A liquid chromatography with tandem mass spectrometry（LC-MS/MS） method has been developed and validated for the measurement of sunitinib in rabbit plasma. After protein precipitation with acetonitrile, samples were ...A liquid chromatography with tandem mass spectrometry（LC-MS/MS） method has been developed and validated for the measurement of sunitinib in rabbit plasma. After protein precipitation with acetonitrile, samples were analyzed on a Zorbax Extend-C18 column（150 mm×4.6 mm, 5μm）. The mobile phase consisted of a mixture of acetonitrile and deionized water（containing 0.05% formic acid） at a ratio of 27：73（v/v）, and the flow rate was set at 0.8 mL /min. The column temperature was maintained at 30 oC. The LC eluate was detected by an electrospray ionization（ESI） source operated in the positive ion mode, and quantification was conducted using MRM of the transitions m/z 399.24→283.01 and m/z 415.19→178.00 for sunitinib and internal standard（IS, diltiazem hydrochloride）, respectively. The calibration curve was linear in the range of 2–600 ng/m L. The lower limit of quantification was 2 ng/mL. The method also exhibited satisfactory results in terms of sensitivity, specificity, accuracy（with relative error ranging from –4.0% to 1.1%）, precision（with intra- and inter-day relative standard deviations ranging from 2.8% to 9.5%）, matrix effect, recovery as well as stability. Taken together, our newly developed method was reliable to monitor sunitinib concentrations in rabbit plasma.展开更多
Lamivudine has been widely used in the treatment of HIV disease. A reliable, sensitive reversed phase high performance liquid chromatography （RP-HPLC） method was developed and validated for lamivudine in rabbit plas...Lamivudine has been widely used in the treatment of HIV disease. A reliable, sensitive reversed phase high performance liquid chromatography （RP-HPLC） method was developed and validated for lamivudine in rabbit plasma. The method was developed on Hypersil BDS C-18 column （250 mm ＊ 4.6 mm, 5 μm） using a mobile phase of 0.25% Triethylamine buffer （pH 3.0）： acetonitrile （70：30, v/v）. The efficient was monitored by UV detector at 256 nm. The total run time was 15 min with a flow rate of 1.0 mL/min. Calibration curve was linear over the concentration range of 25-2000 ng/mL. The retention times of lamivudine and internal standard （Nelfinavir） were 8.78 min and 10.86 min, respectively. The developed RP-HPLC method can be successfully applied for the quantitative pharmacokinetic parameters determination of lamivudine in rabbit model.展开更多
In the present study, free fatty acids(FFAs, including palmitic acid, stearic acid, oleic acid, linoleic acid and arachidonic acid) in rabbit plasma were determined by gas chromatography-mass spectrometry(GC-MS) after...In the present study, free fatty acids(FFAs, including palmitic acid, stearic acid, oleic acid, linoleic acid and arachidonic acid) in rabbit plasma were determined by gas chromatography-mass spectrometry(GC-MS) after trimethylsilylation derivatization using N,O-bis(trimethylsilyl)trifluoroacetamide(BSTFA) – trimethylchlorosilane(TMCS) as derivatization reagent. The experimental conditions, including extraction and silylation reaction, were investigated. The method was experimentally validated. The linearity between fatty acids’ peak areas and their concentrations was obtained with the corelative coefficient(r2) all more than 0.999, and the recoveries were between 82% and 111%. The intra-day variations of FFAs’ in plasma samples at different concentrations were all less than 6%. FFA analysis results of 16 rabbit plasma samples showed that the method could be well applied in the determination of plasma samples in vivo. In contrast to the traditional method of FFA derivatization, the established trimethylsilylation method presented simplicity, high specificity, and completely free from the interference of the esterified fatty acid, such as triacylglyceride. The method could be applied for analyzing FFA profiles in the clinical laboratory or pharmacological research.展开更多
In the present study, we developed and validated a simple and sensitive gradient elution liquid chromatography tandem mass spectrometry （LC-MS/MS） method for quantification of doxorubicin in rabbit plasma. Daunorubi...In the present study, we developed and validated a simple and sensitive gradient elution liquid chromatography tandem mass spectrometry （LC-MS/MS） method for quantification of doxorubicin in rabbit plasma. Daunorubicin was used as an internal standard （IS）. The doxorubicin and IS were extracted with ethyl acetate from plasma samples. The chromatographic separations were achieved on a C18 column （2.1 mm×50 mm, 2.5μm） configured with a C18 guard column （2.1 mm×10 mm, 2.5 μm）. The mobile phase of 0.1% formic acid-water solution and acetonitrile was delivered using a gradient elution program at a flow rate of 0.4 mL/min. The temperature for column was maintained at 40 ℃. The electrospray ionization （ESI） source was operated in the positive ion mode, and the quantification was conducted using multiple reaction monitoring （MRM） of the transitions m/z 544.07→396.96 and m/z 528.06→321.05 for doxorubicin and IS, respectively. The calibration curve of doxorubicin was linear （r 〉 0.999） within the range of 2-600 ng/mL. The lower limit of quantification was 2 ng/mL. The relative errors of intra-day and inter-day accuracies ranged from -2.48% to 0.18% and from -3.78% to 1.94%, respectively. The relative standard deviations of intra-day and inter-day precisions were less than 8.65% and 6.41%, respectively. The method exhibited satisfactory results in terms of specificity, sensitivity, matrix effect, recovery and stability. The newly developed LC-MS/MS method was reliable to monitor doxorubicin concentrations in rabbit plasma.展开更多
A specific, precise and accurate ion-pair HPLC-UV method has been developed and validated for simultaneous determination of phosphocreatine (PCr), and its metabolite creatine (Cr) as well as related ATP in plasma and ...A specific, precise and accurate ion-pair HPLC-UV method has been developed and validated for simultaneous determination of phosphocreatine (PCr), and its metabolite creatine (Cr) as well as related ATP in plasma and red blood cell (RBC) of rabbits. After addition of TMP as IS, the samples were deproteinized with 6% PCA. The analytes were separated on a Kromasil C18 column using a tertiary gradient mobile phase composed of buffer A (0.2% KH2PO4 + 0.08% tetrabutyl ammonium hydrogen sulphate, pH 3.0), buffer B (buffer A adjusted to pH 7.5 with 1 mol/L NaOH) and MeOH. Detection wavelengths were set at 210 nm for PCr and Cr and 260 nm for ATP and TMP. Some blank samples were initially run for baseline subtraction. The linear detection responses were obtained for PCr concentration over a range of 10 - 7500 mg/ml (plasma) and 5 - 2500 mg/ml (RBC) and for both Cr and ATP concentrations of 10 - 1500 mg/ml (plasma) and 5 - 750 mg/ml (RBC) (r > 0.99). The QC samples of 3 analytes showed intra-day and inter-day precisions (RSD) of - 107%. The method was successfully used to simultaneously determine plasma and RBC concentrations of the 3 analytes and to study pharmacokinetics after iv administration of PCr to rabbits.展开更多
Sterols are essential components of the cell membrane lipid bilayer that include molecules such as cholesterol and desmosterol, which are significantly found in the spermatozoa of several animal species. However, the ...Sterols are essential components of the cell membrane lipid bilayer that include molecules such as cholesterol and desmosterol, which are significantly found in the spermatozoa of several animal species. However, the presence of desmosterol in rabbit semen has never been investigated. The aims of this study were to characterize the sterol composition of subfractions of ejaculated rabbit semen and evaluate the in vitro effects of sterol on the spermatozoa acrosome reaction and motility. Two sterols, occurring prevalently in the free form （94.3%）, were identified in whole semen collected from 10 fertile New Zealand White rabbits, specifically desmosterol （58.5% of total sterols） and cholesterol （35.9% of total sterols）. Desmosterol was the predominant sterol found in all subfractions of rabbit semen, varying from 56.7% （in the prostatic secretory granules, PSGs） to 63.8% （in the seminal plasma）. Spermatozoa contained an intermediate proportion of desmosterol （59.8%）, which was asymmetrically distributed between the heads （52.0% of the total content of sterols） and the tails （81.8%）. Results showed that both desmosterol and cholesterol can be transferred from the PSGs to the spermatozoa and are equally effective in inhibiting in vitro spermatozoa capacitation at a concentration higher than 1 mg L^-1. In contrast, neither desmosterol nor cholesterol had a significant effect on spermatozoa motility. Thus, it was concluded that, the various fractions of rabbit seminal fluid differ from each other in sterol composition and quantity, probably due to their different functional properties, and these fractions may undergo significant sterol changes depending on the stage of spermatozoa capacitation.展开更多
It is suggested that the difference in body size between domestic-type rabbits and small pet-type rabbits results in different nutrient requirements. The nutritional requirements of pet rabbits have been assessed, alt...It is suggested that the difference in body size between domestic-type rabbits and small pet-type rabbits results in different nutrient requirements. The nutritional requirements of pet rabbits have been assessed, although such assessments require evaluation throughout the rabbit life span. The present study was conducted under a cecotrophy prevention program with young and adult rabbits. Six male Dutch rabbits were housed individually in a dormitory-type cage, and they were randomly fed graded levels of dietary methionine and threonine, at ratios of 4:0, 3:1, 2:2, 1:3, and 0:4 mixed with two types of feed, 4:0 and 0:4. Four days after switching diets and 4 hrs after starting morning feeding, approximately one milliliter of blood was collected from the vein of the ear. Free amino acid concentrations in the plasma were determined with a high-speed amino acid analyzer. Plasma concentrations of methionine and threonine compared to dietary methionine and threonine levels are shown in young rabbits. The plasma concentration of methionine remained constant at a low level and then increased linearly. The intersection was estimated as 0.16 g/d. In the same manner, the intersection of the plasma threonine value was estimated as 0.27 g/d. These values were calculated as 0.27% and 0.47% of the diets, respectively. In the case of adult rabbits, the baseline was not obtained for dietary methionine and threonine levels. The methionine requirement was estimated as 0.11 g/d. The threonine requirement was estimated as 0.22 g/d. These values were calculated as 0.15% and 0.30% of the diets, respectively. In comparison with young and adult rabbits, both methionine and threonine showed a low baseline level in young rabbits, while their variations in plasma levels of adult rabbits were not determined. The requirement of both amino acids in young rabbits is higher than that in adult rabbits. The dietary values of both amino acids in young rabbits were also higher than those in adult rabbits. Small pet-type adult rabbits required lower amino acid levels than domestic-type rabbits.展开更多
文摘A liquid chromatography with tandem mass spectrometry（LC-MS/MS） method has been developed and validated for the measurement of sunitinib in rabbit plasma. After protein precipitation with acetonitrile, samples were analyzed on a Zorbax Extend-C18 column（150 mm×4.6 mm, 5μm）. The mobile phase consisted of a mixture of acetonitrile and deionized water（containing 0.05% formic acid） at a ratio of 27：73（v/v）, and the flow rate was set at 0.8 mL /min. The column temperature was maintained at 30 oC. The LC eluate was detected by an electrospray ionization（ESI） source operated in the positive ion mode, and quantification was conducted using MRM of the transitions m/z 399.24→283.01 and m/z 415.19→178.00 for sunitinib and internal standard（IS, diltiazem hydrochloride）, respectively. The calibration curve was linear in the range of 2–600 ng/m L. The lower limit of quantification was 2 ng/mL. The method also exhibited satisfactory results in terms of sensitivity, specificity, accuracy（with relative error ranging from –4.0% to 1.1%）, precision（with intra- and inter-day relative standard deviations ranging from 2.8% to 9.5%）, matrix effect, recovery as well as stability. Taken together, our newly developed method was reliable to monitor sunitinib concentrations in rabbit plasma.
文摘Lamivudine has been widely used in the treatment of HIV disease. A reliable, sensitive reversed phase high performance liquid chromatography （RP-HPLC） method was developed and validated for lamivudine in rabbit plasma. The method was developed on Hypersil BDS C-18 column （250 mm ＊ 4.6 mm, 5 μm） using a mobile phase of 0.25% Triethylamine buffer （pH 3.0）： acetonitrile （70：30, v/v）. The efficient was monitored by UV detector at 256 nm. The total run time was 15 min with a flow rate of 1.0 mL/min. Calibration curve was linear over the concentration range of 25-2000 ng/mL. The retention times of lamivudine and internal standard （Nelfinavir） were 8.78 min and 10.86 min, respectively. The developed RP-HPLC method can be successfully applied for the quantitative pharmacokinetic parameters determination of lamivudine in rabbit model.
基金National Natural Science Foundation of China (Grant No. 31671928)Natural Science Foundation of Shanghai (Grant No. 15ZR1440800)。
文摘In the present study, free fatty acids(FFAs, including palmitic acid, stearic acid, oleic acid, linoleic acid and arachidonic acid) in rabbit plasma were determined by gas chromatography-mass spectrometry(GC-MS) after trimethylsilylation derivatization using N,O-bis(trimethylsilyl)trifluoroacetamide(BSTFA) – trimethylchlorosilane(TMCS) as derivatization reagent. The experimental conditions, including extraction and silylation reaction, were investigated. The method was experimentally validated. The linearity between fatty acids’ peak areas and their concentrations was obtained with the corelative coefficient(r2) all more than 0.999, and the recoveries were between 82% and 111%. The intra-day variations of FFAs’ in plasma samples at different concentrations were all less than 6%. FFA analysis results of 16 rabbit plasma samples showed that the method could be well applied in the determination of plasma samples in vivo. In contrast to the traditional method of FFA derivatization, the established trimethylsilylation method presented simplicity, high specificity, and completely free from the interference of the esterified fatty acid, such as triacylglyceride. The method could be applied for analyzing FFA profiles in the clinical laboratory or pharmacological research.
基金National Natural Science Foundation of China（ Grant No.81571779）.
文摘In the present study, we developed and validated a simple and sensitive gradient elution liquid chromatography tandem mass spectrometry （LC-MS/MS） method for quantification of doxorubicin in rabbit plasma. Daunorubicin was used as an internal standard （IS）. The doxorubicin and IS were extracted with ethyl acetate from plasma samples. The chromatographic separations were achieved on a C18 column （2.1 mm×50 mm, 2.5μm） configured with a C18 guard column （2.1 mm×10 mm, 2.5 μm）. The mobile phase of 0.1% formic acid-water solution and acetonitrile was delivered using a gradient elution program at a flow rate of 0.4 mL/min. The temperature for column was maintained at 40 ℃. The electrospray ionization （ESI） source was operated in the positive ion mode, and the quantification was conducted using multiple reaction monitoring （MRM） of the transitions m/z 544.07→396.96 and m/z 528.06→321.05 for doxorubicin and IS, respectively. The calibration curve of doxorubicin was linear （r 〉 0.999） within the range of 2-600 ng/mL. The lower limit of quantification was 2 ng/mL. The relative errors of intra-day and inter-day accuracies ranged from -2.48% to 0.18% and from -3.78% to 1.94%, respectively. The relative standard deviations of intra-day and inter-day precisions were less than 8.65% and 6.41%, respectively. The method exhibited satisfactory results in terms of specificity, sensitivity, matrix effect, recovery and stability. The newly developed LC-MS/MS method was reliable to monitor doxorubicin concentrations in rabbit plasma.
文摘A specific, precise and accurate ion-pair HPLC-UV method has been developed and validated for simultaneous determination of phosphocreatine (PCr), and its metabolite creatine (Cr) as well as related ATP in plasma and red blood cell (RBC) of rabbits. After addition of TMP as IS, the samples were deproteinized with 6% PCA. The analytes were separated on a Kromasil C18 column using a tertiary gradient mobile phase composed of buffer A (0.2% KH2PO4 + 0.08% tetrabutyl ammonium hydrogen sulphate, pH 3.0), buffer B (buffer A adjusted to pH 7.5 with 1 mol/L NaOH) and MeOH. Detection wavelengths were set at 210 nm for PCr and Cr and 260 nm for ATP and TMP. Some blank samples were initially run for baseline subtraction. The linear detection responses were obtained for PCr concentration over a range of 10 - 7500 mg/ml (plasma) and 5 - 2500 mg/ml (RBC) and for both Cr and ATP concentrations of 10 - 1500 mg/ml (plasma) and 5 - 750 mg/ml (RBC) (r > 0.99). The QC samples of 3 analytes showed intra-day and inter-day precisions (RSD) of - 107%. The method was successfully used to simultaneously determine plasma and RBC concentrations of the 3 analytes and to study pharmacokinetics after iv administration of PCr to rabbits.
文摘Sterols are essential components of the cell membrane lipid bilayer that include molecules such as cholesterol and desmosterol, which are significantly found in the spermatozoa of several animal species. However, the presence of desmosterol in rabbit semen has never been investigated. The aims of this study were to characterize the sterol composition of subfractions of ejaculated rabbit semen and evaluate the in vitro effects of sterol on the spermatozoa acrosome reaction and motility. Two sterols, occurring prevalently in the free form （94.3%）, were identified in whole semen collected from 10 fertile New Zealand White rabbits, specifically desmosterol （58.5% of total sterols） and cholesterol （35.9% of total sterols）. Desmosterol was the predominant sterol found in all subfractions of rabbit semen, varying from 56.7% （in the prostatic secretory granules, PSGs） to 63.8% （in the seminal plasma）. Spermatozoa contained an intermediate proportion of desmosterol （59.8%）, which was asymmetrically distributed between the heads （52.0% of the total content of sterols） and the tails （81.8%）. Results showed that both desmosterol and cholesterol can be transferred from the PSGs to the spermatozoa and are equally effective in inhibiting in vitro spermatozoa capacitation at a concentration higher than 1 mg L^-1. In contrast, neither desmosterol nor cholesterol had a significant effect on spermatozoa motility. Thus, it was concluded that, the various fractions of rabbit seminal fluid differ from each other in sterol composition and quantity, probably due to their different functional properties, and these fractions may undergo significant sterol changes depending on the stage of spermatozoa capacitation.
文摘It is suggested that the difference in body size between domestic-type rabbits and small pet-type rabbits results in different nutrient requirements. The nutritional requirements of pet rabbits have been assessed, although such assessments require evaluation throughout the rabbit life span. The present study was conducted under a cecotrophy prevention program with young and adult rabbits. Six male Dutch rabbits were housed individually in a dormitory-type cage, and they were randomly fed graded levels of dietary methionine and threonine, at ratios of 4:0, 3:1, 2:2, 1:3, and 0:4 mixed with two types of feed, 4:0 and 0:4. Four days after switching diets and 4 hrs after starting morning feeding, approximately one milliliter of blood was collected from the vein of the ear. Free amino acid concentrations in the plasma were determined with a high-speed amino acid analyzer. Plasma concentrations of methionine and threonine compared to dietary methionine and threonine levels are shown in young rabbits. The plasma concentration of methionine remained constant at a low level and then increased linearly. The intersection was estimated as 0.16 g/d. In the same manner, the intersection of the plasma threonine value was estimated as 0.27 g/d. These values were calculated as 0.27% and 0.47% of the diets, respectively. In the case of adult rabbits, the baseline was not obtained for dietary methionine and threonine levels. The methionine requirement was estimated as 0.11 g/d. The threonine requirement was estimated as 0.22 g/d. These values were calculated as 0.15% and 0.30% of the diets, respectively. In comparison with young and adult rabbits, both methionine and threonine showed a low baseline level in young rabbits, while their variations in plasma levels of adult rabbits were not determined. The requirement of both amino acids in young rabbits is higher than that in adult rabbits. The dietary values of both amino acids in young rabbits were also higher than those in adult rabbits. Small pet-type adult rabbits required lower amino acid levels than domestic-type rabbits.
