scholarly journals Comparative Analysis of Protein Quantification Methods for the Rapid Determination of Protein Loading in Liposomal Formulations

Pharmaceutics ◽  
2019 ◽  
Vol 11 (1) ◽  
pp. 39 ◽  
Author(s):  
Maryam Hussain ◽  
Neil Forbes ◽  
Yvonne Perrie
Keyword(s):  
Small Molecules ◽  
High Performance ◽  
Rapid Determination ◽  
Correlation Coefficients ◽  
Cationic Liposome ◽  
Protein Quantification ◽  
Protein Loading ◽  
Rp Hplc ◽  
Evaporative Light Scattering ◽  
Bca Assay

Advances in manufacturing processes provide the ability for the high throughput production of liposomes containing a range of moieties, from small molecules to large biologicals (including proteins and nucleic acids for prophylactic and therapeutic applications). Whilst rapid quantification methods for small molecules are generally well established, the ability to rapidly quantify liposomal entrapment of proteins is limited. Indeed, most standard protein quantification techniques (including the BCA assay and Reverse phase-high performance liquid chromatography (RP-HPLC)) measure protein encapsulation indirectly, by measuring the amount of non-incorporated drug, and subtracting from the initial amount of protein added. However, this can give inaccurate and misrepresentative results. To address this, we have developed a range of methods to directly quantify protein entrapment within liposomes. The encapsulation efficiency within neutral, anionic and cationic liposome formulations was determined by three techniques; BCA assay, RP-HPLC and HPLC coupled to an evaporative light scattering detector, (HPLC-ELSD). All three methods are reliable for the quantification of protein, with linear responses and correlation coefficients of 0.99, and LOQ for all three methods being less than 10 µg/mL. Here within, we provide three methods for the rapid and robust quantification of protein loading within liposomal (and other bilayer) vesicle systems.

Improved rapid assay of uric acid in serum by liquid chromatography.

1988 ◽  
Vol 34 (12) ◽  
pp. 2567-2568 ◽  
Author(s):  
M Tanaka ◽  
M Hama
Keyword(s):  
Uric Acid ◽  
High Performance ◽  
Rapid Determination ◽  
Correlation Coefficients ◽  
Gel Permeation Chromatography ◽  
Serum Samples ◽  
Step Procedure ◽  
One Step ◽  
Highly Porous

Abstract This improved method for rapid determination of uric acid in serum is based on high-performance liquid-gel-permeation chromatography, with hydrophilic and highly porous vinyl alcohol copolymer as packing material. It has the following advantages: no need for sample deproteinization or use of a precolumn, more than 500 serum samples can be analyzed without having to regenerate or recondition the analytical apparatus, and the analysis for uric acid is a one-step procedure. Correlation coefficients between this method and other methods are very good (r = 0.998, 0.999).

scholarly journals DEVELOPMENT AND VALIDATION OF REVERSE PHASE-HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY TECHNIQUE FOR THE CONCOMITANT ASSESSMENT OF OMEPRAZOLE AND PIPERINE IN BULK FORM

2018 ◽  
Vol 11 ◽  
Author(s):  
Susithra E ◽  
Pavani Ch
Keyword(s):  
High Performance Liquid Chromatography ◽  
Liquid Chromatography ◽  
High Performance ◽  
Correlation Coefficients ◽  
Literature Study ◽  
Uv Detector ◽  
Reverse Phase ◽  
Rp Hplc ◽  
Relative Standard ◽  
Double Beam

Objective: The immense literature study was carried out and disclosed that here no method arrived for the concomitant assessment of omeprazole and piperine in bulk form by using RP-HPLC. Hence, an effort was assembled to arise a easy, specific, precise, reliable, linear, rapid, and validated reverse phase-high-performance liquid chromatography (RP-HPLC) technique for the simultaneous assessment of omeprazole and piperine in bulk form.Methods: The chromatographic analysis of omeprazole and piperine was performed using a RP-HPLC (WATERS) provided with autosampler and ultraviolet (UV) detector with the software of EMPOWER Version 2. The chosen conditions were isocratic separation with two mobile phase composed of acetonitrile:buffer (phosphate buffer: pH 6.5 ± 0.1) (55:45). Detection was carried out using UV/visible double-beam spectrophotometer at 320 nm. The method was validated as per the ICH guidelines.Results: The retention time for omeprazole and piperine by proposed HPLC method was found to be 2.767 and 4.029 min, respectively. The correlation coefficients are 0.999. The developed chromatographic method was found to be accurate with recovery 99.2–99.8% and was found within the acceptance criteria (i.e., 98.0–102.0%) with acceptable % relative standard deviation of not >2% at each level.Conclusion: Thus, the proposed HPLC procedure for the concomitant assessment of omeprazole and piperine was accurate, precise, linear, robust, simple, and economic. 

Analysis of whey proteins during ripening of Afuega'l Pitu cheese by reversed phase HPLC Análisis por RP-HPLC de las proteínas del suero del queso Afuega'l Pitu durante la maduración

1997 ◽  
Vol 3 (6) ◽  
pp. 445-449 ◽  
Author(s):  
D. González de Llano ◽  
G. Santa-María
Keyword(s):  
Protein Content ◽  
Whey Protein ◽  
High Performance ◽  
Whey Proteins ◽  
Correlation Coefficients ◽  
Reversed Phase ◽  
Soluble Form ◽  
Soluble Nitrogen ◽  
Ripening Time ◽  
Rp Hplc

Evolution of the whey proteins fractions and proteolysis indices from 12 batches of the artisan cheese Afuega'l Pitu variety were studied by reversed phase high performance liquid chro matography (RP-HPLC) over the ripening period. The correlation coefficients between the whey protein contents and ripening time were very low, indicating that the whey protein content did not change as the cheese aged. However, soluble nitrogen content and the percentage of total nitrogen in the soluble form showed correlation coefficients higher than 0.9 with increasing ripening time. Analysis of variance showed significant differences ( p < 0.05) between dairies for most of the variables with the exception of β-lactoglobulin A and bovine serum albumin. These variables were also significantly different between the two manufacture zones. Conversely, no significant differences ( p < 0.05) were observed between the two varieties of cheese, red and white, for whey protein content or for proteolysis indices.

scholarly journals A VALIDATED RP-HPLC METHOD FOR SIMULTANEOUS ESTIMATION OF PYRANTEL PAMOATE AND PRAZIQUANTEL IN BULK AND PHARMACEUTICAL DOSAGE FORM

2019 ◽  
pp. 62-67 ◽  
Author(s):  
Rajesh R. ◽  
JITHU JERIN JAMES
Keyword(s):  
High Performance ◽  
Dosage Form ◽  
Correlation Coefficients ◽  
Hplc Method ◽  
Simultaneous Estimation ◽  
Pharmaceutical Dosage Form ◽  
Pyrantel Pamoate ◽  
Rp Hplc ◽  
Ich Guidelines ◽  
Tablet Dosage

Objective: To develop a simple, accurate and precise reverse-phase high-performance liquid chromatography (RP-HPLC) method and subsequently validate for the simultaneous estimation of praziquantel (PZQ) and pyrantel pamoate (PP) in the pharmaceutical dosage form. Methods: The chromatographic separation was achieved on Phenomenex Luna C18 column (250 mm × 4.6 mm, 5 μm) as stationary phase maintained at an ambient temperature with a mobile phase comprising of water: acetonitrile (20: 80) at a flow rate of 1.0 ml/min and UV detection at 220 nm. Results: The retention time of PZQ and PP was found to be 3.897 min and 1.697 min respectively. The method was validated in terms of specificity, accuracy, precision, linearity and robustness as per ICH guidelines. Linearity was obtained in the concentration range of 20–60 μg/ml for both PZQ and PP with correlation coefficients of 0.987 and 0.998 respectively. The accuracy of the method was determined using a recovery test and found as 98.44 % to 100.35 %. All parameters are found to be within the acceptable limit. Conclusion: The developed RP-HPLC method was simple, rapid, accurate, precise for the simultaneous estimation of PZQ and PP in bulk and tablet dosage form.

scholarly journals Method Development and Its Validation for Simultaneous Estimation of Domperidone and Esomeprazole by RP-HPLC in Combination Tablet Dosage Form

2016 ◽  
Vol 1 (01) ◽  
pp. 46-55
Author(s):  
V. Tejasvi Reddy ◽  
S. Hari Hara Kumar ◽  
M Akiful Haque ◽  
Vasudha Bakshi
Keyword(s):  
High Performance ◽  
Method Development ◽  
Limit Of Detection ◽  
Potassium Dihydrogen Phosphate ◽  
Correlation Coefficients ◽  
Simultaneous Estimation ◽  
Dihydrogen Phosphate ◽  
Limit Of Quantification ◽  
Combination Tablet ◽  
Rp Hplc

A simple and validated reverse phase high performance liquid chromatography (RP-HPLC) method was developed for simultaneous estimation of Esomeprazole and Domperidone. Different HPLC Chromatographic conditions were used to find out the optimum chromatographic condition for best elution of drugs. Good chromatographic separation was achieved on Develosil ODS HG-5 RP C18 (15cmX4.6mm, 5m Particle size), analytical column using a mixture of Potassium dihydrogen phosphate buffer pH: acetonitrile in the ratio of 45:55 v/v used as mobile phase at the flow rate of 1 ml/min and detector wavelength at 284nm. The validation of the proposed method was carried out for specificity, linearity and range, accuracy, precision, limit of detection and limit of quantification. Linearity range was found to be 0-140 μg/ml and 0-150 μg/ml with correlation coefficients of 0.995 and 0.994 and retention time of 2.59 and 3.85 min for Domperidone and Esomeprazole respectively. The LOD was found to be 0.341 g/ml and 2.031 g/ml and LOQ was found to be 1.023 g/ml and 6.093 g/ml respectively. The result shows the developed method is yet another suitable method for assay and stability studies which can help in the analysis of Domperidone and Esomeprazole in different formulations.

Development of RP-HPLC Method for Estimation of Valsartan and Hydrochlorothiazide in Tablets

2013 ◽  
Vol 6 (4) ◽  
pp. 2240-2244
Author(s):  
Cylma Menezes ◽  
Vishal M ◽  
Shyamkumar B ◽  
Reema N
Keyword(s):  
High Performance ◽  
Dynamic Range ◽  
Correlation Coefficients ◽  
High Performance Liquid Chromatographic ◽  
Reversed Phase ◽  
Hplc Method ◽  
Simultaneous Estimation ◽  
Rp Hplc ◽  
Validation Parameters ◽  
Liquid Chromatographic

A simple, efficient and reproducible reversed phase high performance liquid chromatographic (RP-HPLC) method was developed and validated for the simultaneous estimation of valsartan and hydrochlorothiazide in bulk and in tablets. A column having 250 x 4.6 mm i.d. (Kromasil C18) in isocratic mode with mobile phase containing 50 mM potassium dihydrogen o-phosphate buffer (triethylamine 0.2%), (pH 3.7 adjusted with o-phosphoric acid): acetonitrile (56:44 v/v) was used. The flow rate was 1.0 ml/min and effluent was monitored at 232 nm.  The retention time of valsartan and hydrochlorothiazide was 10.15 and 3.78 min respectively. All calibration curves showed good linear correlation coefficients within the tested limits (r2 > 0.9995). The linearity dynamic range was found to be 20-150 µg/ml and 5-45 µg/ml for valsartan and hydrochlorothiazide respectively. Percentage recoveries for valsartan and Hydrochlorothiazide were 100.45 % and 98.75 % respectively. All the analytical validation parameters were determined and found in the limit as per the International Conference on Harmonization (ICH) guidelines which indicates the validity of the method. The developed method was found to be accurate, precise and robust for the simultaneous estimation of valsartan and hydrochlorothiazide in bulk and in tablets.

Simultaneous Determination of L-Ascorbic Acid and L-Ascorbylpalmitate Using RP-HPLC Method

2013 ◽  
Vol 716 ◽  
pp. 465-469
Author(s):  
Ye Lin Tian ◽  
Jun Kai Wang ◽  
Ping Sheng Leng ◽  
Yun Liu
Keyword(s):  
Ascorbic Acid ◽  
Simultaneous Determination ◽  
Methyl Palmitate ◽  
High Performance ◽  
Correlation Coefficients ◽  
Reversed Phase ◽  
Hplc Method ◽  
Enzymatic Transesterification ◽  
Rp Hplc

A rapid reversed phase highperformance liquid chromatography (RP-HPLC) method was developed for the simultaneous analysis of L-ascorbic acid and L-ascorbylpalmitate (AP). The chromatography was performed on a SSI model 2300-525 high performance liquid chromatographequipped with anAlltechApollo C18 column at 30 oC.The mobile phase was acetonitrile-water (90:10,v/v) with the flow rate of 1.0 mL/min. UV detection wavelength was 250 nm. This method permits the simultaneous determination of ascorbic acid and AP in the synthesis of AP transesterified with methyl palmitate and ascorbic acid. The detection limit of ascorbic acid and AP was 0.07μg/mL and 0.12μg/mL, respectively. The recovery was 90.59 ± 3.04% for ascorbic acid and 101.3 ± 4.81% for AP. The linearity range for ascorbic acid and AP was in the range of 0.1 - 0.7 mg/mL and 0.4 - 4.0 mg/mL, respectively. Correlation coefficients (R2) were 0.9910 for ascorbic acid and 0.9986 for AP. The proposed method could be used for routine quality control of AP synthesis with methyl palmitate and ascorbic acidby enzymatic transesterification.

Rapid determination of fosetyl-aluminium in commercial pesticide formulations by high-performance liquid chromatography

2014 ◽  
Vol 68 (6) ◽  
Author(s):  
Helen Karasali ◽  
Konstantinos Kasiotis ◽  
Kyriaki Machera
Keyword(s):  
Standard Deviation ◽  
Relative Standard Deviation ◽  
High Performance ◽  
Limit Of Detection ◽  
Rapid Determination ◽  
Hplc Method ◽  
Negative Ion ◽  
Rp Hplc ◽  
Relative Standard

AbstractAn isocratic reversed-phase high-performance liquid chromatographic (RP-HPLC) method with diode array detection (DAD) was developed for the determination of aluminium tris(ethyl phosphonate) (fosetyl-aluminium, fosetyl-Al) in plant-protection products. The method involves extraction of the active ingredient by sonication of the sample with water and direct measurement by RPHPLC. The isocratic RP-HPLC method for the analysis of fosetyl-Al thus developed was then validated for specificity, linearity, precision, and accuracy. The chromatographic peak confirmation was performed by LC-MS using electron spray ionisation in the negative-ion mode. The repeatability of the method, expressed as relative standard deviation (RSD, %), was found to be 0.5 % and the limit of detection was 0.035 mg mL−1. The average recoveries of the three fortification levels varied from 96.7 % to 100.6 % and the RSDs ranged between 2.6 % and 6.3 %. The precision of the method was also considered to be acceptable as the experimental repeatability relative standard deviation (RSDr) was lower than the RSDr, calculated using the Horwitz equation. The method is rapid, simple, accurate, cost-effective, and provides a new and reliable means for the analysis of fosetyl-Al in formulated products.

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