| 1. |
- Blessborn, Daniel, et al.
(författare)
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Determination of Lumefantrine after Capillary Sampling onto Sampling Paper
- 2007
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Ingår i: The Royal Society of Tropical Medicine and Hygiene. - London.
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Konferensbidrag (refereegranskat)abstract
- The antimalarial lumefantrine was first synthesised and registered in China and is now commercially available as a coformulated product together with artemether (Coartem®/Riamet®). This combination is well tolerated and has proven highly efficacious for treatment of uncomplicated falciparum malaria. Lumefantrine is highly lipophilic with an extensive protein binding (99.9%). The day 7 plasma lumefantrine level has been shown to be an important determinant of treatment efficacy. To date no method has been published for the determination of lumefantrine after capillary sampling onto filter paper for field use. The aim of this work was to develop a method with adequate sensitivity for quantification of lumefantrine in capillary blood sampled onto filter paper. The method has been validated according to the current FDA guideline for bioanalytical method validation. Method: Whatman 31 ET Chr filter paper was pre-treated with an organic acid before sampling capillary blood to enable a high recovery of lumefantrine. Lumefantrine was extracted from the filter paper, then further purified using solid phase extraction and finally quantified with HPLC. Results: The between day variation is below 10 % over the range 0.4 to 25 µmol/l. The lower limit of quantification is 0.25 µmol/l in 100 µl capillary blood. No decrease in Lumefantrine concentration in dried blood spot is seen after 3 months at 37o C. The field sampling for lumefantrine assay with pre-treated Whatman 31 ET Chr has been tested in Tanzania with good results. Discussion: The field sampling for lumefantrine concentration assay with pre-treated Whatman 31 ET Chr has been evaluated and proven to be a valid method for field studies. The day 7 level after treatment can lumefantrine be accurately estimated in capillary blood to follow up compliance and efficacy. Validation data will be presented.
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| 2. |
- Blessborn, Daniel, et al.
(författare)
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Development and validation of an automated solid-phase extraction and liquid chromatographic method for determination of lumefantrine in capillary blood on sampling paper
- 2007
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Ingår i: Journal of Pharmaceutical and Biomedical Analysis. - : Elsevier BV. - 0731-7085 .- 1873-264X. ; 45:2, s. 282-287
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Tidskriftsartikel (refereegranskat)abstract
- A bioanalytical method for the determination of lumefantrine in 100 μl 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 μM. The lower limit of quantification was 0.25 μM in 100 μl capillary blood. No decrease in lumefantrine concentration in dried blood spot is seen after 4 months storage at 22 °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.
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| 3. |
- Blessborn, Daniel, et al.
(författare)
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A new approach to evaluate stability of amodiaquine and its metabolite in blood and plasma
- 2006
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Ingår i: Journal of Pharmaceutical and Biomedical Analysis. - : Elsevier BV. - 0731-7085 .- 1873-264X. ; 41:1, s. 207-212
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Tidskriftsartikel (refereegranskat)abstract
- A stability study for amodiaquine (AQ) and desethylamodiaquine (AQm) in whole blood and plasma is reported. AQ, AQm and chloroquine (CQ) were simultaneously analysed and the ratios AQ/CQ and AQm/CQ were used to ensure correct interpretation of the stability results. CQ was stable in whole blood and plasma at all tested temperatures enabling it to be a stability marker in stability studies. Simultaneous analysis of compounds, of which at least one is already known to be stable, permits a within sample ratio to be used as a stability indicator, The new approach significantly reduced bias when compared to the traditional approach. AQ and AQm were stable in plasma at -86 degrees C and -20 degrees C for 35 days, at 4 degrees C for 14 days and at 22 degrees C for 1 day. AQ and AQm were stable in blood at -86 degrees C and 4 degrees C for 35 days, at -20 degrees C and 22 degrees C for 7 days and at 37 degrees C for 1 day.
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| 4. |
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| 5. |
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| 6. |
- Blessborn, Daniel
(författare)
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Development of Analytical Methods for Measurement of Drugs against Malaria in Plasma and Whole Blood
- 2006
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Licentiatavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The aim of this thesis was to develop analytical methods for measuring drugs in blood and/or plasma. Solid phase extraction was used for the enrichment and purification of the drugs from the sample matrix. Automatic extraction procedures using a solid phase extraction robot to reduce the workload of the analyst and to minimize variations in the extraction procedure were developed. To determine sample concentrations, liquid chromatography was used together with UV absorbance detection through out this work. Determination of Pyronaridine in whole blood utilised a weak cation exchanger to extract Pyronaridine from blood. From the beginning there were problems in separating Pyronaridine from the internal standard but by adding sodium perclorate as an ion-pairing agent, good separation was achieved. For the simultaneous quantification of the highly lipophilic Atovaquone and the strong basic drug Proguanil with metabolites, a novel mixed mode solid phase extraction column was used. It combines the properties of a carboxylic acid (CBA) column and a non-polar octyl-silica (C8) column to extract the compounds from plasma. Their different physiochemical properties also required a gradient separation on the liquid chromatography system. Stability is an important factor when developing new methods. A new approach was used to evaluate the stability of Amodiaquine in blood and plasma. This included the use of a stability marker, in this case Chloroquine, a stable compound which was added together with Amodiaquine when preparing the stability samples. When using the ratio of Amodiaquine and the stability marker, between-run variations and variations associated with the preparation of new stock solutions and calibration standards were eliminated.
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| 7. |
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| 8. |
- Blessborn, Daniel
(författare)
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Development of Analytical Methods for the Determination of Antimalarials in Biological Fluids
- 2009
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The aim of this thesis was to develop analytical methods for measuring antimalarial drugs in biological fluids. Solid phase extraction (SPE) was used for the enrichment and purification of the drugs. Automatic extraction procedures using a SPE robot were developed to reduce the workload for the analyst and to minimize variations in the extraction procedure. Liquid chromatography (LC) with either UV or mass spectrometric (MS) detection was used to determine sample concentrations. Determination of Pyronaridine in whole blood utilised a weak cation exchanger to extract Pyronaridine from blood. To improve LC separation between Pyronaridine and the internal standard, ion-pairing was utilized. For the simultaneous quantification of the highly lipophilic Atovaquone and the strong basic drug Proguanil with metabolites, a novel mixed mode solid phase extraction column was used. It combines the properties of a carboxylic acid (CBA) column and a non-polar octyl-silica (C8) column to extract the compounds from plasma; it also required a gradient LC separation. Stability is an important factor when developing new methods. A new approach was used to evaluate the stability of Amodiaquine in blood and plasma. This included the use of a stability marker, a stable compound which was added together with Amodiaquine when preparing the stability samples. This eliminated between-run variations and variations associated with preparation of new stock solutions. Lumefantrine (LF) is one of the active components in a new drug combination recommended by the World Health Organization as a replacement for older drugs which have lost their effect. The first of the two methods described for this compound is the determination of LF and a possible metabolite in plasma with a calibration range suitable for pharmacokinetic studies. In the second method, a capillary sampling technique is used where the blood is dried on a sampling paper and sent to the laboratory where the extraction and determination of LF concentrations take place. This method facilitates sample collection and will enable drug efficacy studies conducted in rural settings. To monitor a current change in treatment policy and self medication, a screening assay was developed. Its purpose is to be a complement to interviewing patients about their previous medication (in the previous few weeks) and to detect some of the more common drugs which might have been used.
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| 9. |
- Lindegårdh, Niklas, et al.
(författare)
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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
- 2005
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Ingår i: Journal of Pharmaceutical and Biomedical Analysis. - : Elsevier BV. - 0731-7085 .- 1873-264X. ; 37:5, s. 1081-1088
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Tidskriftsartikel (refereegranskat)abstract
- 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 μ g/mL, respectively, and for DLF 4.5 and 1.5% at 0.039 and 0.777 μ g/mL, respectively. The between-day precisions for LF were 12.0 and 2.9% at 0.042 and 8.02 μ g/mL, respectively, while for DLF 0.7 and 1.2% at 0.039 and 0.777 μ g/mL, respectively. The limit of quantification was 0.024 and 0.021 μ g/mL for LF and DLF, respectively. Different amounts of lipids in plasma did not affect the absolute recovery of LF or DLF.
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| 10. |
- Lindegårdh, Niklas, et al.
(författare)
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Simultaneous quantitation of the highly lipophilic atovaquone and hydrophilic strong basic proguanil and its metabolites using a new mixed-mode SPE approach and steep-gradient LC
- 2005
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Ingår i: Journal of Chromatographic Science. - : Oxford University Press (OUP). - 0021-9665 .- 1945-239X. ; 43:5, s. 259-266
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Tidskriftsartikel (refereegranskat)abstract
- A bioanalytical method is described for the simultaneous quantitative analysis of the highly lipophilic atovaquone and the strong basic proguanil with metabolites in plasma. The drugs are extracted from protein precipitated plasma samples on a novel mixed-mode solid-phase extraction (SPE) column containing carboxypropyl and octyl silica as functional groups. The analytes are further separated and quantitated using a steep-gradient liquid chromatographic method on a Zorbax SB-CN column with UV detection at 245 nm. Two different internal standards (IS) are used in the method to compensate for both types of analytes. A structurally similar IS to atovaquone is added with acetonitrile to precipitate proteins from plasma. A structurally similar IS to proguanil and its metabolites is added with phosphate buffer before samples are loaded onto the SPE columns. A single elution step is sufficient to elute all analytes. The method is validated according to published guidelines and shows excellent performance. The within-day precisions, expressed as relative standard deviation, are lower than 5% for all analytes at three tested concentrations within the calibration range. The between-day precisions are lower than 13% for all analytes at the same tested concentrations. The limit of quantitation is 25 nM for the basic substances and 50 nM for atovaquone. Several considerations regarding development and optimization of a method for determination of analytes with such a difference in physiochemical properties are discussed.
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| 11. |
- Römsing, Susanne, et al.
(författare)
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A METHOD FOR SCREENING SOME OF THE MOST COMMON ANTIMALARIAL DRUGS OF TODAY
- 2008
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Ingår i: Pharmaceutical & Biomedical Analysis. - Gdansk, Poland.
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Konferensbidrag (refereegranskat)abstract
- Malaria is an infectious decease caused by a one-celled parasite called Plasmodium. The most common types are falciparum and vivax malaria where falciparum is the deadliest form of malaria infection. Malaria is present in almost all tropical and sub-tropical regions around the world and there are at least 300 million cases of malaria in the world with over a million deaths every year. Some of the most frequently used antimalarial drugs are chloroquine, amodiaquine and sulphadoxine-pyrimethamine but drug resistance of the plasmodium falciparum parasite is spreading. The situation is especially serious in Southeast Asia where multi-drug resistant falciparum malaria is widespread. One factor that adds to drug resistance is poor compliance with dosing schedules. There are also increasing problems with counterfeit drugs that contain none or to low amount of antimalarial drug and this will also speed up drug resistance. It has recently become more common to combine different antimalarial drugs, preferably using drugs with different mechanisms of action, in order to increase efficacy and reduce the spread of malarial drug resistance within the malaria parasite populations. The aim of the developed screening method is to determine drug use in areas where a change in treatment policy has taken place since this would be more accurate than interviewing patients. Several of the drugs included in this screening method have relative long half-life e.g.chloroquine, pyrimethamine, mefloquine, and lumefantrine that are detectable at least 7 days after administration. Solid phase extraction is used as sample clean-up of plasma samples and separation is performed with gradient-LC and UV-detection at two different wavelengths. The developed method will be presented together with detection limits of the various drugs.
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