Rapid determination of gemcitabine in plasma and serum using reversed‐phase HPLC

Abstract

Gemcitabine (2'2'-difluorodeoxycytidine) is a pyrimidine analog used in the treatment of a variety of solid tumors. After intravenous (i.v.) administration, it is rapidly inactivated to 2'-deoxy-2',2'-difluorouridine (dFdU). A sensitive analytical method for the quantitation of gemcitabine is required for the assessment of alternative dosage and treatment schemes. A rapid and robust RP-HPLC assay for analysis of gemcitabine in human and animal plasma and serum was developed and validated using 2'-deoxyuridine (dU) and 5-fluoro-2'-deoxyuridine (5FdU) as internal standards. It is based on protein precipitation, the use of an Atlantis dC18 column of 100 mm length (inner diameter, 4.6 mm; particle size, 3 microm) and isocratic elution using a 10 mM phosphate buffer, pH 3.0, followed by isocratic elution with the same buffer containing 3% of ACN. For gemcitabine, RSD values for intraday and interday precision were < 4.4 and 5.3%, respectively, the LOQ was 20 ng/mL, and the assay was linear in the range of 0.020-20 microg/mL with an accuracy of > or =89%. The recovery for gemcitabine, dU and 5FdU was 86-98%. The assay was applied to determine gemcitabine levels in plasma samples of patients collected during and shortly after conventional infusion of 25-30 mg/kg body mass (levels: 2.0-18.9 microg/mL) and rats that received lower doses (1.5 mg/kg) via i.v., subcutaneous and oral drug administration (levels: 0.20-2.60 microg/mL). It could also be applied to estimate dFdU levels in human plasma.

Combinatorial Pharmacologic Effects of Gemcitabine and its Metabolite dFdU

“…[12,13] Hertel and colleagues performed hydrolytic transformation of the cytosine ring of gemcitabine (1) into the uracil ring, using acetic acid in aqueous solution. A lengthy multistep purification procedure was required, resulting in a 19 % yield.…”

Development and validation of a stability indicating isocratic HPLC method for gemcitabine with application to drug release from poly lactic-co-glycolic acid nanoparticles and enzymatic degradation studies

“…Lanz et al . determined the LOD and LOQ of 10 ng/ml and 20 ng/ml, respectively, by using signal to noise ratio method [10]. While Xu et al .…”

“…Very few HPLC methods deal with the determination of gemcitabine in the presence of degradation products from forced degradation studies, and those that do are time‐consuming, with complicated mobile phases and gradient elution leading to base line shifting [9,10]. To date, digestive enzymatic degradation studies of gemcitabine have not been reported.…”

Gold nanoparticles and radiofrequency in experimental models for hepatocellular carcinoma

“…Isotype control AuNP-IgG-gemcitabine-mPEG (AIGP) nanoparticles were synthesized similarly by using isotype control human IgG in place of C225. Gemcitabine loading was determined by measuring unbound gemcitabine that was removed after filter centrifugation using reverse phase high performance liquid chromatography (HPLC) described elsewhere (14). Briefly, the UV absorbance of chromatographic peaks was recorded (Waters 1525 HPLC system, Milford, MA) at 275nm after separation through a Phenomenex Luna 5uC18(2) 100A column (250 x 4.6 mm, particle size 5 microns silica) using 0.1% TFA-water/acetonitrile at ambient temperature.…”