Data from a total of 4028 Abbreviated New Drug Application submissions were collected to identify bioanalytical deficiencies. Of the three categories of bioanalytical deficiencies (method, validation and report), the majority of the deficiencies were from the bioanalytical method validation section. Globally, the percentage of bioanalytical method validation deficiencies was 62%. The approval of generic drugs would be accelerated if these deficiencies were avoided by generic companies by adhering to the guidance and therefore submitting a more complete application.
Journal of Pharmacology and Experimental Therapeutics, Jun 4, 2007
Persistent behavioral abnormalities have been commonly associated with acute organophosphate (OP)... more Persistent behavioral abnormalities have been commonly associated with acute organophosphate (OP) pesticide poisoning; however, relatively little is known about the consequences of chronic OP exposures that are not associated with acute cholinergic symptoms. In this study, the behavioral and neurochemical effects of chronic, intermittent, and subthreshold exposures to the OP pesticide, chlorpyrifos (CPF), were investigated. Rats were injected with CPF s.c. (dose range, 2.5-18.0 mg/kg) every other day over the course of 30 days and then were given a 2-week CPF-free washout period. In behavioral experiments conducted during the washout period, dosedependent decrements in a water-maze hidden platform task and a prepulse inhibition procedure were observed, without significant effects on open-field activity, Rotorod performance, grip strength, or a spontaneous novel object recognition task. After washout, levels of CPF and its metabolite 3,5,6-trichloro-2-pyridinol were minimal in plasma and brain; however, cholinesterase inhibition was still detectable. Furthermore, the 18.0 mg/kg dose of CPF was associated with (brain region-dependent) decreases in nerve growth factor receptors and cholinergic proteins including the vesicular acetylcholine transporter, the high-affinity choline transporter, and the ␣ 7-nicotinic acetylcholine receptor. These deficits were accompanied by decreases in anterograde and retrograde axonal transport measured in sciatic nerves ex vivo. Thus, low-level (intermittent) exposure to CPF has persistent effects on neurotrophin receptors and cholinergic proteins, possibly through inhibition of fast axonal transport. Such neurochemical changes may lead to deficits in information processing and cognitive function. This work was supported by the National Institute of Environmental Health Sciences (ES012241 to A.V.T.). Salary support (to J.J.B.) also was contributed through a Merit Review Award from the Veterans Administration. Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
Rapid Communications in Mass Spectrometry, 2006
A method has been developed to quantify chlorpyrifos (O,O‐diethyl‐O‐[3,5,6,‐trichloro‐2‐pyridyl] ... more A method has been developed to quantify chlorpyrifos (O,O‐diethyl‐O‐[3,5,6,‐trichloro‐2‐pyridyl] phosphorothionate) and its metabolites chlorpyrifos‐oxon (O,O‐diethyl‐O‐[3,5,6,trichloro‐2‐pyridinyl] phosphate) and TCP (3,5,6,‐trichloro‐2‐pyridinol) in rat brain tissue by coupled‐column liquid chromatography/electrospray ionization tandem mass spectrometry (LC/LC/ESI‐MS/MS). Rat brains were homogenized and treated by protein precipitation using ice‐cold acetonitrile. The supernatant was directly injected onto the coupled‐column system. Sample clean‐up was achieved on a Zorbax Extend‐C18 column (2.1 × 50 mm, 5 µm) using a mobile phase of acetonitrile/water with 0.0025% formic acid (40:60, v/v). The compounds were separated isocratically on a Zorbax Eclipse XDB C8 column (2.0 × 150 mm, 5 µm) using a mobile phase of acetonitrile/water with 0.0025% formic acid (75:25, v/v). Chlorpyrifos and chlorpyrifos‐oxon were detected in positive ion mode using multiple reaction monitoring (MRM). TCP was detected in negative ion mode using precursor‐to‐precursor transition monitoring. The method was validated and the specificity, linearity, limit of quantitation (LOQ), precision, accuracy, stability, and recoveries were determined. Calibration curves for all three analytes yielded correlation coefficients of 0.993 or greater. The LOQs were 25.3 ng/g for chlorpyrifos and 6.3 ng/g for chlorpyrifos‐oxon and TCP. All precision relative standard deviations (RSDs) were less than 16% for the LOQ and less than 11% for the other QC samples. This method was successfully applied to six rats that were injected subcutaneously with chlorpyrifos. Copyright © 2006 John Wiley & Sons, Ltd.
Journal of Liquid Chromatography & Related Technologies, 2007
A method has been developed to quantify chlorpyrifos (O,O‐diethyl O‐[3,5,6,‐trichloro‐2‐pyridyl] ... more A method has been developed to quantify chlorpyrifos (O,O‐diethyl O‐[3,5,6,‐trichloro‐2‐pyridyl] phosphorothionate) and its metabolites chlorpyrifos‐oxon (O,O‐diethyl‐O[3,5,6,trichloro‐2‐pyridyl] phosphate) and TCP (3,5,6,‐trichloro‐2‐pyridinol) in rat blood by liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI‐MS/MS). Rat blood was treated by liquid‐liquid extraction and the analytes were separated by gradient elution on an Agilent Zorbax Extended‐C8 column (2.0×150 mm, 5 µm). Chlorpyrifos and chlorpyrifos‐oxon were detected in
Biomedical Chromatography, 2007
Time-of-flight (TOF) instruments have recently gained popularity in quantitative analyses. Normal... more Time-of-flight (TOF) instruments have recently gained popularity in quantitative analyses. Normally, TOF mass spectrometers are used for accurate mass measurements for empirical formula verification. However, over the past decade, they have been used quantitatively as well. Because of the fast separations and narrow peaks that result from gas chromatography separations, scanning mass spectrometers are not ideal detectors. TOF mass spectrometers, however, have the ability to collect spectra at a faster rate. Two-dimensional gas chromatography has also been introduced to further resolve peaks from complex matrices. Two-dimensional gas chromatography results in a faster separation as well as narrower peaks. This paper reviews the methods currently in the literature for the quantitation of compounds using one- and two-dimensional gas chromatography and TOF mass spectrometry detection.
Journal of Chromatography B, Nov 1, 2004
2 ,3-Dideoxycytidine (DDC) is a nucleoside reverse transcriptase inhibitor that has been shown to... more 2 ,3-Dideoxycytidine (DDC) is a nucleoside reverse transcriptase inhibitor that has been shown to inhibit the human immunodeficiency virus (HIV). DDC is a candidate for treatment of pregnant women to prevent prenatal transmission of HIV/AIDS to their unborn children. A quick and simple high-performance liquid chromatography (HPLC) method has been developed and validated for the determination of DDC concentrations in samples collected from a pregnant rat model (maternal plasma, amniotic fluid, placental and fetal tissues). Extraction of DDC and its internal standard 2 ,3-dideoxy-3-thiacytidine (3TC) in plasma and amniotic fluid was carried out by protein precipitation. Extraction from placental and fetal homogenates was achieved by solid phase extraction using Waters Oasis HLB solid phase extraction cartridges. Chromatographic separation was achieved on a Waters Spherisorb S3W silica column (4.6 mm × 100 mm) equipped with a Phenomenex guard column. The mobile phase used was 10% methanol in water with 22 mM formic acid. The flow rate was 0.5 ml/min, and the detection wavelength was optimized at 275 nm. Under these chromatographic conditions, DDC eluted around 12 min, and 3TC eluted around 10 min. The calibration curves for each day of validation and analysis showed good linear response through the range of 0.15-75.0 g/ml in each of the four matrices. The relative recovery for DDC in each of the matrices ranged from 87.8% to 103.0%. Acceptable intra-and inter-day assay precision (<15% R.S.D.) and accuracy (<15% error) were observed over 0.15-75.0 g/ml for all four matrices.
Biomedical Chromatography, 2007
Over the last decade, time‐of‐flight (TOF) instruments have increasingly been used as quantitatio... more Over the last decade, time‐of‐flight (TOF) instruments have increasingly been used as quantitation tools. In addition, because of their high resolving power, they can be used for verification of empirical formulas. Historically, TOF instruments have had limited quantitation capabilities because of their narrow dynamic range. However, recent advances have improved these limitations. This review covers the rationale for using TOF for LC detection, and describes the many methods currently in the literature for the quantitation of pharmaceuticals, environmental pollutants, explosives and many phytochemicals. Copyright © 2007 John Wiley & Sons, Ltd.
Humans can be exposed to central nervous system agents environmentally and/or therapeutically. Ch... more Humans can be exposed to central nervous system agents environmentally and/or therapeutically. Chronic exposure to these agents can cause damage to the central nervous system. In order to study the neurological effects of these agents, sensitive and selective analytical methods must be developed and validated in biological matrices, such as brain tissue, plasma, and whole blood. Chlorpyrifos (CPF) is an organophosphate pesticide that inhibits acetylcholinesterase, which is an enzyme that is necessary for normal function of the nervous system. CPF is metabolized by cytochrome P450 into chlorpyrifos-oxon (CPF-O) and TCP. CPF-O is about 3,000 times more potent than CPF in its inhibition of acetylcholinesterase activity, while TCP is non-toxic and is eliminated by the kidneys. Therapeutically, chronic exposure to antipsychotic drugs can result in abnormalities in motor function. Additionally, it is important to determine the extent to which a correlation exists between plasma and brain levels of these agents and cognitive function. Chapter 1 is the introduction and literature review that describes the layout of the dissertation and reviews the literature for analytical methods using liquid chromatography with time-of-flight mass spectrometry (TOF-MS) for quantitation. Chapter 2 reviews the literature for analytical methods using gas chromatography and TOF-MS v ACKNOWLEDGEMENTS I would first like to thank God for bringing me this far and for blessing me with an abundance of loving and supportive family members and friends. I thank my parents, James and Starletta Williamson, my brother James, and everyone else in my family that has continued to support me during my academic career. I love you all and am so grateful to have you in my life. I would like to thank my graduate advisor, Dr. Michael Bartlett. It has been an excellent experience learning and working under your direction. I feel extremely prepared for a career in analytical chemistry; whether it be in academia or industry. I would also like to thank my committee members, Drs. Beach, Capomacchia, Orlando and White. I owe a special thanks to Dr. Capomacchia for providing me with the opportunity to study here at UGA with the Bridges to Doctorate grant. I thank my friends from home and my new friends I have made during my time here at UGA. Thank you for being there for me if I needed to vent, needed to pull an allnighter studying, wanted you to try a new recipe I found, or if I just wanted to chat. I would like to thank my past and present labmates for their training and incite; Amy, Shonetta, Guodong, Yongzhen, and Meng. I could not have done this alone, thank you all for your support. vi
Journal of Liquid Chromatography & Related Technologies, Sep 30, 2008
Abstract Tandem mass spectrometry (MS/MS) is frequently used for the detection of small molecules... more Abstract Tandem mass spectrometry (MS/MS) is frequently used for the detection of small molecules in biological samples because of the high specificity and sensitivity associated with these instruments. Time-of-flight mass spectrometers (TOF-MS) are typically used ...
Journal of Pharmacology and Experimental Therapeutics, 2007
Persistent behavioral abnormalities have been commonly associated with acute organophosphate (OP)... more Persistent behavioral abnormalities have been commonly associated with acute organophosphate (OP) pesticide poisoning; however, relatively little is known about the consequences of chronic OP exposures that are not associated with acute cholinergic symptoms. In this study, the behavioral and neurochemical effects of chronic, intermittent, and subthreshold exposures to the OP pesticide, chlorpyrifos (CPF), were investigated. Rats were injected with CPF s.c. (dose range, 2.5-18.0 mg/kg) every other day over the course of 30 days and then were given a 2-week CPF-free washout period. In behavioral experiments conducted during the washout period, dosedependent decrements in a water-maze hidden platform task and a prepulse inhibition procedure were observed, without significant effects on open-field activity, Rotorod performance, grip strength, or a spontaneous novel object recognition task. After washout, levels of CPF and its metabolite 3,5,6-trichloro-2-pyridinol were minimal in plasma and brain; however, cholinesterase inhibition was still detectable. Furthermore, the 18.0 mg/kg dose of CPF was associated with (brain region-dependent) decreases in nerve growth factor receptors and cholinergic proteins including the vesicular acetylcholine transporter, the high-affinity choline transporter, and the ␣ 7-nicotinic acetylcholine receptor. These deficits were accompanied by decreases in anterograde and retrograde axonal transport measured in sciatic nerves ex vivo. Thus, low-level (intermittent) exposure to CPF has persistent effects on neurotrophin receptors and cholinergic proteins, possibly through inhibition of fast axonal transport. Such neurochemical changes may lead to deficits in information processing and cognitive function. This work was supported by the National Institute of Environmental Health Sciences (ES012241 to A.V.T.). Salary support (to J.J.B.) also was contributed through a Merit Review Award from the Veterans Administration. Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
Experiments using inductively coupled plasma-mass spectrometry (ICP-MS) as a detection system for... more Experiments using inductively coupled plasma-mass spectrometry (ICP-MS) as a detection system for reversed-phase chromatographic methods have been conducted for the analyses of various pharmaceutical compounds. Interfacing the ICP-MS instrument to the high performance liquid chromatography (HPLC) system was straightforward. The eluent from the HPLC column was pumped through a photodiode array (PDA) and then directly into the ICP-MS nebulizer using standard sample introduction components at flow rates ranging from 0.5 to 1.5 mL/min. Two separate sets of experiments were carried out. Initial speciation experiments were conducted on the B supplements cyanocobalamin (vitamin B 12 ), biotin, and thiamine (vitamin B 1 ) using isocratic separations. ICP-MS chromatograms were generated by monitoring cobalt at m/z 59, phosphorusatm/z31,andsulfuratm/z34simultaneously. Linearregression data were obtained for cyanocobalamin, thiamine hydrochloride, and biotin. Elemental detection was used to re...
Bioanalysis, 2014
The US FDA published A Guidance for Industry: Bioanalytical Method Validation in May 2001. Despit... more The US FDA published A Guidance for Industry: Bioanalytical Method Validation in May 2001. Despite the publication of the guidance, companies continue to submit bioequivalence studies with bioanalytical deficiencies that preclude Abbreviated New Drug Application approval. The Divisions of Bioequivalence in the FDA's Office of Generic Drugs conducted a survey of the bioequivalence submissions over a 10-year period (2001-2011) to identify the most commonly occurring bioanalytical deficiencies. Data from a total of 4028 Abbreviated New Drug Application submissions were collected to identify bioanalytical deficiencies. Of the three categories of bioanalytical deficiencies (method, validation and report), the majority of the deficiencies were from the bioanalytical method validation section. Globally, the percentage of bioanalytical method validation deficiencies was 62%. The approval of generic drugs would be accelerated if these deficiencies were avoided by generic companies by adh...
Journal of Liquid Chromatography & Related Technologies, 2007
A method has been developed to quantify chlorpyrifos (O,O‐diethyl O‐[3,5,6,‐trichloro‐2‐pyridyl] ... more A method has been developed to quantify chlorpyrifos (O,O‐diethyl O‐[3,5,6,‐trichloro‐2‐pyridyl] phosphorothionate) and its metabolites chlorpyrifos‐oxon (O,O‐diethyl‐O[3,5,6,trichloro‐2‐pyridyl] phosphate) and TCP (3,5,6,‐trichloro‐2‐pyridinol) in rat blood by liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI‐MS/MS). Rat blood was treated by liquid‐liquid extraction and the analytes were separated by gradient elution on an Agilent Zorbax Extended‐C8 column (2.0×150 mm, 5 µm). Chlorpyrifos and chlorpyrifos‐oxon were detected in
Journal of Chromatography B, 2004
2 ,3-Dideoxycytidine (DDC) is a nucleoside reverse transcriptase inhibitor that has been shown to... more 2 ,3-Dideoxycytidine (DDC) is a nucleoside reverse transcriptase inhibitor that has been shown to inhibit the human immunodeficiency virus (HIV). DDC is a candidate for treatment of pregnant women to prevent prenatal transmission of HIV/AIDS to their unborn children. A quick and simple high-performance liquid chromatography (HPLC) method has been developed and validated for the determination of DDC concentrations in samples collected from a pregnant rat model (maternal plasma, amniotic fluid, placental and fetal tissues). Extraction of DDC and its internal standard 2 ,3-dideoxy-3-thiacytidine (3TC) in plasma and amniotic fluid was carried out by protein precipitation. Extraction from placental and fetal homogenates was achieved by solid phase extraction using Waters Oasis HLB solid phase extraction cartridges. Chromatographic separation was achieved on a Waters Spherisorb S3W silica column (4.6 mm × 100 mm) equipped with a Phenomenex guard column. The mobile phase used was 10% methanol in water with 22 mM formic acid. The flow rate was 0.5 ml/min, and the detection wavelength was optimized at 275 nm. Under these chromatographic conditions, DDC eluted around 12 min, and 3TC eluted around 10 min. The calibration curves for each day of validation and analysis showed good linear response through the range of 0.15-75.0 g/ml in each of the four matrices. The relative recovery for DDC in each of the matrices ranged from 87.8% to 103.0%. Acceptable intra-and inter-day assay precision (<15% R.S.D.) and accuracy (<15% error) were observed over 0.15-75.0 g/ml for all four matrices.
Humans can be exposed to central nervous system agents environmentally and/or therapeutically. Ch... more Humans can be exposed to central nervous system agents environmentally and/or therapeutically. Chronic exposure to these agents can cause damage to the central nervous system. In order to study the neurological effects of these agents, sensitive and selective ...
Biomedical Chromatography, 2007
Time-of-flight (TOF) instruments have recently gained popularity in quantitative analyses. Normal... more Time-of-flight (TOF) instruments have recently gained popularity in quantitative analyses. Normally, TOF mass spectrometers are used for accurate mass measurements for empirical formula verification. However, over the past decade, they have been used quantitatively as well. Because of the fast separations and narrow peaks that result from gas chromatography separations, scanning mass spectrometers are not ideal detectors. TOF mass spectrometers, however, have the ability to collect spectra at a faster rate. Two-dimensional gas chromatography has also been introduced to further resolve peaks from complex matrices. Two-dimensional gas chromatography results in a faster separation as well as narrower peaks. This paper reviews the methods currently in the literature for the quantitation of compounds using one- and two-dimensional gas chromatography and TOF mass spectrometry detection.
Biomedical Chromatography, 2007
Over the last decade, time-of-flight (TOF) instruments have increasingly been used as quantitatio... more Over the last decade, time-of-flight (TOF) instruments have increasingly been used as quantitation tools. In addition, because of their high resolving power, they can be used for verification of empirical formulas. Historically, TOF instruments have had limited quantitation capabilities because of their narrow dynamic range. However, recent advances have improved these limitations. This review covers the rationale for using TOF for LC detection, and describes the many methods currently in the literature for the quantitation of pharmaceuticals, environmental pollutants, explosives and many phytochemicals.
Journal of Liquid Chromatography & Related Technologies, 2008
Abstract Tandem mass spectrometry (MS/MS) is frequently used for the detection of small molecules... more Abstract Tandem mass spectrometry (MS/MS) is frequently used for the detection of small molecules in biological samples because of the high specificity and sensitivity associated with these instruments. Time-of-flight mass spectrometers (TOF-MS) are typically used ...