Lumefantrine Plasma Research Articles

Malaria prevalence, severity and treatment outcome in relation to day 7 lumefantrine plasma concentration in pregnant women.

BackgroundDay 7 plasma concentrations of lumefantrine (LF) can serve as a marker to predict malaria treatment outcome in different study populations. Two main cut-off points (175 and 280 ng/ml) are used to indicate plasma concentrations of LF, below which treatment failure is anticipated. However, there is limited data on the cumulative risk of recurrent parasitaemia (RP) in relation to day 7 LF plasma concentrations in pregnant women. This study describes the prevalence, severity, factors influencing treatment outcome of malaria in pregnancy and day 7 LF plasma concentration therapeutic cut-off points that predicts treatment outcome in pregnant women.MethodsThis was a one-arm prospective cohort study whereby 89 pregnant women with uncomplicated Plasmodium falciparum malaria receiving artemether-lumefantrine (ALu) participated in pharmacokinetics and pharmacodynamics study. Blood samples were collected on days 0, 2, 7, 14, 21 and 28 for malaria parasite quantification. LF plasma concentrations were determined on day 7. The primary outcome measure was an adequate clinical and parasitological response (ACPR) after treatment with ALu.ResultsThe prevalence of malaria in pregnant women was 8.1 % (95 % CI 6.85–9.35) of whom 3.4 % (95 % CI 1.49–8.51) had severe malaria. The overall PCR-uncorrected treatment failure rate was 11.7 % (95 % CI 0.54–13.46 %). Low baseline hemoglobin (<10 g/dl) and day 7 LF concentration <600 ng/ml were significant predictors of RP. The median day 7 LF concentration was significantly lower in pregnant women with RP (270 ng/ml) than those with ACPR (705 ng/ml) (p = 0.016). The relative risk of RP was 4.8 folds higher (p = 0.034) when cut-off of <280 ng/ml was compared to ≥280 ng/ml and 7.8-folds higher (p = 0.022) when cut-off of <600 ng/ml was compared to ≥600 ng/ml. The cut-off value of 175 ng/ml was not associated with the risk of RP (p = 0.399).ConclusionsPregnant women with day 7 LF concentration <600 ng/ml are at high risk of RP than those with ≥600 ng/ml. To achieve effective therapeutic outcome, higher day 7 venous plasma LF concentration ≥600 ng/ml is required for pregnant patients than the previously suggested cut-off value of 175 or 280 ng/ml for non-pregnant adult patients.

Bioanalytical Method Development and Validation for the Estimation of Lumefantrine and Desbutyl Lumefantrine in Human Plasma by LC-MS/MS

Bioanalytical Method Development and Validation for the Estimation of Lumefantrine and Desbutyl Lumefantrine in Human Plasma by LC-MS/MS

Artemether-lumefantrine treatment of uncomplicated Plasmodium falciparum malaria: a systematic review and meta-analysis of day 7 lumefantrine concentrations and therapeutic response using individual patient data.

BackgroundAchieving adequate antimalarial drug exposure is essential for curing malaria. Day 7 blood or plasma lumefantrine concentrations provide a simple measure of drug exposure that correlates well with artemether-lumefantrine efficacy. However, the ‘therapeutic’ day 7 lumefantrine concentration threshold needs to be defined better, particularly for important patient and parasite sub-populations.MethodsThe WorldWide Antimalarial Resistance Network (WWARN) conducted a large pooled analysis of individual pharmacokinetic-pharmacodynamic data from patients treated with artemether-lumefantrine for uncomplicated Plasmodium falciparum malaria, to define therapeutic day 7 lumefantrine concentrations and identify patient factors that substantially alter these concentrations. A systematic review of PubMed, Embase, Google Scholar, ClinicalTrials.gov and conference proceedings identified all relevant studies. Risk of bias in individual studies was evaluated based on study design, methodology and missing data.ResultsOf 31 studies identified through a systematic review, 26 studies were shared with WWARN and 21 studies with 2,787 patients were included. Recrudescence was associated with low day 7 lumefantrine concentrations (HR 1.59 (95 % CI 1.36 to 1.85) per halving of day 7 concentrations) and high baseline parasitemia (HR 1.87 (95 % CI 1.22 to 2.87) per 10-fold increase). Adjusted for mg/kg dose, day 7 concentrations were lowest in very young children (<3 years), among whom underweight-for-age children had 23 % (95 % CI −1 to 41 %) lower concentrations than adequately nourished children of the same age and 53 % (95 % CI 37 to 65 %) lower concentrations than adults. Day 7 lumefantrine concentrations were 44 % (95 % CI 38 to 49 %) lower following unsupervised treatment. The highest risk of recrudescence was observed in areas of emerging artemisinin resistance and very low transmission intensity. For all other populations studied, day 7 concentrations ≥200 ng/ml were associated with >98 % cure rates (if parasitemia <135,000/μL).ConclusionsCurrent artemether-lumefantrine dosing recommendations achieve day 7 lumefantrine concentrations ≥200 ng/ml and high cure rates in most uncomplicated malaria patients. Three groups are at increased risk of treatment failure: very young children (particularly those underweight-for-age); patients with high parasitemias; and patients in very low transmission intensity areas with emerging parasite resistance. In these groups, adherence and treatment response should be monitored closely. Higher, more frequent, or prolonged dosage regimens should now be evaluated in very young children, particularly if malnourished, and in patients with hyperparasitemia.Electronic supplementary materialThe online version of this article (doi:10.1186/s12916-015-0456-7) contains supplementary material, which is available to authorized users.

CYP2B6*6 genotype and high efavirenz plasma concentration but not nevirapine are associated with low lumefantrine plasma exposure and poor treatment response in HIV-malaria-coinfected patients.

We investigated the influence of efavirenz (EFV)- or nevirapine (NVP)-based antiretroviral therapy (ART) on lumefantrine plasma exposure in HIV-malaria-coinfected patients and implication of pharmacogenetic variations. A total of 269 HIV patients with uncomplicated falciparum malaria on NVP-based ART (NVP-arm), EFV-based ART (EFV-arm) or not receiving ART (control-arm) were enrolled and treated with artemether-lumefantrine. Day-7 lumefantrine, baseline EFV and NVP plasma concentrations, and CYP2B6*6,*18, CYP3A4*1B, CYP3A5*3,*6,*7, ABCB1 c.3435C>T and ABCB1 c.4036A>G genotypes were determined. The median day-7 lumefantrine plasma concentration was significantly lower in the EFV-arm compared with that in NVP- and control-arm. High EFV plasma concentrations and CYP2B6*6/*6 genotype significantly correlated with low lumefantrine plasma concentrations and high rate of recurrent parasitemia. No significant effect of NVP-based ART on lumefantrine exposure was observed. In conclusion, owing to long-term CYP3A induction, EFV-based ART cotreatment significantly reduces lumefantrine plasma exposure leading to poor malaria treatment response, which is more pronounced in CYP2B6 slow metabolizers.

Development and validation of a LC-MS/MS method for the quantitation of lumefantrine in mouse whole blood and plasma.

A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for the quantitation of the antimalarial drug, lumefantrine (LF), in mouse whole blood and plasma. The analyte was extracted using a protein precipitation method followed by chromatographic separation on a Phenomenex Luna, PFP (50mm×2.0mm, 5μm) analytical column with a mobile phase consisting of acetonitrile and 0.1% formic acid (formic acid:water, 1:1000, v/v) at a ratio of 3:7 (v/v), delivered at a constant flow rate of 0.5ml/min. Stable isotope labeled lumefantrine (D9-LF) was used as the internal standard. Multiple reaction monitoring was performed using the transitions m/z 530.1→m/z 347.9 and m/z 539.1→m/z 347.9 for the quantification of LF and D9-LF, respectively. Calibration curves were constructed over the concentration range 15.6-4000ng/ml. The mean intra- and inter-assay accuracy values for the analysis of LF in WB was 103% (%CV=5.5) and 99.5% (%CV=5.5), respectively. The mean intra- and inter-assay accuracy values for the analysis of LF in plasma was 93.7% (%CV=3.5) and 93.9% (%CV=5.5), respectively. No significant matrix effect was observed during the method validation. The validated method was applied to an absorption study in mice, to determine and compare LF concentrations in whole blood and plasma samples. Results of the statistical analysis using a linear mixed effects growth curve model concluded that there was no significant difference (p-value=0.668) between WB and plasma LF concentrations. This method utilizes a small sample volume of 20μl, facilitating low blood collection volumes and a short chromatographic run time of 3min which allows for high sample throughput analysis.

Bio-Analytical Method Development and Validation for Estimation of Lume fantrine in Human Plasma by Using Lc-Ms/Ms

Lumefantrine and Glimepiride (IS) were extracted from human plasma by Precipitation followed by Solid phase extraction using Orochem (30 mg/1 CC) solid phase extraction cartridge. The chromatographic separation was performed on Hypurity C18 (50 cm×4.6 mm), 5 μ column. The mobile phase consisted of Acetonitrile: 2 mM Ammonium Acetate (pH: 3.5) (90:10, % v/v) was delivered at rate of 0.600 mL/min with Splitter. Detection and quantitation were performed by a triple quadrupole equipped with electro spray ionization and multiple reaction monitoring in positive ionization mode (API 3000). The most intense [M-H]- transition for Lume fantrine at m/z 528.0→510.0 and for IS at m/z 491.2→352.0 were used for quantification. The developed method was successfully applied for bioequivalence study of Lume fantrine. The method was found to linear over the range of 100-20000 ng/mL (r≥0.992). The lower limit of quantitation (LLOQ) was 100 ng/mL. The extraction recovery was above 75% for analyte and above 90% for IS. The intra and inter-day accuracy was found to 92.27% - 104.00%. The intra and inter-day precision expressed as % CV were 1.76% - 6.47%, respectively. The stability testing was also investigated and it was found that both drug and IS were quite stable. A simple, rapid, sensitive, accurate and precise LC-MS/MS method has been developed for the quantification of Lumefantrine from human plasma using Protein precipitation followed by SPE method. The method exhibited good linear response over the selected concentration range 100-20000 ng/mL. Selectivity and sensitivity were sufficient for detecting and quantifying Lumefantrine in human plasma. These features coupled with a short run time at 3.50 min compared to reported methods, facilitated a high analysis throughput, with the ability to quantify a larger number of clinical samples in a shorter time frame.

Method development and validation for simultaneous determination of lumefantrine and its major metabolite, desbutyl lumefantrine in human plasma using RP-HPLC/UV detection

A simple, specific, precise and rapid RP-HPLC-UV method was developed for simultaneous determination of lumefantrine and its metabolite desbutyl lumefantrine in human plasma. Experimental parameters were optimized and the method was validated according to standard guidelines. The method showed adequate separation for lumefantrine and desbutyl lumefantrine and best resolution was achieved with Supelco Discovery HS C18 RP (150mm×4.6mm, 5μm) column using acetonitrile and 0.05% trifluroacetic acid (70:30, v/v) as a mobile phase pumped at a flow rate of 1.0ml/min and wavelength of 335nm. The method was linear over the concentration range of 10-12,000ng/ml. The lower limit of detection (LLOD) and lower limit of quantification (LLOQ) for lumefantrine were 10.0 and 18.0ng/ml, while for desbutyl lumefantrine were 7.5 and 15.0ng/ml, respectively. The proposed method was efficiently applied for determination of lumefantrine and desbutyl lumefantrine concentrations in plasma samples for pharmacokinetic studies.

Comparison of bioavailability between the most available generic tablet formulation containing artemether and lumefantrine on the Tanzanian market and the innovator’s product

BackgroundExistence of anti-malarial generic drugs with low bioavailability marketed on sub-Saharan Africa raises a concern on patients achieving therapeutic concentrations after intake of such products. This work compared bioavailability of one generic tablet formulation with innovator’s product. Both were fixed dose combination tablet formulations containing artemether and lumefantrine.MethodologyThe study was conducted in Dar Es Salaam, Tanzania, in which a survey of the most abundant generic containing artemether-lumefantrine tablet formulation was carried out in retail pharmacies. The most widely available generic (Artefan®, Ajanta Pharma Ltd, Maharashtra, India) was sampled for bioavailability comparison with Coartem® (Novartis Pharma, Basel, Switzerland) - the innovator’s product. A randomized, two-treatment cross-over study was conducted in 18 healthy Tanzanian black male volunteers. Each volunteer received Artefan® (test) and Coartem® (as reference) formulation separated by 42 days of drug-free washout period. Serial blood samples were collected up to 168 hours after oral administration of a single dose of each treatment. Quantitation of lumefantrine plasma levels was done using HPLC with UV detection. Bioequivalence of the two products was assessed in accordance with the US Food and Drug Authority (FDA) guidelines.ResultsThe most widely available generic in pharmacies was Artefan® from India. All eighteen enrolled volunteers completed the study and both test and reference tablet formulations were well tolerated. It was possible to quantify lumefantrine alone, therefore, the pharmacokinetic parameters reported herein are for lumefantrine. The geometric mean ratios for Cmax, AUC0-t and AUC0-∞ were 84% in all cases and within FDA recommended bioequivalence limits of 80% – 125%, but the 90% confidence intervals were outside FDA recommended limits (CI 49–143%, 53 - 137%, 52 - 135% respectively). There were no statistical significant differences between the two formulations with regard to PK parameters (P > 0.05).ConclusionsAlthough the ratios of AUCs and Cmax were within the acceptable FDA range, bioequivalence between Artefan® and Coartem® tablet formulations was not demonstrated due to failure to comply with the FDA 90% confidence interval criteria. Based on the observed total drug exposure (AUCs), Artefan® is likely to produce a similar therapeutic response as Coartem®.

Comparable Lumefantrine Oral Bioavailability when Co‐administered With Oil‐Fortified Maize Porridge or Milk in Healthy Volunteers

Co-administration of artemether-lumefantrine with milk is recommended to improve lumefantrine (L) absorption but milk may not be available in resource-limited settings. This study explored the effects of cheap local food in Uganda on oral bioavailability of lumefantrine relative to milk. In an open-label, four-period crossover study, 13 healthy adult volunteers were randomized to receive a single oral dose of artemether-lumefantrine (80 mg artemether/480 mg lumefantrine) with water, milk, maize porridge or maize porridge with oil on separate occasions. Plasma lumefantrine was assayed using high-performance liquid chromatography with ultraviolet detection. Pharmacokinetic exposure parameters were determined by non-compartmental methods using WinNonlin. Peak concentrations (Cmax ) and area under concentration-time curve restricted to 48 hr after single dosing (AUC(0-48) ) were selected for relative bioavailability evaluations using confidence interval approach for average bioequivalence. Lumefantrine exposure was comparable in milk and maize porridge plus oil study groups. When artemether-lumefantrine was administered with maize porridge plus oil, average bioequivalence ranges (means ratios 90% CI, 0.84-1.88 and 0.85-1.69 for Cmax and AUC(0-48) , respectively) were within and exceeded acceptance ranges relative to milk (90% CI, 0.80-1.25). Both fasted and maize porridge groups demonstrated similarly much lower ranges of lumefantrine exposures (bioinequivalence) relative to milk. If milk is not available, it is thus possible to recommend fortification of carbohydrate-rich food with little fat (maize porridge plus vegetable oil) to achieve similarly optimal absorption of lumefantrine after artemether-lumefantrine administration.

Exploration of in vivo efficacy of artemether-lumefantrine against uncomplicated Plasmodium falciparum malaria in under fives in Tabora region, Tanzania

BackgroundTanzania adopted artemether-lumefantrine (AL) as first-line drug for uncomplicated malaria in 2006. Recently, there was an anecdotal report on high malaria recurrence rate following AL treatment in in the (urban and peri-urban), western part of Tanzania. The current report is an exploratory study to carefully and systematically assess AL efficacy in the area.MethodsBetween June and August 2011, a total of 1,126 patients were screened for malaria, 33 had malaria, of which 20 patients met inclusion criteria and were enrolled and treated with standard dose of AL as recommended in the WHO protocol. Treated patients were followed up for 28 days to assess treatment responses. Before treatment (Day 0) and post-treatment (Day 7) plasma lumefantrine levels were determined to assess prior AL use and ascertain parasites exposure to adequate plasma leveles of lumefantrine, respectively.ResultsThe cure rate was 100%. All Day 0 plasma lumefantrine were below HPLC detectable level. The median Day 7 lumefantrine concentration was 404, (range, 189–894 ng/ml). Six out of 20 patients (30%) were gametocytaemic and all cleared gametocytes by Day 14. One patient showed an increase in gametocytes from four on Day 0 to 68, per 500 WBC on Day 2.ConclusionArtemether lumefantrine is highly efficacious against uncomplicated Plasmodium falciparum malaria. The elevation of gametocytaemia despite AL treatment needs to be evaluated in a larger study.

A fast and reliable reversed phase high performance liquid chromatography method for simultaneous determination of selected anti-retroviral and lumefantrine in human plasma

A fast and reliable high performance liquid chromatography (HPLC) method with UV diode array detection for simultaneous quantitative analysis of the anti-retroviral drugs, nevirapine (NVP) and efavirenz (EFV) and the anti-malarial, lumefantrine (LUM) in human plasma has been developed and validated. The sample preparation consisted of a plasma protein precipitation with 0.5% acetic acid acetonitrile solution containing the internal standard halofantrine (HALO) prior the LC-analysis. Chromatographic separation was carried out on a Acclaim Polar Advantage C(16), column (150 mm × 4.6 mm, particle size, 3 μm) using a gradient of mobile phase made of 0.01% TFA in 0.1M ammonium acetate (solvent A) and 0.1% TFA in acetonitrile (solvent B). The separation of NVP, EFV, LUM and HALO was achieved within 17 min at a flow rate of 1.0 mL/min and detections were initially performed at three wavelengths, 275 nm (NVP), 255 nm (EFV), and 300 nm (LUM). The method selectivity was demonstrated in six different human plasma batches. In addition, several concomitant drugs were analyzed under our experimental conditions and none of them co-eluted with EFV, NVP and LUM. This demonstrated that our method is highly selective. Calibration graphs plotted with seven concentrations in duplicate for each compound were linear between the selected ranges with a regression coefficient (R(2)) greater than 0.998. Absolute extraction recovery for NVP, EFV and LUM were 99%, 98.6 and 102%, respectively. Inter- and intra-day coefficients of variation for LUM, EFV and NVP were ≤10%. The lower limits of quantification were 0.125 μg/mL for LUM and 0.250 μg/mL for both EFV and NVP. Intra- and inter-assay relative standard deviation values were found to be less than 15% at the concentrations examined (0.125-10.0 μg/mL for LUM and 0.250-15.0 μg/mL for both EFV and NVP). The present method was successfully implemented in Tanzania and only one wavelength (255 nm) was used to measure samples of patients receiving either NVP or EFV in combination with LUM. The concentration found in human plasma samples for all three compounds were within the calibration range. This makes our method particularly applicable and useful to resource-limited settings.

Quantification of Lumefantrine in Human Plasma Using LC-MS/MS and Its Application to a Bioequivalence Study.

An analytical method based on protein precipitation has been developed and validated for analysis of lumefantrine in human plasma. Artesunate was used as an internal standard for lumefantrine. Inertsil ODS column provided chromatographic separation of analytes followed by detection with mass spectrometry. The method involves simple isocratic chromatographic condition and mass spectrometric detection in the positive ionization mode using an API-3000 system. The total run time was 2.5 minutes. The proposed method has been validated with linear range of 200–20000 ng/mL for lumefantrine. The intrarun and interrun precision values are within 6.66% and 5.56%, respectively, for lumefantrine at the lower limit of quantification level. The overall recovery for lumefantrine and artesunate was 93.16% and 91.05%, respectively. This validated method was used successfully for analysis of plasma samples from a bioequivalence study.

Interlaboratory Development and Cross Validation of a Chromatographic Method for Determination of Lumefantrine in Human Plasma-A Proficient Capacity Assessment of Bioanalytical Laboratories in East Africa

Background: Bioanalytical laboratories in developing countries face many challenges. The objective of this work was to assess the capacity of bioanalytical laboratories in emerging countries in setting and validating analytical methods. An HPLC method for determination of lumefantrine in human plasma was used to assess three medical university laboratories of Tanzania, Uganda and Kenya. Methodology: Bioanalytical experts from Analytical Clinical Concept, Leidersbach, in Germany (ACC GmbH) developed the HPLC method and assigned the 3 laboratories to set up and validate the method. The laboratories were tasked to determine the concentrations of blinded plasma samples spiked with lumefantrine and had to submit their analysis reports to ACC for evaluation within 6 weeks. Each laboratory was provided with reference standard, internal standard columns and precolumns. Spiked plasma samples were shipped under dry ice from Germany to “Gesellschaft fuer Internationale Zusammenarbeit” (GIZ) local office of each participating country. All other requirements were procured by individual laboratories. Results: The results from MUHAS Bioanalytical laboratory (Tanzania) Laboratory met the criteria set by ACC laboratory. The results were within the range set by ACC laboratory and most calibration curves had good linearity with coefficient of correlation always > 0.990. The inter-day precision and accuracy (Relative standard deviation=RD of recovery) were always < 15%. The relative deviation of the results obtained compared to assigned concentrations for blinded plasma samples were between -15% and -25%. The Laboratory met the stipulated time line and the obtained validation results were within the range set by the ACC experts. The results from other laboratories were also satisfactory. Conclusion: The results indicate that with further little infrastructural and technical assistance the capacity of these bioanalytical laboratories in conducting bio-analyses will be more strengthened and can serve as centers for training bioanalytics and running bioequivalence studies in the region.

Population Pharmacokinetics of Artemether, Lumefantrine, and Their Respective Metabolites in Papua New Guinean Children with Uncomplicated Malaria

There are sparse published data relating to the pharmacokinetic properties of artemether, lumefantrine, and their active metabolites in children, especially desbutyl-lumefantrine. We studied 13 Papua New Guinean children aged 5 to 10 years with uncomplicated malaria who received the six recommended doses of artemether (1.7 mg/kg of body weight) plus lumefantrine (10 mg/kg), given with fat over 3 days. Intensive blood sampling was carried out over 42 days. Plasma artemether, dihydroartemisinin, lumefantrine, and desbutyl-lumefantrine were assayed using liquid chromatography-mass spectrometry or high-performance liquid chromatography. Multicompartmental pharmacokinetic models for a drug plus its metabolite were developed using a population approach that included plasma artemether and dihydroartemisinin concentrations below the limit of quantitation. Although artemether bioavailability was variable and its clearance increased by 67.8% with each dose, the median areas under the plasma concentration-time curve from 0 h to infinity (AUC(0-∞)s) for artemether and dihydroartemisinin (3,063 and 2,839 μg · h/liter, respectively) were similar to those reported previously in adults with malaria. For lumefantrine, the median AUC(0-∞) (459,980 μg · h/liter) was also similar to that in adults with malaria. These data support the higher dose recommended for children weighing 15 to 35 kg (35% higher than that for a 50-kg adult) but question the recommendation for a lower dose in children weighing 12.5 to 15 kg. The median desbutyl-lumefantrine/lumefantrine ratio in the children in our study was 1.13%, within the range reported for adults and higher at later time points because of the longer desbutyl-lumefantrine terminal elimination half-life. A combined desbutyl-lumefantrine and lumefantrine AUC(0-∞) weighted on in vitro antimalarial activity was inversely associated with recurrent parasitemia, suggesting that both the parent drug and the metabolite contribute to the treatment outcome of artemether-lumefantrine.

Desbutyl-Lumefantrine Is a Metabolite of Lumefantrine with Potent In Vitro Antimalarial Activity That May Influence Artemether-Lumefantrine Treatment Outcome

Desbutyl-lumefantrine (DBL) is a metabolite of lumefantrine. Preliminary data from Plasmodium falciparum field isolates show greater antimalarial potency than, and synergy with, the parent compound and synergy with artemisinin. In the present study, the in vitro activity and interactions of DBL were assessed from tritium-labeled hypoxanthine uptake in cultures of the laboratory-adapted strains 3D7 (chloroquine sensitive) and W2mef (chloroquine resistant). The geometric mean 50% inhibitory concentrations (IC(50)s) for DBL against 3D7 and W2mef were 9.0 nM (95% confidence interval, 5.7 to 14.4 nM) and 9.5 nM (95% confidence interval, 7.5 to 11.9 nM), respectively, and those for lumefantrine were 65.2 nM (95% confidence interval, 42.3 to 100.8 nM) and 55.5 nM (95% confidence interval, 40.6 to 75.7 nM), respectively. An isobolographic analysis of DBL and lumefantrine combinations showed no interaction in either laboratory-adapted strain but mild synergy between DBL and dihydroartemisinin (sums of the fractional inhibitory concentrations of 0.92 [95% confidence interval, 0.87 to 0.98] and 0.94 [95% confidence interval, 0.90 to 0.99] for 3D7 and W2mef, respectively). Using a validated ultra-high-performance liquid chromatography-tandem mass spectrometry assay and 94 day 7 samples from a previously reported intervention trial, the mean plasma DBL was 31.9 nM (range, 1.3 to 123.1 nM). Mean plasma DBL concentrations were lower in children who failed artemether-lumefantrine treatment than in those with an adequate clinical and parasitological response (ACPR) (P = 0.053 versus P > 0.22 for plasma lumefantrine and the plasma lumefantrine-to-DBL ratio, respectively). DBL is more potent than the parent compound and mildly synergistic with dihydroartemisinin. These properties and the relationship between day 7 plasma concentrations and the ACPR suggest that it could be a useful alternative to lumefantrine as a part of artemisinin combination therapy.

Measurement of lumefantrine and its metabolite in plasma by high performance liquid chromatography with ultraviolet detection

Artemether-lumefantrine (ARM-LUM) has in recent years become the first-line treatment for uncomplicated malaria in many Sub-Saharan African countries. Vigorous monitoring of the therapeutic efficacy of this treatment is needed. This requires high-quality studies following standard protocols; ideally, such studies should incorporate measurement of drug levels in the study patients to exclude the possibility that insufficient drug levels explain an observed treatment failure. Several methods for measuring lumefantrine (LUM) in plasma by HPLC are available; however, several of these methods have some limitations in terms of high costs and limited feasibility arising from large required sample volumes and demanding sample preparation. Therefore, we set out to develop a simpler reversed phase high performance liquid chromatography (RP-HPLC) method based on UV detection for simultaneous measurement of LUM and its major metabolite the desbutyl LUM (DL) in plasma. Halofantrine was used as an internal standard. Liquid–liquid extraction of samples was carried out using hexane–ethyl acetate (70:30, v/v). Chromatographic separation was carried out on a Synergi Polar-RP column (250 mm × 300 mm, particle size 4 μm). The mobile phase consisted of acetonitrile–0.1 M ammonium acetate buffer adjusted to pH 4.9 (85:15%, v/v). Absorbance of the compounds was monitored at 335 nm using a reference wavelength of 360 nm. Absolute extraction recovery for LUM and DL were 88% and 90%, respectively. Inter- and intraday coefficients of variation for LUM and DL were ≤10%. The lower limits of quantification for LUM and DL were 12.5 and 6.5 ng/ml, respectively. After validation, the methodology was transferred to a local laboratory in Tanga Tanzania and samples from a small subset of malaria patients were analysed for LUM. The method appears to be applicable in settings with limited facilities.

Liquid chromatography–tandem mass spectrometry for the simultaneous quantitation of artemether and lumefantrine in human plasma: Application for a pharmacokinetic study

A liquid chromatography–electrospray ionization tandem mass spectrometry (HPLC–ESI-MS/MS) method for the simultaneous quantitation of artemether and lumefantrine in human plasma was developed and validated. Artesunate was used as an internal standard (IS). The analytes were extracted by a protein precipitation procedure and separated on a reversed-phase Zorbax SB-Ciano column with a mobile phase composed of methanol and 10 mM aqueous ammonium acetate containing 0.2% (v/v) acetic acid and 0.1% (v/v) formic acid. Multiple reaction monitoring was performed using the transitions m/ z 316 → m/ z 267, m/ z 530 → m/ z 348 and m/ z 402 → m/ z 267 to quantify artemether, lumefantrine and artesunate, respectively. Calibration curves were constructed over the range of 10–1000 ng/mL for artemether and 10–18,000 ng/mL for lumefantrine. The lower limit of quantitation was 10 ng/mL for both drugs. The mean R.S.D. values for the intra-run precision were 2.6% and 3.0% and for the inter-run precision were 3.6% and 4.6% for artemether and lumefantrine, respectively. The mean accuracy values were 102.0% and 101.2% for artemether and lumefantrine, respectively. No matrix effect was detected in the samples. The validated method was successfully applied to determine the plasma concentrations of artemether and lumefantrine in healthy volunteers, in a one-dose pharmacokinetic study, over the course of 11 days.

LC-Tandem Mass Spectrometry Method for Quantification of Lumefantrine in Human Plasma and Its Application to Bioequivalence Study

A rapid, specific and robust assay based on protein precipitation and liquid chromatography-electrospray ionization tandem mass spectrometry (LC–ESI–MS–MS) has been developed and validated for the quantitative analysis of lumefantrine, an antimalarial drug, in human plasma using piperazine bis chloroquinoline as internal standard (ISTD). The precursor to product ion transitions of m/z 528.2 → 510.3 and m/z 409.2 → 205.1 were used to measure the analyte and the ISTD, respectively. Analysis was performed on C8 column (50 mm × 4.6 mm, 5 μm) with 10 mM ammonium acetate/acetonitrile/0.05% acetic acid (10/85/5, v/v) as mobile phase at a flow rate of 0.6 mL min−1. The assay exhibited a linear dynamic range of 0.21–25.05 μg mL−1. The RSD% of intra-day and inter-day assay was ≤15%. The application of this assay was demonstrated in a bioequivalence study and will be ideal for pharmacokinetic studies in pediatric population.

The effect of food consumption on lumefantrine bioavailability in African children receiving artemether-lumefantrine crushed or dispersible tablets (Coartem®) for acute uncomplicatedPlasmodium falciparummalaria

Artemether-lumefantrine (AL) is first-line treatment for uncomplicated malaria in many African countries. Concomitant food consumption may affect absorption of lumefantrine but data in the most important target population, i.e. children, are lacking. Therefore, we evaluated the effect of food intake on oral lumefantrine bioavailability in African children with malaria. In a randomised, investigator-blinded, multicentre phase III efficacy trial, 899 infants and children with acute uncomplicated Plasmodium falciparum malaria received six doses of AL according to body weight over 3 days either as crushed tablets (Coartem) or as dispersible tablets. Single blood samples were obtained for lumefantrine plasma concentration determination in a subset of 621 patients, and a two-compartment pharmacokinetic model was constructed. The mean observed lumefantrine plasma concentration for crushed tablet and dispersible tablet, respectively, was 100% and 55% higher with a concomitant meal at the time of dose intake than when taken alone. Similarly, consumption of milk (the most common meal) increased model-estimated lumefantrine bioavailability by 57% (90% CI: 29-96%) with crushed tablets and 65% (90% CI: 28-109%) with dispersible tablets compared to no food. The 28-day PCR-corrected cure rate (primary study endpoint) in the evaluable population was 582/587 [99.1% (95% CI: 98.0-99.7%)] and was not related to food intake. AL was highly efficacious. Concomitant food intake increased lumefantrine absorption in children with malaria.

A modified method for determination of lumefantrine in human plasma by HPLC-UV and combination of protein precipitation and solid-phase extraction: application to a pharmacokinetic study.

An HPLC-UV method was developed and validated for the determination of lumefantrine in human plasma. Lumefantrine and its internal standard halofantrine were extracted from plasma samples using protein precipitation with acetonitrile (0.2% perchloric acid) followed by solid-phase extraction with Hypersep C(8) cartridges. Chromatographic separation was performed on a Zorbax SB-CN HPLC column (3.0 x 150 mm, 3.5 microm) with water/methanol (0.1% TFA) as the mobile phases in a gradient elution mode. Detection was performed using UV/vis detector at lambda = 335 nm. The method showed to be linear over a range of 50-10,000 ng/mL with acceptable intra- and inter-day precision and accuracy. The mean recoveries were 88.2% for lumefatrine and 84.5% for the I.S. The internal standard halofantrine is readily available from commercial sources. This method was successfully applied to a pharmacokinetic interaction study between a first-line antimalarial combination (artemether-lumefantrine) and antiretroviral therapy.