Comparison of plasma, venous and capillary blood levels of piperaquine in patients with uncomplicated falciparum malaria

PURPOSE: Dihydroartemisinin-piperaquine (DP) is a fixed-dose artemisinin-based combination treatment. Field pharmacokinetic studies would be simplified and facilitated by being able to use small volume capillary assays rather than venous blood. The aim of this study was to describe the relationship between piperaquine concentrations measured in capillary blood, venous blood and venous plasma. METHODS: Samples of plasma, whole blood obtained by venesection and capillary blood were taken simultaneously from patients with uncomplicated Plasmodium falciparum malaria treated with DP between 0 and 9 weeks after treatment. Piperaquine concentrations in venous and capillary samples were measured using solid phase extraction and analysis by liquid chromatography with ultraviolet detection. RESULTS: A total of 161 sets of the three measures were obtained from 54 patients. Piperaquine concentrations in the venous blood samples were approximately twofold higher and those in the capillary blood samples were threefold higher than the corresponding venous plasma concentrations. Capillary blood piperaquine concentrations were approximately 1.7-fold higher than venous blood concentrations, and this difference also increased with time. CONCLUSION: Differences in whole blood and plasma levels of piperaquine suggest compartmentalisation of the drug within blood cells, as also occurs with the structurally related quinoline chloroquine. The relationship between piperaquine concentrations in the venous plasma, venous blood and capillary blood is variable and unpredictable at low concentrations. However, within the range of concentrations usually present in patients between 3 and 21 days after treatment with currently recommended doses, the relationship between capillary and venous whole blood is predictable; consequently, capillary blood sampling can be used in field assessments

Sentinel network for monitoring in vitro susceptibility of Plasmodium falciparum to antimalarial drugs in Colombia: a proof of concept

Drug resistance is one of the principal obstacles blocking worldwide malaria control. In Colombia, malaria remains a major public health concern and drug-resistant parasites have been reported. In vitro drug susceptibility assays are a useful tool for monitoring the emergence and spread of drug-resistant Plasmodium falciparum. The present study was conducted as a proof of concept for an antimalarial drug resistance surveillance network based on in vitro susceptibility testing in Colombia. Sentinel laboratories were set up in three malaria endemic areas. The enzyme linked immunosorbent assay-histidine rich protein 2 and schizont maturation methods were used to assess the susceptibility of fresh P. falciparum isolates to six antimalarial drugs. This study demonstrates that an antimalarial drug resistance surveillance network based on in vitro methods is feasible in the field with the participation of a research institute, local health institutions and universities. It could also serve as a model for a regional surveillance network. Preliminary susceptibility results showed widespread chloroquine resistance, which was consistent with previous reports for the Pacific region. However, high susceptibility to dihydroartemisinin and lumefantrine compounds, currently used for treatment in the country, was also reported. The implementation process identified critical points and opportunities for the improvement of network sustainability strategies.PAHO [057-1-3144141]; COLCIENCIAS [ID 2229-405-20319]info:eu-repo/semantics/publishedVersio

Development and validation of a liquid chromatographic-tandem mass spectrometric method for determination of piperaquine in plasma stable isotope labeled internal standard does not always compensate for matrix effects.

A bioanalytical method for the analysis of piperaquine in human plasma using off-line solid-phase extraction and liquid chromatography coupled to positive tandem mass spectroscopy has been developed and validated. It was found that a mobile phase with high pH (i.e. 10) led to better sensitivity than mobile phase combinations with low pH (i.e. 2.5-4.5) despite the use of positive electrospray and a basic analyte. The method was validated according to published FDA guidelines and showed excellent performance. The within-day and between-day precisions expressed as R.S.D., were lower than 7% at all tested concentrations (4.5, 20, 400 and 500ng/mL) and below 10% at the lower limit of quantification (LLOQ) (1.5ng/mL). The calibration range was 1.5-500ng/mL with a limit of detection (LOD) at 0.38ng/mL. Validation of over-curve samples ensured that it would be possible with dilution if samples went outside the calibration range. Matrix effects were thoroughly evaluated both graphically and quantitatively. Matrix effects originating from the sample clean-up (i.e. solid-phase extraction) procedure rather than the plasma background were responsible for the ion suppression seen in this study. Salts remaining from the buffers used in the solid-phase extraction suppressed the signals for both piperaquine and its deuterated internal standard. This had no effect on the quantification of piperaquine. Triethylamine residues remaining after evaporation of the solid-phase extraction eluate were found to suppress the signals for piperaquine and its deuterated internal standard differently. It was found that this could lead to an underestimation of the true concentration with 50% despite the use of a deuterated internal standard

Development and validation of a solid-phase extraction-liquid chromatographic method for determination of amoxicillin in plasma.

A bioanalytic method for the determination of amoxicillin in plasma by hydrophilic interaction solid-phase extraction and liquid chromatography has been developed and validated. Plasma was precipitated with acetonitrile before samples were loaded onto a zwitterionic hydrophilic interaction liquid chromatography (ZIC-HILIC) solid-phase extraction column. Amoxicillin was analyzed by liquid chromatography on an Aquasil (150 x 4.6 mm) LC column with mobile-phase acetonitrile: phosphate buffer (pH 2.5; 0.1 mol/L) (7:93, v/v) and UV detection at 230 nm. A regression model using 1/concentration weighting was found the most appropriate for quantification. The intraassay precision for plasma was 3.3% at 15.0 microg/mL and 10.9% at 0.200 microg/mL. The interassay precision for plasma was 1.8% at 15.0 microg/mL and 7.5% at 0.200 microg/mL. The total-assay precision for plasma over 4 days using a total of 20 replicates was 13.2%, 5.5%, and 3.8% at 0.200 microg/mL, 3.00 microg/mL, and 15.0 microg/mL, respectively. The lower limit of quantification and the limit of detection were 0.050 microg/mL and 0.025 microg/mL, respectively, for 100 microL plasma. Long-term storage stability studies of amoxicillin in plasma indicate that a temperature of -80 degrees C is necessary to prevent degradation of amoxicillin

High throughput assay for the determination of lumefantrine in plasma.

A high throughput bioanalytical assay for the determination of lumefantrine in plasma has been developed and validated extensively. The within-day precisions for lumefantrine were 5.2, 3.5 and 2.5% at 200, 2000 and 15000 ng/mL, respectively. The between-day precisions were 4.0, 2.8 and 3.1% at 200, 2000 and 15000 ng/mL, respectively. The lower limits of quantification (LLOQ) and the limits of detection (LOD) were 25 and 10 ng/mL, respectively using 0.250 mL plasma. The average recovery of lumefantrine was 85% and independent upon concentration. The use of 96-well plate format and short chromatographic run has increased the daily sample throughput four times. The assay is particularly suitable for large therapeutic drug monitoring studies using day 7 sampling

High throughput assay for the determination of lumefantrine in plasma.

A high throughput bioanalytical assay for the determination of lumefantrine in plasma has been developed and validated extensively. The within-day precisions for lumefantrine were 5.2, 3.5 and 2.5% at 200, 2000 and 15000 ng/mL, respectively. The between-day precisions were 4.0, 2.8 and 3.1% at 200, 2000 and 15000 ng/mL, respectively. The lower limits of quantification (LLOQ) and the limits of detection (LOD) were 25 and 10 ng/mL, respectively using 0.250 mL plasma. The average recovery of lumefantrine was 85% and independent upon concentration. The use of 96-well plate format and short chromatographic run has increased the daily sample throughput four times. The assay is particularly suitable for large therapeutic drug monitoring studies using day 7 sampling

Development and validation of an automated solid-phase extraction and liquid chromatographic method for determination of lumefantrine in capillary blood on sampling paper.

A bioanalytical method for the determination of lumefantrine in 100 microl blood applied onto sampling paper, by solid-phase extraction and liquid chromatography, has been developed and validated. Whatman 31 ET Chr sampling paper was pre-treated with 0.75 M tartaric acid before sampling capillary blood to enable a high recovery of lumefantrine. Lumefantrine was extracted from the sampling paper, then further purified using solid-phase extraction and finally quantified with HPLC. The between-day variation was below 10% over the range 0.4-25 microM. The lower limit of quantification was 0.25 microM in 100 microl capillary blood. No decrease in lumefantrine concentration in dried blood spot is seen after 4 months storage at 22 degrees C. The method was also evaluated in field samples from patients in Tanzania after treatment with lumefantrine/artemether. Lumefantrine could be estimated accurately enough to assess bioavailability and treatment compliance on day 7 (i.e. 4 days after the last dose) after a standard regimen with the lumefantrine/artemether combination

Development and validation of a bioanalytical method using automated solid-phase extraction and LC-UV for the simultaneous determination of lumefantrine and its desbutyl metabolite in plasma.

A bioanalytical method for the determination of lumefantrine (LF) and its metabolite desbutyl-lumefantrine (DLF) in plasma by solid-phase extraction (SPE) and liquid chromatography has been developed. Plasma proteins were precipitated with acetonitrile:acetic acid (99:1, v/v) containing a DLF analogue internal standard before being loaded onto a octylsilica (3 M Empore) SPE column. Two different DLF analogues were evaluated as internal standards. The compounds were analysed by liquid chromatography UV detection on a SB-CN (250 mm x 4.6 mm) column with a mobile phase containing acetonitrile-sodium phosphate buffer pH (2.0; 0.1 M) (55:45, v/v) and sodium perchlorate 0.05 M. Different SPE columns were evaluated during method development to optimise reproducibility and recovery for LF, DLF and the two different DLF analogues. The within-day precisions for LF were 6.6 and 2.1% at 0.042 and 8.02 microg/mL, respectively, and for DLF 4.5 and 1.5% at 0.039 and 0.777 microg/mL, respectively. The between-day precisions for LF were 12.0 and 2.9% at 0.042 and 8.02 microg/mL, respectively, while for DLF 0.7 and 1.2% at 0.039 and 0.777 microg/mL, respectively. The limit of quantification was 0.024 and 0.021 microg/mL for LF and DLF, respectively. Different amounts of lipids in plasma did not affect the absolute recovery of LF or DLF

How much fat is necessary to optimize lumefantrine oral bioavailability?

BACKGROUND: Artemether-lumefantrine (AL) is the only fixed, artemisinin-based combination antimalarial drug which is registered internationally and deployed on a large scale. Absorption of the hydrophobic lipophilic lumefantrine component varies widely between individuals and is greatly increased by fat coadministration; but patients with acute malaria are frequently nauseated and anorexic, making dietary advice difficult to comply with. The aim of this study was to describe the dose-response relationship between coadministration of fat and relative lumefantrine bioavailability, in order to determine the minimum amount of fat necessary to optimize absorption. METHOD: We conducted a multiple crossover pharmacokinetic study in 12 healthy volunteers. This compared the area under the plasma concentration-time curve (AUC) for lumefantrine after administration of a single dose of AL in the fasting state given with 0, 10, 40, 150 and 500 ml of soya milk corresponding to 0, 0.32, 1.28, 4.8 and 16 g of fat. All volumes of milk supplements were tested in all subjects with a 3- to 4-week washout period in-between. RESULTS: A dose-response relationship was demonstrated between the volume of soya milk administered and lumefantrine bioavailability. AL administration with soya milk increased the lumefantrine AUC more than five fold. The population mean estimated volume of soya milk required to obtain 90% of maximum effect (in terms of lumefantrine AUC) was 36 ml (corresponding to 1.2 g of fat). CONCLUSIONS: Coadministration of artemether-lumefantrine with a relatively small amount of fat (as soya milk) was required to ensure maximum absorption of lumefantrine in healthy adult volunteers

Development and validation of an automated solid phase extraction and liquid chromatographic method for the determination of piperaquine in urine.

A sensitive and specific bioanalytical method for determination of piperaquine in urine by automated solid-phase extraction (SPE) and liquid chromatography (LC) has been developed and validated. Buffered urine samples (containing internal standard) were loaded onto mixed phase (cation-exchange and octylsilica) SPE columns using an ASPEC XL SPE robot. Chromatographic separation was achieved on a Chromolith Performance RP-18e (100 mm x 4.6 mm I.D.) LC column with phosphate buffer (pH 2.5; 0.1 mol/L)-acetonitrile (92:8, v/v). Piperaquine was analysed at a flow rate of 3 mL/min with UV detection at 347 nm. A linear regression model on log-log transformed data was used for quantification. Within-day precision for piperaquine was 1.3% at 5000 ng/mL and 6.6% at 50 ng/mL. Between-day precision for piperaquine was 3.7% at 5000 ng/mL and 7.2% at 50 ng/mL. Total-assay precision for piperaquine over 4 days using five replicates each day (n = 20) was 4.0%, 5.2% and 9.8% at 5000, 500 and 50 ng/mL, respectively. The lower limit of quantification (LLOQ) was set to 3 ng/mL using 1 mL of urine, which could be lowered to 0.33 ng/mL when using 9 mL of urine and an increased injection volume