Ultrafiltration-based Extraction and LC-MS/MS Quantification of Phenylalanine in Human Blood Sample for Metabolite Target Analysis

2020 ◽  
Vol 17 (1) ◽  
pp. 81-86
Author(s):  
Mustafa Çelebier ◽  
Tuba Reçber ◽  
Emirhan Nemutlu ◽  
Sedef Kır
Keyword(s):  
Ischemic Stroke ◽  
Formic Acid ◽  
Flow Rate ◽  
Mobile Phase ◽  
Heavy Isotope ◽  
Column Temperature ◽  
Human Blood Sample ◽  
Linear Concentration Range ◽  
Metabolite Target Analysis ◽  
Linear Concentration

Background: Phenylalanine is a significant biomarker for various diseases like phenylketonuria, gastric cancers, and ischemic stroke according to recent studies. Methods: In the present study; a simple, sensitive, selective and novel analytical method was validated by using an ultrafiltration-based extraction and LC-MS/MS quantification of phenylalanine in human plasma using 13C phenylalanine heavy isotope. Amicon® Ultra Centrifugal Filter was used for ultrafiltration. Parameters affecting LC separation and MS/MS detection were investigated and optimized. Chromatographic separation was achieved on a Merck SeQuant ZIC-HILIC (100x4.6 mm, 5 μm) at a column temperature of 40°C using a mobile phase of mixture of acetonitrile containing 0.1% formic acid and water containing 0.1% formic acid (50:50 v/v) at a flow rate of 0.35 mL/min. The transitions m/z 167→121 for 13C phenylalanine, m/z 166→120 for phenylalanine itself were monitored using the MRM mode. Result: The assay was linear concentration range of 0.0025 μg/mL to 1.20 μg/mL (R2=0.999). The developed method was validated according to FDA guidelines. The method was found linear, sensitive, precise, accurate, and selective.

scholarly journals Quantitative Determination of Azithromycin in Human Plasma by Liquid Chromatography–Mass Spectrometry and its Application in Pharmackokinetic Study

2012 ◽  
Vol 11 (1) ◽  
pp. 55-63 ◽  
Author(s):  
Maizbha Uddin Ahmed ◽  
Mohammad Safiqul Islam ◽  
Tasmin Ara Sultana ◽  
AGM Mostofa ◽  
Muhammad Shahdaat Bin Sayeed ◽  
...  
Keyword(s):  
Mobile Phase ◽  
Solid Phase ◽  
Internal Standard ◽  
Extraction Process ◽  
Serum Samples ◽  
Limit Of Quantification ◽  
Relative Standard ◽  
Linear Concentration Range ◽  
Linear Concentration

Azithromycin is an effective and well-known antimicrobial agent. In the present study, a simple, sensitive and specific LC/MS/MS method has been developed and validated for the quantification of Azithromycin in  human serum samples using Clarithromycin as internal standard. Azithromycin was extracted from biological matrix  by using solid phase extraction process. The chromatographic separation was performed on Luna C18 (3 ?, 2x150   mm) column with a mobile phase consisting of 35 mM ammonium acetate buffer (mobile phase-A) and acetonitrile  and methanol in ratio of 90:10 ( as mobile phase-B) at a flow rate of 0.25 mL/min. The method was validated over a  linear concentration range of 0.5?50.0 ng/mL and limit of quantification (LOQ) was 0.5 ng/mL with a coefficient of  correlation (r2) = 0.9998. The intra-day and inter-day precision expressed as relative standard deviation were 1.64% – 8.43% and 2.32% – 9.92%, respectively. The average recovery of azithromycin from serum was 98.11%. The method  was successfully applied to a pharmacokinetic study after oral administration of Azithromycin 200 mg/5 ml suspension in healthy Bangladeshi volunteers. DOI: http://dx.doi.org/10.3329/dujps.v11i1.12488 Dhaka Univ. J. Pharm. Sci. 11(1): 55-63, 2012 (June)

scholarly journals A New Validated Stability Indicating RP-HPLC Method for Simultaneous Quantification of Formoterol Fumarate Dihydrate and Mometasone Furoate in Metered Dose Inhalation Aerosol

2021 ◽  
Vol 33 (11) ◽  
pp. 2723-2728
Author(s):  
Surya Prakash Mamillapalli ◽  
Gourabattina Lakshmi Prasanna ◽  
B. Venkata Subbaiah ◽  
N. Annapurna
Keyword(s):  
Flow Rate ◽  
Mobile Phase ◽  
Hplc Method ◽  
Mometasone Furoate ◽  
Simultaneous Estimation ◽  
Column Temperature ◽  
Stability Indicating ◽  
Metered Dose ◽  
Formoterol Fumarate ◽  
Dose Inhalation

Stability indicating reversed phase-HPLC method for simultaneous estimation of mometasone furoate (MAF) and formoterol fumarate (FFD) in metered dose inhalation aerosol (MDI) dosage formulation has been developed and discussed in the present work. The chromatographic separation was achieved using Hypersil ODS column (250 mm × 4.6 mm, 3 μm) using an isocratic separation mode at a flow rate of 1.2 mL/min, column temperature of 50 ºC. The system operates with a mobile phase comprising of solution-A (buffer): Solution-B (acetonitrile) mixed in the ratio of 70:30 %v/v at a UV detection wavelength of 214 nm. Retention times of mometasone furoate and formoterol fumarate found to be about 3 min and 7 min, respectively. All possible degradation products of both compounds were monitored at 214 nm and spectral purity along with % mass balance is assessed using PDA detector. Both analyte were subjected to force degradation studies, found all degradants were resolved from analyte peaks and also other process-related impurities. The proposed method is validated for specificity, linearity, accuracy, precision and robustness as per ICH guidelines and found to be adequate. Method stood to be robust with variation in column temperature, flow rate, pH of buffer and organic content in mobile phase.

scholarly journals Development and validation of a rapid selective high-throughput LC-MS/MS method for the determination of triclabendazole sulfoxide concentrations in sheep plasma and its use in bioequivalence studies

2021 ◽  
Author(s):  
Lénárd Farczádi ◽  
Álmos Dósa ◽  
Orsolya Melles ◽  
Laurian Vlase
Keyword(s):  
Formic Acid ◽  
Flow Rate ◽  
Mobile Phase ◽  
Liver Fluke ◽  
Internal Standard ◽  
Gradient Elution ◽  
Sample Processing ◽  
Development And Validation ◽  
Analytical Separation

AbstractTriclabendazole is one of the main drugs used to treat liver fluke in livestock. A rapid LC-MS/MS method was developed and validated to determine ovine plasma levels of triclabendazole sulfoxide.A Gemini NX-C18 column was used to achieve analytical separation, with gradient elution of a mobile phase composed of 0.1% formic acid in acetonitril and 0.1% formic acid in water at flow rate of 0.6 mL/min. MRM with positive ESI ionization was used for the detection of triclabendazole sulfoxide (m/z 360.10 from m/z 376.97). Fenbendazole was used as internal standard. Plasma protein precipitation with acetonitrile was used for sample processing.The method was validated with regards to selectivity, linearity (r > 0.9939), within run and between run precision (CV < 8.9%) and accuracy (bias < 8.9%) over the concentration range 1–100 µg/mL plasma.The method developed is simple, selective and can be applied in bioequivalence and bioavailability studies.

scholarly journals HPLC method for simultaneous determination of impurities and degradation products in Cardiazol

Pharmacia ◽  
2020 ◽  
Vol 67 (1) ◽  
pp. 29-37
Author(s):  
Iryna Drapak ◽  
Borys Zimenkovsky ◽  
Liudas Ivanauskas ◽  
Ivan Bezruk ◽  
Lina Perekhoda ◽  
...  
Keyword(s):  
Flow Rate ◽  
Mobile Phase ◽  
Degradation Products ◽  
Phase 1 ◽  
Simultaneous Detection ◽  
Hplc Method ◽  
Column Temperature ◽  
Degree Of Separation ◽  
Determination Of Impurities

Aim. The aim of study was to develop a simple and accurate procedure that could be applied for the determination of impurities and degradation products in cardiazol. Materials and methods. Separation in samples was carried out with Acquity H-class UPLC system (Waters, Milford, USA) equipped with Acquity UPLC BEH C18 column (2.1 × 50 mm, 1.7 μm) (Waters, Milford, USA). Xevo TQD triple quadrupole mass spectrometer detector (Waters Millford, USA) was used to obtain MS/MS data. Mobile phase A: 0.1% solution of trifluoroacetic acid R in water R; Mobile phase B: acetonitrile R. Samples were chromatographed in gradient mode (Table 1). Flow rate of the mobile phase: 1 ml / min. Column temperature: 30 °С. Detection: at 240 nm wavelength. Injection volume: 10 μl. Results. The retention time of the main substance is about 18.5 minutes. The order of the peak, the retention times and relative retention times: impurity B (12.04, 0.65); impurity А (18.5; 0.98); Cardiazol (18.87; 1.00). The LOD and LOQ values obtained were in the range of 30 ng/mL to 100 ng/mL and 80 ng/mL to 310 ng/mL respectively (with respect to sample concentration of 2 mg/ml). Linearity was established in the range of LOQ level to 0.2% having regression coefficients in the range of 0.9996 to 0.9999. The change in the temperature of the column affects the degree of separation of cardiazol and the impurity A, and thus, with a decrease of 5 ° C, the degree of separation is (1.06), while with increasing this index (3.43). When changing the flow rate of the mobile phase, the degree of separation changes in the following order, with a decrease to 0.9 ml / min separation (1.90), with an increase in speed to 1.1 ml / min (2.45). When the number of mobile phase B decreases by 5%, the degree of separation varies by (2.65), with an increase of 5% (1.82). In comparison with the chromatogram of the tested solution, the substance is not resistant to the action of peroxide, alkaline and acid decomposition. Conclusion. 1) HPLC method was developed and validated for the simultaneous detection and quantitation of impurities formed during the synthesis of cardiazol. 2) The method proved to be sensitive, selective, precise, linear, accurate and stability-indicating.

scholarly journals THE OPTIMIZATION OF HPLC FOR QUANTITATIVE ANALYSIS OF ACID ORANGE 7 AND SUDAN II IN COSMETIC PRODUCTS USING BOX BEHNKEN DESIGN

2019 ◽  
pp. 130-137 ◽  
Author(s):  
NOVALINA BR PURBA ◽  
ABDUL ROHMAN ◽  
SUDIBYO MARTONO
Keyword(s):  
Flow Rate ◽  
Retention Time ◽  
Mobile Phase ◽  
High Performance ◽  
Hplc Method ◽  
Acid Orange 7 ◽  
Column Temperature ◽  
Box Behnken Design ◽  
Acid Orange

Objective: The objective of this study was to optimize high-performance liquid chromatography (HPLC) method for the determination of acid orange 7 (AO7) and sudan II (SII) in blusher product based on response surface methodology using box behnken design (BBD) approach. Methods: Some factors responsible for HPLC separation including column temperature, mobile phase composition, flow rate were optimized using BBD. The responses evaluated were peak area, retention time, and tailing factor. AO7 and SII in blusher product has different properties, therefore both analytes were analysed using C18 column (Thermo Synergy Gold 250 mm x 4.6 mm i.d.,5 µm) using Shimadzu LC 20AD chromatograph equipped with photo-diode array (PDA) detector at 300-650 nm. The mobile phase used was acetonitrile-water (1:1 v/v), and acetonitrile composition was optimized at 35-50% for separation AO7 (ACN1), and 80-90% for SII (ACN2), delivered at the flow rate of 0.9–1 ml/min, using column temperature at 30-40 °C. Results: BBD showed that separation of AO7 was influenced by the concentration of ACN1, flow rate and column temperature. These factors affected retention time, peak area, and tailing factor with peak area was the most significant. Tailing factor was not significantly affected by each factor, and retention time was slightly effected. Otherwise, Sudan II was affected by all these factors except ACN1. The optimal condition obtained based BBD was ACN1 43%, ACN2 90%, the flow rate of 0.9 ml/min and a column temperature of 40 °C. Conclusion: BBD can be used to get optimum condition for analysis of AO7 and SII in blusher product.

Determination of Paeoniflorin and Paeonol in Houyinan Tablet by UPLC

2012 ◽  
Vol 581-582 ◽  
pp. 68-72
Author(s):  
Chu Qin Yu ◽  
Hua Qing Lin ◽  
Yue Han Hou ◽  
Zhong Feng Shi ◽  
Di Shi Lin
Keyword(s):  
Quality Control ◽  
Simultaneous Determination ◽  
Flow Rate ◽  
Mobile Phase ◽  
Gradient Elution ◽  
The Other ◽  
Column Temperature ◽  
Average Recovery ◽  
Injection Volume

In this study, our purpose was to establish a UPLC method for the simultaneous determination of Paeoniflorin and Paeonol in Houyinan Tablet. The separation was performed on Acquity BEH C18 column(2.1mm×100mm,1.7μm), the mobile phase was acetonitrile-water with gradient elution at a flow rate of 0.2 mL•min-1, the detection wavelength was 230nm, the column temperature was 30°Cand the injection volume was 2μL. Paeoniflorin and Paeonol reached effective separation with the other components in this chromatographic conditions. Paeoniflorin and Paeonol were linear within the range of 0.0406~0.4064μg(r=0.9999) and 0.0426~0.4256μg (r=0.9999), respectively. The average recovery was 99.82% and 100.6%. The results of method validation indicated that the method was simple,quick,accurate, specific and less solvent consumption. It can be used for the quality control of Houyinan Tablet.

scholarly journals DEVELOPMENT AND VALIDATION OF A FAST AND SENSITIVE UHPLC-PDA METHOD FOR THE QUANTIFICATION OF URSOLIC ACID IN POLY(L-LACTIC ACID) NANOCAPSULES

2020 ◽  
pp. 161-165
Author(s):  
BRUNA CARLETTO ◽  
AMANDA MARTINEZ LYRA ◽  
ADRIANA YURIKO KOGA ◽  
ANDRESSA NOVATSKI ◽  
RUBIANA MARA MAINARDES ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Ursolic Acid ◽  
Flow Rate ◽  
Mobile Phase ◽  
High Performance ◽  
Limits Of Detection ◽  
Column Temperature ◽  
Relative Standard ◽  
Development And Validation ◽  
Detection And Quantification

Objective: The aim of the present study is to develop and validation of a ultra-high performance liquid chromatography (UHPLC) method to determine the ursolic acid content and its encapsulation efficiency (EE) in lipid-core nanocapsules prepared from poly (L-lactic acid). Methods: A simple UHPLC-PDA method was developed and validated for the quantitative determination of ursolic acid in poly(L-lactic acid) nanocapsules. The chromatographic conditions used were: RP-C18 column, isocratic mobile phase containing acetonitrile:water (92:8, v/v), flow rate of 0.8 ml/min, column temperature of 50°C, and detection at 203 nm. The following parameters were evaluated: Specificity, linearity, limits of detection and quantification, precision, accuracy, and robustness. Results: The method was specific to the ursolic acid and linear (r=0.9998) in the range of 10–100 μg/ml. The limits of detection and quantification were 1.35 and 4.10 μg/ml, respectively. The precision was demonstrated by a relative standard deviation less than 2%. Adequate accuracy (98.35%±0.82) was obtained. Changes in flow rate, mobile phase, and column temperature did not significantly alter the peak area and the retention time of the ursolic acid. The mean EE was 99.89%. Conclusion: The method proved to be fast, sensitive, and simple for quantifying ursolic acid in nanocapsules and was successfully used for determining the EE.

scholarly journals Pharmacokinetics study of potential anti-CML drug Cyclobentinib (CB1107) by HPLC–MS/MS

2021 ◽  
Vol 3 (1) ◽  
pp. 169-175
Author(s):  
Xinghua Zhao ◽  
◽  
Jiaojiao Zhang ◽  
Yutong Liang ◽  
Jie Li ◽  
...  
Keyword(s):  
Formic Acid ◽  
Mobile Phase ◽  
Time Curve ◽  
Column Temperature ◽  
Mobile Phases ◽  
Ionization Reaction ◽  
Liquid Chromatography Tandem Mass ◽  
Different Doses

Purpose: A simple, sensitive and specific HPLC–MS/MS method was established to analysis the pharmacokinetics of CB1107 in mouses. Methods: A simple, selective, and sensitive high-throughput liquid chromatography-tandem mass spectrometry (LC-MS-MS) method has been developed and validated for quantitative determination of CB1107 in rat serum.Chromatographic separation was achieved on a Zorbax Extend C18 Rapid Resolution HD column (4.6 mm × 50 mm, 1.8 μm). The column temperature was maintained at 35℃ and at flow rate of 0.6 mL/min. Injection volume was 20 μL. The mobile phases consisted of 0.1% formic acid in water (mobile phase A)and 0.1% formic acid in acetonitrile (mobile phase B), and total run time was 30min. MS-MS detection was performed in the selected monitoring mode of electrospray positive ionization reaction. Results: The pharmacokinetic characteristics of CB1107 in mice belong to the two-compartment model.When the doses were 400 mg/kg, 600 mg/kg and 800 mg/kg, corresponding area under the plasma concentration-time curve (AUC) respectively were 20.011±1.24 mg/h/L, 26.778±2.19 mg/h/L, 38.82±1.44 mg/h/L, suggesting that CB1107 have a good absorption in the body.And the AUC of three doses are proportional, indicating that CB1107 conforms to linear pharmacokinetics in vivo. Conclusion: This method was successfully applied to study the pharmacokinetics at three different doses of CB1107 after oral administration in mouses. In this study, the bioactivity mechanism of CB1107, by the pharmacokinetic investigation of CB1107 in vivo.

scholarly journals RP-HPLC Method Development and Validation for the Estimation of Lansoprazole in Presence of Related Substances by QbD Approach

2021 ◽  
pp. 138-150
Author(s):  
Sachin B. Gholve ◽  
Jaiprakash N. Sangshetti ◽  
Omprakash G. Bhusnure ◽  
Ram S. Sakhare ◽  
Pratap H. Bhosale ◽  
...  
Keyword(s):  
Flow Rate ◽  
Mobile Phase ◽  
Method Development ◽  
Hplc Method ◽  
Drug Substance ◽  
Gastric Ulcers ◽  
Mobile Phase Flow Rate ◽  
Column Temperature ◽  
Rp Hplc

A rapid specific RP-HPLC method has been developed for the determination of Lansoprazole impurities in the drug substance. The control of pharmaceutical impurities is currently a critical issue in the pharmaceutical industry. The International Council for Harmonization (ICH) has formulated a workable guideline regarding the control of impurities. The objective of the recent study was to develop and validate a HPLC method for the quantitative determination of process-related impurities of Lansoprazole in pharmaceutical drug substance. Lansoprazole, 2-[[[3-methyl-4-(2,2,2-trifluoroethoxy)-2-pyridinyl] methyl]-sulfinyl]- 1H-benzimidazole is an proton pump inhibitor used in the management of gastric ulcers. Chromatographic identification of the impurities was carried out by response surface methodology, applying a three-level Box Behnken design with three center points. Three factors selected were a mobile phase, flow rate, column temperature. Evaluation of the main factor, their interaction, and the quadric effect on peak resolution were done on Waters Symmetry C8, 250 x 4.6mm, 5µm column is used for the development of the method. The mobile phase consists of buffer and acetonitrile. The flow rate of the mobile phase was 1.0 ml/min with gradient elution. The column temperature is ambient and the detection wavelength is 235 nm. The injection volume was 10 µL. The method was validated as per ICH guidelines for linearity in the range of 50-150 µg/ml and the LOD & LOQ values obtained were 0.437×10-4 and 0.1325×10-3 µg/ml respectively which specifies the method's sensitivity. The proposed method was successfully used to determine the Lansoprazole impurities in drug substances.

scholarly journals HPLC method validation for quantification of tetrahydrocurcumin in bulk drug and formulation

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Roopam Raut ◽  
Jessy Shaji
Keyword(s):  
Flow Rate ◽  
Mobile Phase ◽  
Wide Spectrum ◽  
Cost Effective ◽  
Hplc Method ◽  
Uv Detector ◽  
Column Temperature ◽  
Ich Guidelines ◽  
Recovery Study ◽  
Bulk Drug

Abstract Background Tetrahydrocurcumin (THC), the active metabolite of curcumin, is gaining popularity amongst scientist due to its wide spectrum of pharmacological activities, better stability and colourless nature. The objective of this study was to develop a sensitive, cost-effective RP-HPLC method for the estimation of THC in bulk drug substance and formulation. Results Efficient chromatographic separation was achieved on Hypersil BDS, C18 column, 250 mm × 4.6 mm, 5 μm column by isocratic elution with mobile phase comprising of acetonitrile: methanol: water (40:23:37% V/V); adjusted to a pH of 3.0 ± 0.05. The flow rate of the mobile phase was 1.0 ml/min with a column temperature of 25 °C. UV detector was used for the analysis and detection was carried out at 280 nm. The developed method was validated according to ICH guidelines with respect to system suitability, linearity, accuracy, precision and robustness. The theoretical plates were found to be more than 5800. The method showed linearity over the range of 4 to 60 μg/ml with R2 = 0.9998. The accuracy of the method in terms of recovery study was 98.23-99.99%. The %RSD for intra-day and inter-day precision were 0.272 and 0.275, respectively. The method was found to be robust with respect to change in wavelength, flow rate and column temperature. Conclusion The analytical method was found satisfactory on validation as per ICH guidelines. Hence, it can be routinely used for quantification of THC in bulk drug and formulation.

Sign in / Sign up

Export Citation Format

Share Document