Polyclonal Antibody-Based ELISA and HPLC Methods for the Determination of Fumonisins in Corn: A Comparative Study

1996 ◽  
Vol 59 (8) ◽  
pp. 893-897 ◽  
Author(s):  
ERIC W. SYDENHAM ◽  
SONJA STOCKENSTRÖM ◽  
PIETER G. THIEL ◽  
JOHN P. RHEEDER ◽  
M. BRUNO DOKO ◽  
...  
Keyword(s):  
Comparative Study ◽  
Polyclonal Antibody ◽  
High Performance ◽  
Linear Regression Analysis ◽  
Correlation Coefficients ◽  
Hplc Method ◽  
Enzyme Linked Immunosorbent Assay ◽  
Regression Slopes ◽  
The Individual ◽  
The Comparative Study

The performance of an experimental polyclonal antibody (PAb)-based competitive direct enzyme-linked immunosorbent assay (CD-ELISA) developed for the analysis of fumonisins in corn was assessed by comparison with an established high-performance liquid chromatography (HPLC) method. The comparative study was conducted using a series of 20 corn samples naturally contaminated with combined fumonisin levels ranging from <0.05 to >5 μg/g (ppm). Linear regression analysis between the results generated by HPLC and CD-ELISA provided correlation coefficients (r) and regression slopes (b) of r = 0.960, b = 1.493 (P < 0.001); r = 0.865, b = 3.903 (P < 0.001); and r = 0.832, b = 0.107 (P < 0.001) for the individual fumonisins B1 (FB1), B2 (FB2) and B3 (FB3), respectively, while corresponding values of r = 0.967, b = 1.059 (P < 0.001) were obtained for the combined FB1, FB2, and FB3 concentrations. In 3 of 18 fumonisin-positive corn samples, combined fumonisin levels determined by CD-ELISA were between 85 and 100% higher than those determined in the same extracts by HPLC, while in 13 other samples, CD-ELISA results were between 1.8 and 53% higher than those determined by HPLC. Conversely, in 2 of 18 samples, CD-ELISA results were lower than those determined by HPLC. The differences recorded between HPLC and the experimental PAb-based CD-ELISA were far less than those previously recorded for other mono- and polyclonal antibody-based CD-ELISA systems. The results indicate that the experimental PAb-based CD-ELISA may be effectively applied for the initial screening for fumonisins in corn.

scholarly journals Elisa and HPLC analyses of deoxynivalenol in maize and wheat

2011 ◽  
pp. 25-32 ◽  
Author(s):  
Jovana Matic ◽  
Igor Jajic ◽  
Bojana Saric ◽  
Aleksandra Misan ◽  
Sasa Krstovic ◽  
...  
Keyword(s):  
High Performance ◽  
Screening Method ◽  
Correlation Coefficients ◽  
Hplc Method ◽  
Enzyme Linked Immunosorbent Assay ◽  
Elisa Method ◽  
Masked Mycotoxins ◽  
The Family ◽  
Positive Results ◽  
Hplc Analyses

Deoxynivalenol (DON) is a part of the family of mycotoxins called trichothecenes which are produced by a number of different Fusarium mold species. The presence of DON in 25 wheat and 25 maize samples was examined by Enzyme Linked Immunosorbent Assay (ELISA) and High Performance Liquid Chromatography (HPLC) methods. The presence of DON was detected and determined in 5 (20%) maize and 6 (25%) wheat samples by both of the methods. Correlation between ELISA and HPLC results was established, with the correlation coefficients (r) of 0.9691 and 0.9735 for wheat and maize samples, respectively. The results obtained by ELISA method were significantly higher than those obtained by HPLC method. This fact can be explained by the presence of conjugated or masked mycotoxins in the samples, especially DON-3-glucoside (DON-3-Glc), which could not be determined by HPLC method due to the lack of external standards. Contrary to this, being insufficiently selective towards masked DON, ELISA method measures total DON content of a sample. According to the obtained results, ELISA can be used as a reliable screening method, but the confirmation of positive results must be done by HPLC method.

scholarly journals A comparison between enzyme immunoassay and HPLC for ochratoxin A detection in green, roasted and instant coffee

2007 ◽  
Vol 50 (2) ◽  
pp. 349-359 ◽  
Author(s):  
Simone Fujii ◽  
Elisabete Yurie Sataque Ono ◽  
Ricardo Marcelo Reche Ribeiro ◽  
Fernanda Garcia Algarte Assunção ◽  
Cássia Reika Takabayashi ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Ochratoxin A ◽  
High Performance ◽  
Screening Tool ◽  
Correlation Coefficients ◽  
Enzyme Linked Immunosorbent Assay ◽  
Elisa Assay ◽  
Quality And Safety ◽  
Green Coffee ◽  
Instant Coffee

An indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) for ochratoxin A (OTA) detection in green, roasted and instant coffees was developed using anti-OTA monoclonal antibody. Immunological reagents prepared were OTA-BSA (4.76 mg/mL), anti-OTA.7 MAb (2x10³-fold dilution) and HRP-anti IgG (10³-fold dilution). The detection limit was 3.73 ng OTA/g and correlation coefficients (r) between this immunoassay and high performance liquid chromatography were 0.98 for green coffee, 0.98 for roasted and 0.86 for instant. OTA levels detected by ic-ELISA were higher than by HPLC, with ELISA/HPLC ratio of 0.66 - 1.46 (green coffee), 0.96 - 1.11 (roasted) and 0.93 - 1.82 (instant). ELISA recoveries for OTA added to coffee (5 - 70 ng/g) were 81.53 % for green coffee, 46.73 % for roasted and 64.35 % for instant, while recoveries by HPLC were 80.54 %, 45.91 % and 55.15 %, respectively. Matrices interferences were minimized by samples dilution before carrying out the ELISA assay. The results indicate that MAb-based ic-ELISA could be a simple, sensitive and specific screening tool for OTA detection, contributing to quality and safety of coffee products.

ANALYTICAL METHOD DEVELOPMENT AND VALIDATION FOR THE SIMULTANEOUS ESTIMATION OF ABACAVIR SULPHATE AND LAMIVUDINE IN TABLET DOSAGE FORM BY RP-HPLC

INDIAN DRUGS ◽  
2015 ◽  
Vol 52 (08) ◽  
pp. 12-16
Author(s):  
S Vidyadhara ◽  
◽  
L. S Reddyvalam ◽  
T. Koduri ◽  
P. K. Borra ◽  
...  
Keyword(s):  
High Performance ◽  
Dosage Form ◽  
Method Development ◽  
Correlation Coefficients ◽  
High Performance Liquid Chromatographic ◽  
Hplc Method ◽  
Simultaneous Estimation ◽  
Development And Validation ◽  
Tablet Dosage ◽  
Liquid Chromatographic

A simple, accurate, precise high-performance liquid chromatographic (HPLC) method has been developed and validated for the simultaneous determination of abacavir sulphate (ABA) and lamivudine (LAM) in combined dosage form. Separation was performed on a C18 column [Agilent ODS UG 5 column, 250 mm x 4.5 mm], with methanol: water (50:50 V/V) isocratic elution using a flow rate of 1mL/min. Good sensitivity was observed with UV detection at 277 nm. After method development, the interference of other active compounds and excipients, repeatability and linearity, were investigated. Retention times of LAM and ABA were found to be 3.3 and 6.3 min, respectively. The method was validated over the range from 2.5-12.5 μg/mL for LAM and 5-25 μg/mL for ABA with correlation coefficients of 0.9997 and 0.9996, respectively. This method was shown to be accurate, robust, selective, linear, and repeatable and can be successfully employed in routine quality control for the simultaneous analysis of ABA and LAM in tablets.

Measurement of carbamazepine and its epoxide metabolite by high-performance liquid chromatography, and a comparison of assay techniques for the analysis of carbamazepine.

1977 ◽  
Vol 23 (12) ◽  
pp. 2283-2287 ◽  
Author(s):  
G W Mihaly ◽  
J A Phillips ◽  
W J Louis ◽  
F J Vajda
Keyword(s):  
High Performance Liquid Chromatography ◽  
Liquid Chromatography ◽  
High Performance ◽  
Plasma Concentrations ◽  
Correlation Coefficients ◽  
High Performance Liquid Chromatographic ◽  
Biologically Active ◽  
Gas Liquid Chromatography ◽  
Carbamazepine Therapy ◽  
Regression Slopes

Abstract We describe a modified high-performance liquid-chromatographic method for the simultaneous analysis of carbamazepine andits biologically active metabolite, carbamazepine-10, 11-epoxide. Concentrations of both these compounds in the plasma of 35 epileptic patients receiving chronic carbamazepine therapy are presented. Concentrations of carbamazepine in plasma were related to those of carbamazepine-10, 11-epoxide (r - 0.495, P less than 0.05). Total daily doses of carbamazepine were better correlated with plasma concentrations of carbamazepine-10, 11-epoxide (r = 0.714, P less than 0.001) than of carbamazepine (r = 0.269, P greater than 0.05). Close correlations were found between results of the three assay procedures we used to measure plasma carbamazepine concentrations: high-performance liquid chromatography, gas-liquid chromatography, and enzyme immunoassay. Correlation coefficients exceeded 0.97 and regression slopes were near unity, indicating that all three procedures were individually specific for the quantification of plasma carbamazepine.

Measurement of hemoglobin A1 by liquid chromatography and by agar gel electrophoresis compared.

1981 ◽  
Vol 27 (3) ◽  
pp. 476-479 ◽  
Author(s):  
E J Hayes ◽  
R E Gleason ◽  
J S Soeldner ◽  
M Wacks ◽  
L Blankstein
Keyword(s):  
Liquid Chromatography ◽  
High Performance ◽  
Chromatographic Method ◽  
Linear Regression Analysis ◽  
Hplc Method ◽  
Diabetic Patients ◽  
Sample Population ◽  
Agar Gel ◽  
Electrophoretic Method ◽  
Agar Gel Electrophoresis

Abstract We compare measurement of total fast hemoglobin (HbA1) by "high-performance" liquid chromatography and by electrophoresis on agar gel. Blood samples were obtained from a diverse population (n = 222): offspring of two diabetic parents, diabetic patients with and without retinopathy, diabetic and non-diabetic pregnant women, patients in the coronary-care unit, and normal persons. Precision studies with a normal and an above-normal A1 sample resulted in overall CVs of 9.0% and 4.6% for the electrophoretic method and 4.4% and 2% for the chromatographic method. Linear regression analysis of values for total fast hemoglobin for the complete sample population and for each subgroup showed results of the electrophoretic method to be in excellent agreement with those by the chromatographic method. We conclude that the agar gel electrophoretic method offers a reproducible means for HbA1 determination that is comparable to the HPLC method in terms of accuracy and is highly suited for routine laboratory use.

Determination of Aniline, 4-Aminoazobenzene, and 2-Naphthol in the Color Additive D&C Red No. 17 Using Ultra-High-Performance Liquid Chromatography

2018 ◽  
Vol 101 (6) ◽  
pp. 1961-1966 ◽  
Author(s):  
H H Wendy Yang ◽  
Adrian Weisz
Keyword(s):  
High Performance ◽  
Ammonium Acetate ◽  
Correlation Coefficients ◽  
Hplc Method ◽  
Chromatography Method ◽  
Relative Standard ◽  
Data Points ◽  
Color Additive ◽  
The U.S

Abstract Specifications in the U.S. Code of Federal Regulations for the color additive D&C Red No. 17 (R17, Colour Index No. 26100) limit the levels of the dye’s intermediates, aniline (AN), 2-naphthol (β-naphthol, BN), and 4-aminoazobenzene (4AAB), to 0.2, 0.2, and 0.1%, respectively. The present work reports the development and application of an ultra-HPLC method for the quantitative determination of these impurities in R17. A 1.7 μm particle size C-18 column was used with 0.2 M ammonium acetate and acetonitrile as the eluents. AN, BN, and 4AAB were quantified by using six-point calibration curves with data points (w/w) ranging from 0.01 to 0.25% for AN, 0.01 to 0.24% for BN, and 0.01 to 0.19% for 4AAB. The correlation coefficients ranged from 0.9992 to 0.9999. Limits of detection for the analytes ranged from 0.002 to 0.01%. Recoveries of the analytes ranged from 99.5 to 102%. Relative standard deviations ranged from 0.482 to 1.262%. The new method was applied to analyze portions from 22 batches of R17 submitted to the U.S. Food and Drug Administration for certification. It was found to be simpler to implement, faster, and more sensitive than the older gravity-elution column chromatography method, which it has replaced.

Production and Characterization of Antibodies against Fumonisin B1

1996 ◽  
Vol 59 (9) ◽  
pp. 992-997 ◽  
Author(s):  
FENG-YIR YU ◽  
FUN S. CHU
Keyword(s):  
Detection Limit ◽  
Polyclonal Antibodies ◽  
Fumonisin B1 ◽  
Correlation Coefficients ◽  
Keyhole Limpet Hemocyanin ◽  
Hplc Method ◽  
Enzyme Linked Immunosorbent Assay ◽  
Indirect Competitive Elisa ◽  
Antibody Titers

Polyclonal antibodies against fumonisin B1 (FmB1) were produced in rabbits after immunizing the animals with either FmBl-keyhole limpet hemocyanin (KLH) or FmB1 bovine serum albumin (BSA). A direct competitive enzyme-linked immunosorbent assay (dc-ELISA) and an indirect competitive ELISA (idc-ELISA) were used for the characterization of the antibodies and for analysis of the toxin in corn samples. The antibody titers in the serum of rabbits immunized with FmBl-KLH were considerably higher than in those immunized with FmBl-BSA. The antibodies from the rabbits immunized with FmBl-KLH were further characterized. The concentrations causing 50% inhibition of binding of FmB1-horseradish peroxidase (HRP) to the antibodies by FmB1, FmB2 and FmB3 in the ELISA were found to be 0.45, 0.72, and 25 ng/ml, respectively. The detection limit of FmBl, based on 95% confidence at 5% of inhibition of binding of FmBl-HRP conjugate, in buffer of the dc-ELISA was found to be 0.05 ng/ml. In the presence of a matrix such as corn, the detection limit was less than 50 ppb. The overall analytical recoveries of FmBl (50 to 1,000 ng/g) added to the ground corn and then extracted with CH3CN/H2O (1/1, vol/vol) with cleanup and without cleanup in the dc ELISA were found to be 70.5 and 85.9%, respectively. A good correlation was found between the FmBl levels in 2 starch and 10 naturally contaminated corn samples analyzed by the dc-ELISA and the high-pressure liquid chromatography (HPLC) method. The correlation coefficients between ELISA and HPLC were found to be 0.955 (y [ELISA] = 1.3 1x [HPLC] + 77 ppb; P < 0.001) and 0.811 (y = 1.13x + 34 ppb; P < 0.01) for the sample without and with cleanup treatment, respectively.

High-Performance Liquid-Chromatographic Measurement of Plasma Creatinine in Newborns

1992 ◽  
Vol 38 (1) ◽  
pp. 101-103 ◽  
Author(s):  
Paul H Scott
Keyword(s):  
High Performance ◽  
Linear Regression Analysis ◽  
Internal Standard ◽  
High Performance Liquid Chromatographic ◽  
Hplc Method ◽  
Exchange Chromatography ◽  
Plasma Creatinine ◽  
Standard Curve ◽  
Analytical Recovery ◽  
Liquid Chromatographic

Abstract This HPLC method for measuring plasma creatinine is based on cation-exchange chromatography and is particularly suitable for use with specimens from babies. A short chromatographic run is performed after simple protein precipitation with zinc sulfate and addition of an internal standard, N-methylnicotinamide. The standard curve for the method is linear up to 200 mumol/L, and analytical recovery of added creatinine is between 101% and 103%. Between-batch precision (CV) is less than 3% for mean creatinine values of 103 and 164 mumol/L. The method is free of interference from other metabolic components and drugs commonly used in neonates in routine clinical practice. Using specimens from neonates, I compared this method with a routinely used automated alkaline picrate method (from Randox Labs., performed on a Cobas MIRA analyzer). Linear-regression analysis yielded a correlation coefficient of 0.90, a slope of 1.00, and an intercept of +0.8 mumol/L. This HPLC method for creatinine should be of use in those circumstances where the alkaline picrate method is known to produce dubious results; however, the latter method is probably more suitable for routine use.

scholarly journals A comparative study concerning chromatographic retention and computed partition coefficients of some precursors of peraza crown ethers

2010 ◽  
Vol 8 (6) ◽  
pp. 1203-1209 ◽  
Author(s):  
Cristina Onişor ◽  
Gabriela Blăniţă ◽  
Maria Coroş ◽  
Monica Bucşa ◽  
Mircea Vlassa ◽  
...  
Keyword(s):  
Comparative Study ◽  
Crown Ethers ◽  
Partition Coefficients ◽  
High Performance ◽  
Correlation Coefficients ◽  
Reversed Phase ◽  
Neural Network Modeling ◽  
Log P ◽  
Chromatographic Retention ◽  
P Values

AbstractRetention indices for some precursors of peraza crown ethers were determined by reversed phase high-performance thin layer chromatography on RP-18 plates with methanol-water in different volume proportions as mobile phase. The Log P values for the same compounds were calculated using different computer programs: SciQSAR, SciLogP, Chem3D Ultra 8.0, XLOGP (based on atom contributions), Chemaxon and KOWWIN (based on atom/fragment contributions), cLogP (based on fragmental contributions), ALOGPS and IAlogP (based on atom-type electrotopological-state indices and neural network modeling). A comparative study concerning lipophilic parameters (RM0, b and ϕ0) and computed partition coefficients has been developed. Taking into account the correlation coefficients between determined and calculated Log P values, it seems that RM0 and b are less suitable than ϕ0 for estimating lipophilicity of the compounds investigated, and cLogP and ALOGPS provide the best correlations with experimental values.

Analysis of spectinomycin in fermentation broth by reversed-phase chromatography

2009 ◽  
Vol 63 (6) ◽  
Author(s):  
Hong Yan ◽  
Pei Xu ◽  
Hai Huang ◽  
Juan Qiu
Keyword(s):  
High Performance ◽  
Limit Of Detection ◽  
Fermentation Broth ◽  
Correlation Coefficients ◽  
High Performance Liquid Chromatographic ◽  
Reversed Phase ◽  
Hplc Method ◽  
Standard Curve ◽  
Relative Standard ◽  
Liquid Chromatographic

AbstractA pre-column derivatized high-performance liquid chromatographic (HPLC) method with ultraviolet-visible detection was developed to measure the concentrations of spectinomycin in fermentation broth. Derivatization reagents, 2,4-dinitrophenylhydrazine in acetonitrile (5 mg mL−1) and trifluoroacetic acid in acetonitrile (0.8 mol L−1), were added to an aliquot of the fermentation broth, and the mixture was incubated for 60 min at 70°C. The resulting derivative was separated from other compounds by isocratic elution in a reversed-phase column Zorbax SB-C18 (250 mm × 4.6 mm, 5 µm). Mobile phase consisted of acetonitrile, tetrahydrofuran, and water (φ r = 40: 35: 25) and the flow rate was 1.0 mL min−1. The detection wavelength was 415 nm. The standard curve for spectinomycin sulfate was linear with correlation coefficients of 0.9997 in the range of 25 µg mL−1 to 600 µg mL−1. The relative standard deviation values ranged from 0.43 % to 2.18 % depending on the concentration of samples. The average recovery was 101.5 %. The limit of detection was 50 ng mL−1.

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