| 1. |
- Salas, Emir, et al.
(författare)
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A heterogeneous enzymatic assay for quantification of Plasmepsin II activity and the evaluation of its inhibitors
- 2004
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Ingår i: Journal of Pharmaceutical and Biomedical Analysis. - 0731-7085 .- 1873-264X. ; 34:4, s. 833-840
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Tidskriftsartikel (refereegranskat)abstract
- The emergence and worldwide spreading of Plasmodium falciparum strains that shown to be resistant to traditional drugs is considered a very serious health problem, given the high mortality and morbidity rate of Malaria. In the search for new drugs against this parasite, Hb hydrolyzing enzymes, such as Plasmepsin II (Plm II), have been classified as very promising targets for therapeutic attacks. In this work, it is developed a cheap and high-throughput heterogeneous enzymatic assay for measuring Plasmepsin II activity in order to use it as a tool in the discovery of new inhibitors of this enzyme. In this assay, Plasmepsin II acts upon a solid-phase bound synthetic peptide (DU2) whose sequence comprises the cleavage site F(33)-L(34) present in Hb alpha-chain. The peptide surface density is quantified by means of a classical ELISA-based procedure. In order to estimate the kinetic constants of the system and to quantify both, enzymatic and inhibitory activity, it was used a model for the kinetics of enzyme quasi-saturable systems previously developed by our group, that fitted very well to the experimental data. It was used Pepstatin as a model inhibitor of Plasmepsin II and the resulting dose-response relation agreed with the expected behavior for the Pepstatin-Plasmepsin II pair under the employed experimental conditions.
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| 2. |
- Wiberg, Kent, et al.
(författare)
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Rapid determination of lidocaine solutions with non-column chromatographic diode array UV spectroscopy and multivariate calibration
- 2003
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Ingår i: Journal of Pharmaceutical and Biomedical Analysis. - 0731-7085 .- 1873-264X. ; 30:5, s. 1575-1586
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Tidskriftsartikel (refereegranskat)abstract
- A new method for the rapid determination of pharmaceutical solutions is proposed. A conventional HPLC system with a Diode Array Detector (DAD) was used with no chromatographic column connected. As eluent, purified water (Milli Q) was used. The pump and autosampler of the HPLC system were mainly utilised as an automatic and convenient way of introducing the sample into the DAD. The method was tested on the local anaesthetic compound lidocaine. The UV spectrum (245–290 nm) from the samples analysed in the detector was used for multivariate calibration for the determination of lidocaine solutions. The content was determined with PLS regression. The effect on the predictive ability of three factors: flow, data-collection rate and rise time as well as two ways of exporting a representative UV spectrum from the DAD file collected was investigated by means of an experimental design comprising 11 experiments. For each experiment, 14 solutions containing a known content of lidocaine were analysed (0.02–0.2 mg ml−1). From these 14 samples two calibration sets and two test sets were made and as the response in the experimental design the Root Mean Square Error of Prediction (RMSEP) values from the predictions of the two test sets were used. When the factor setting giving the lowest RMSEP was found, this setting was used when analysing a new calibration set of 12 lidocaine samples (0.1–0.2 mg ml−1). This calibration model was validated by two external test sets, A and B, analysed on separate occasions for the evaluation of repeatability (test set A) and determination over time (test set B). For comparison, the reference method, liquid chromatography, was also used for analysis of the ten samples in test set B. This comparison of the two methods was done twice on different occasions. The results show that in respect of accuracy, precision and repeatability the new method is comparable to the reference method. The main advantages compared with liquid chromatography are the much shorter time of analysis (<30 s) as well as the automatic and simple analytical procedure and the low consumption of organic solvents.
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| 3. |
- Wiberg, Kent, et al.
(författare)
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Use of control sample for estimation of prediction error in multivariate determination of lidocaine solutions with non-column chromatographic diode array UV spectroscopy
- 2003
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Ingår i: Journal of Pharmaceutical and Biomedical Analysis. - 0731-7085 .- 1873-264X. ; 33:5, s. 859-869
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Tidskriftsartikel (refereegranskat)abstract
- The aim of this study was to investigate the ability of a control sample, of known content and identity, to diagnose and correct errors in the predictions when the same multivariate calibration model was used for analysis of new samples over time. A calibration set consisting of 16 samples with a known content of lidocaine was analysed and two external test sets, A and B, were used for the validation. Test set A contained 15 samples with different concentrations of lidocaine and test set B contained three samples with different lidocaine content, which were analysed six times in order to obtain a measure of repeatability. The multivariate calibration was done with PLS regression on UV spectra collected between 245 and 290 nm. A representative UV spectrum was exported from the collected DAD files by two methods, average spectrum over the whole file and average spectrum over the sample plug. Test set A was analysed further on another three occasions together with a control sample. The results showed that the control sample could be used to give a diagnosis and estimate of the prediction error. Moreover, the measured prediction error of the control sample could also be used to correct the predictions, thereby reducing the prediction error. Finally, some practical considerations regarding use of the proposed DAD method with a control sample are presented. The procedure suggested could lead to an efficient analytical approach where the same calibration model could be used over time without recalibration, which may be attractive in industrial quality control or screening analysis in pharmaceutical research.
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| 4. |
- Abbasy, Leila, et al.
(författare)
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Development of a reliable bioanalytical method based on prostate specific antigen trapping on the cavity of molecular imprinted polymer towards sensing of PSA using binding affinity of PSA-MIP receptor : A novel biosensor
- 2020
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Ingår i: Journal of Pharmaceutical and Biomedical Analysis. - : Elsevier. - 0731-7085 .- 1873-264X. ; 188
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Tidskriftsartikel (refereegranskat)abstract
- In this study, electrically-conducting poly [Toluidine Blue (PTB)] was applied as artificial receptor. It was organized by molecular imprinting approaches and via electrochemical technique for the sensitive monitoring of prostate-specific antigen (PSA). The protein-imprinted PTB was electropolymerized in a pre-formed glutaraldehyde-cysteamine (GA-Cys A) matrix on the surface of gold electrode, which significantly boosted the stability against degradation of the Molecular Imprinted Polymer (MIP) on the surface of pre-modified gold electrode. Moreover, the MIP bio-receptor ability towards protein recognition was explored by some electrochemical techniques. The binding affinity of MIP system was considerably upper than that of non-imprinted polymer (NIP) system, indicating the success of the method in generating imprinted materials that was specifically use to PSA protein. The incubation of the MIP modified electrode in various concentration of PSA (from 1-60 μg/L) resulted in the increase of the Fe (CN)63-/4- redox peak current. The bio-device also showed linear response from 1-60 μg/L and LLOQ of 1 μg/L by using DPV technique, leading to PSA monitoring in clinical samples. The proposed MIP-based biosensor was satisfactorily applied to the determination of PSA in human plasma samples. Therefore, the developed bio-device provides a new approach for sensitive, simple, rapid, and cost-effective monitoring of 1 μg/L of PSA. Notably, this approach could appear as an appropriate candidate for point-of-care (POC) use in clinical and biomedical analyses.
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| 5. |
- Abdel-Khalik, Jonas, et al.
(författare)
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Incorporation of (14)C-cholesterol in human adrenal corticocarcinoma H295R cell line and online-radiodetection of produced (14)C-steroid hormone metabolites
- 2017
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Ingår i: Journal of Pharmaceutical and Biomedical Analysis. - 0731-7085 .- 1873-264X. ; 145, s. 569-575
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Tidskriftsartikel (refereegranskat)abstract
- This study demonstrates the addition of (14)C-cholesterol to the human cell line H295R will in-situ form radiolabeled steroid hormones allowing for new mechanistic and metabolic insights. The aim of the present study was to in-situ radiolabel steroid hormones from cell line-incorporated (14)C-cholesterol using the OECD guideline 456, H295R steroidogenesis in-vitro assay. Radiodetection of the steroid metabolites of the steroidogenic pathway allows for an improved understanding of the various enzymatic mechanisms involved without necessarily being dependent on quantification. Generated radiolabeled steroids were analyzed using HPLC hyphenated with a Flow Scintillation Analyzer (FSA). H295R cells were incubated with radiolabeled cholesterol and cell media were collected and prepared by solid phase extraction and analyzed with HPLC-FSA. For successful radiolabeling of the steroids in the steroidogenesis of H295R cells, radioactive cholesterol may potentially only need to be added just before the cells are incubated for 72h in well plates. Based on the obtained HPLC-FSA chromatograms, and confirmation of the observations by studies in the literature, a qualitative time profile for the production of steroid hormones was estimated. Multiple radiolabeled steroid hormones were identified by means of analytical standards and UV (ultraviolet) co-chromatography, though the elucidation of multiple metabolites remains unresolved. Although online radiodetection proved to suffer from suboptimal sensitivity, the concept of radiolabeling the steroidogenesis in H295R cells with (14)C-cholesterol and detecting the radiolabeled steroid hormones online was proved and may assist in further toxicological studies.
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| 6. |
- Abdel-Khalik, Jonas, et al.
(författare)
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Incorporation of (14)C-cholesterol in human adrenal corticocarcinoma H295R cell line and online-radiodetection of produced (14)C-steroid hormone metabolites
- 2017
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Ingår i: Journal of Pharmaceutical and Biomedical Analysis. - : Elsevier. - 0731-7085 .- 1873-264X. ; 145, s. 569-575
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Tidskriftsartikel (refereegranskat)abstract
- This study demonstrates the addition of (14)C-cholesterol to the human cell line H295R will in-situ form radiolabeled steroid hormones allowing for new mechanistic and metabolic insights. The aim of the present study was to in-situ radiolabel steroid hormones from cell line-incorporated (14)C-cholesterol using the OECD guideline 456, H295R steroidogenesis in-vitro assay. Radiodetection of the steroid metabolites of the steroidogenic pathway allows for an improved understanding of the various enzymatic mechanisms involved without necessarily being dependent on quantification. Generated radiolabeled steroids were analyzed using HPLC hyphenated with a Flow Scintillation Analyzer (FSA). H295R cells were incubated with radiolabeled cholesterol and cell media were collected and prepared by solid phase extraction and analyzed with HPLC-FSA. For successful radiolabeling of the steroids in the steroidogenesis of H295R cells, radioactive cholesterol may potentially only need to be added just before the cells are incubated for 72h in well plates. Based on the obtained HPLC-FSA chromatograms, and confirmation of the observations by studies in the literature, a qualitative time profile for the production of steroid hormones was estimated. Multiple radiolabeled steroid hormones were identified by means of analytical standards and UV (ultraviolet) co-chromatography, though the elucidation of multiple metabolites remains unresolved. Although online radiodetection proved to suffer from suboptimal sensitivity, the concept of radiolabeling the steroidogenesis in H295R cells with (14)C-cholesterol and detecting the radiolabeled steroid hormones online was proved and may assist in further toxicological studies.
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| 7. |
- Abrahamsson, Pernilla, 1972-, et al.
(författare)
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An assessment of calibration and performance of the microdialysis system
- 2005
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Ingår i: Journal of Pharmaceutical and Biomedical Analysis. - : Elsevier. - 0731-7085 .- 1873-264X. ; 39:3-4, s. 730-734
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Tidskriftsartikel (refereegranskat)abstract
- To improve the reliability of microdialysis measurements of tissue concentrations of metabolic substances, this study was designed to test both the performance and the internal validity of the microdialysis methods in the hands of our research group. The stability of the CMA 600 analyser was tested with a known glucose solution in 72 standard microvials and in 48 plastic vials. To evaluate if variation in sampling time makes any difference in sample concentration (recovery), sampling times of 10, 20 and 30 min were compared in vitro with a constant flow rate of 1 microl/min. For testing of sampling times at different flow rates, an in vitro study was performed in which a constant sample volume of 10 microl was obtained. With the no net flux method, the actual concentration of glucose and urea in subcutaneous tissue was measured. The CMA 600 glucose analysis function was accurate and stable with a coefficient of variability (CV) of 0.2-0.55%. There was no difference in recovery for the CMA 60 catheter for glucose when sampling times were varied. Higher flow rates resulted in decreased recovery. Subcutaneous tissue concentrations of glucose and urea were 4.4 mmol/l and 4.1 mmol/l, respectively. To conclude, this work describes an internal validation of our use of the microdialysis system by calibration of vials and catheters. Internal validation is necessary in order to be certain of adequate sampling times, flow rates and sampling volumes. With this in mind, the microdialysis technique is useful and appropriate for in vivo studies on tissue metabolism.
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| 8. |
- Abrahamsson, Pernilla, 1972-, et al.
(författare)
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Optimised sample handling in association with use of the CMA 600 analyser
- 2008
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Ingår i: Journal of Pharmaceutical and Biomedical Analysis. - : Elsevier. - 0731-7085 .- 1873-264X. ; 48:5, s. 940-945
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Tidskriftsartikel (refereegranskat)abstract
- A large degree of variability for batched analysis of serially collected microdialysis samples measured with the CMA 600 analyser has been described. This study was designed to identify sources of variability related to sample handling. Standard concentrations of four solutes were placed in microdialysis vials and then stored and analysed at intervals. Results were analysed for variability related to vial and cap type, duration and temperature of storage, centrifugation and re-analysis. The main results were that centrifugation of samples reduced variability. When a batch of 24 samples was analysed, the use of crimp caps reduced evaporation. Samples in glass vials with crimp caps could be stored in a refrigerator for up to 14 days without large variability in concentration compared to plastic vials which demonstrated variability already when stored for more than 1 day. We conclude that variability in microdialysis results can occur in relation to storage and analysis routines if routines are not optimised concerning evaporation. Centrifugation before analyses, glass vials with crimp caps even during frozen storage, and attention to minimal times for samples to be uncapped during analysis all contribute to minimise variability in the handling and analysis of microdialysis samples.
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| 9. |
- Adler, Camille, et al.
(författare)
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Flow-through cross-polarized imaging as a new tool to overcome the analytical sensitivity challenges of a low-dose crystalline compound in a lipid matrix
- 2015
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Ingår i: Journal of Pharmaceutical and Biomedical Analysis. - : Elsevier BV. - 0731-7085 .- 1873-264X. ; 115, s. 20-30
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Tidskriftsartikel (refereegranskat)abstract
- Assessing the physical state of a low-dose active compound in a solid lipid or polymer matrix is analytically challenging, especially if the matrix exhibits some crystallinity. The aim of this study was first to compare the ability of current methods to detect the presence of a crystalline model compound in lipid matrices. Subsequently, a new technique was introduced and evaluated because of sensitivity issues that were encountered with current methods. The new technique is a flow-through version of cross-polarized imaging in transmission mode. The tested lipid-based solid dispersions (SDs) consisted of beta-carotene (BC) as a model compound, and of Gelucire 50/13 or Geleol mono- and diglycerides as lipid matrices. The solid dispersions were analyzed by (hyper) differential scanning calorimetry (DSC), X-ray powder diffraction (XRPD), and microscopic techniques including atomic force microscopy (AFM). DSC and XRPD could analyze crystalline BC at concentrations as low as 3% (w/w) in the formulations. However, with microscopic techniques crystalline particles were detected at significantly lower concentrations of even 0.5% (w/w) BC. A flow-through cross-polarized imaging technique was introduced that combines the advantage of analyzing a larger sample size with high sensitivity of microscopy. Crystals were detected easily in samples containing even less than 0.2% (w/w) BC. Moreover, the new tool enabled approximation of the kinetic BC solubility in the crystalline lipid matrices. As a conclusion, the flow-through cross-polarized imaging technique has the potential to become an indispensable tool for characterizing low-dose crystalline compounds in a lipid or polymer matrix of solid dispersions. (C) 2015 Elsevier B.V. All rights reserved.
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| 10. |
- Amini, Ahmad
(författare)
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Identification of ε-caprolactam, melamine and urea in polyvinylpyrrolidone powders by micellar electrokinetic chromatography
- 2014
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Ingår i: Journal of Pharmaceutical and Biomedical Analysis. - : Elsevier BV. - 0731-7085 .- 1873-264X. ; 91, s. 12-16
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Tidskriftsartikel (refereegranskat)abstract
- A sodium dodecyl sulfate micellar electrokinetic chromatography (SDS-MEKC) method for the simultaneous separation and identification of ɛ-caprolactam, melamine and urea deliberately added to polyvinylpyrrolidone (povidone) products has been developed. All samples to be analyzed contained paracetamol as an internal marker (IM). The optimized separations were performed in 50 mM phosphate buffer (pH 7.0) containing 2% (w/v) sodium dodecyl sulfate (SDS) in fused silica capillaries with UV absorption detection at 200 nm. The method was validated with respect to repeatability and intermediate precision, selectivity and robustness with satisfactory results. The relative migration times (RMT) were found to be between 0.03% and 0.13% for intra-day precision and between 0.50% and 0.60% for inter-day precision in four days. The detection limits were determined to be 1.3 (11.5 μM), 0.4 (3.5 μM) and 41 μg/ml (0.4 mM) for ɛ-caprolactam, melamine and urea, respectively.
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