Considerations in the preparation of laboratory samples for the analysis of ochratoxin A in wheat

2012 ◽  
Vol 5 (2) ◽  
pp. 107-116 ◽  
Author(s):  
S.A. Tittlemier ◽  
M. Roscoe ◽  
C. Kobialka ◽  
R. Blagden
Keyword(s):  
Ochratoxin A ◽  
Whole Grain ◽  
Laboratory Sample ◽  
Test Portion ◽  
Homogeneous Sample ◽  
Ambient Temperatures ◽  
Accuracy And Precision ◽  
Relative Standard ◽  
The Mean ◽  
Ground Wheat

A process used to prepare the test portion of ground wheat from the whole grain laboratory sample for ochratoxin A (OTA) analysis using dry comminution with homogenisation and sub-sampling via a rotary sample divider was developed and evaluated. With respect to OTA content, the developed process produced a homogeneous sample of ground wheat from 10 kg of whole grain. Relative standard deviations of the mean OTA concentration for five naturally contaminated wheat samples processed using the developed method ranged from 9% to 19% over a relevant concentration range of 1.7 to 7.6 mg/kg. Additional studies demonstrated that OTA was stable in ground wheat with moisture content between 12 to 13% for at least a year when stored at ambient temperatures. Further examination of the developed comminution and dividing procedure demonstrated that higher concentrations were measured in smaller sized particles, indicating that the accuracy and precision of OTA analyses could be affected by the particle size of ground wheat.

An alternative to slurry mixing to minimise sample preparation variance for determination of ochratoxin A in wheat

2010 ◽  
Vol 3 (2) ◽  
pp. 147-156 ◽  
Author(s):  
T. Nowicki ◽  
M. Roscoe
Keyword(s):  
Sample Preparation ◽  
Ochratoxin A ◽  
Mixing Time ◽  
Test Sample ◽  
Laboratory Sample ◽  
Analytical Process ◽  
Accuracy And Precision ◽  
Primary Sample ◽  
Surveillance Programs ◽  
Ground Wheat

Many countries have established maximum limits for ochratoxin A (OTA) in cereal grains and implemented surveillance programs for OTA in wheat shipments. For shipment to some countries, certification for OTA content is mandatory. These control activities require the capability to measure OTA in bulk grain shipments with accuracy and precision. It is known that the nugget effect caused by the heterogeneous nature of mycotoxin contamination in agricultural commodities creates major challenges for generating representative test results due to the potential for high variances in the sampling / sample preparation phases of the analytical process. The water-slurry mixing approach to sample preparation has greatly minimises variances associated with this phase of the analytical process, but this is not a practical technique for all laboratories. The potential magnitude of variances for subsampling raw and ground grain for the dry-milling approach to sample preparation and the means for reducing variances to acceptable levels is not fully understood. We investigated the repeatability of OTA measurements in 2 kg laboratory samples subsampled from 20 kg samples of raw wheat and in 100 g test portions subsampled from 2 kg of ground wheat. In addition, the effect of mixing time on the repeatability of OTA results was investigated prior to subsampling to obtain test portions for analysis. Results show that for subsampling of a primary sample of raw wheat using a conventional sample divider the variability of OTA results decreases with increasing weight of the laboratory sample relative to the weight of the primary sample. In order to improve repeatability, the proportion of primary sample separated out to produce a laboratory sample should be as large as operationally feasible and ideally about 50% of the weight of the primary sample. Four factors are identified for separating out a test sample from a raw wheat laboratory sample.

Comparison of Liquid Chromatographic Method to AOAC Microbiological Method for Determination of L-Tryptophan in Tablets and Capsules

1993 ◽  
Vol 76 (2) ◽  
pp. 414-417 ◽  
Author(s):  
Henry S Kim ◽  
Gerald Angyal
Keyword(s):  
Reversed Phase ◽  
Precolumn Derivatization ◽  
Microbiological Method ◽  
Test Portion ◽  
Liquid Chromatographic Method ◽  
Relative Standard ◽  
Phase Liquid ◽  
The Mean ◽  
Liquid Chromatographic

Abstract A reversed-phase liquid chromatographic (LC) method coupled with precolumn derivatization of L-tryptophan with phenylisothiocyanate was compared to the AOAC microbiological method for determining L-tryptophan in tablets and capsules. For the microbiological method, the concentrations of L-tryptophan were 4-8% lower in autoclaved test samples (hot method) than in test samples that were not autoclaved (cold method). When L-tryptophan values obtained by the LC method were compared to those obtained by the cold microbiological method, no significant differences were observed (P > 0.05). The mean relative standard deviations were 2.9% for the LC method and 1.6% for the cold microbiological method. The mean recoveries of standard L-tryptophan added before analysis were 99% for the LC method and 101 % for the cold microbiological method. These results demonstrate that both methods are reliable for determining free L-tryptophan contained in tablets and capsules. However, the LC method has the advantages of using a smaller test portion and having a shorter analysis time.

scholarly journals Combined Phenyl Silane and Immunoaffinity Column Cleanup with Liquid Chromatography for Determination of Ochratoxin A in Roasted Coffee: Collaborative Study

2001 ◽  
Vol 84 (2) ◽  
pp. 444-450 ◽  
Author(s):  
A Catherine Entwisle ◽  
Alison C Williams ◽  
Peter J Mann ◽  
Joanne Russell ◽  
Philip T Slack ◽  
...  
Keyword(s):  
Liquid Chromatography ◽  
Standard Deviation ◽  
Ochratoxin A ◽  
Relative Standard Deviation ◽  
Collaborative Study ◽  
Immunoaffinity Column ◽  
Roasted Coffee ◽  
Test Portion ◽  
Low Level ◽  
Relative Standard

Abstract A collaborative study was conducted to evaluate a liquid chromatography (LC) method for ochratoxin A using sequential phenyl silane and immunoaffinity column cleanup. The method was tested at 3 different levels of ochratoxin A in roasted coffee, which spanned the range of possible future European regulatory limits. The test portion was extracted with methanol and sodium bicarbonate by shaking for 30 min. The extract was filtered, centrifuged, and then cleaned up on a phenyl silane column before being eluted from the washed column with methanol–water. The eluate was diluted with phosphate-buffered saline (PBS) and applied to an ochratoxin A immunoaffinity column, which was washed with water. The ochratoxin A was eluted with methanol, the solvent was evaporated, and the residue was redissolved in injection solvent. After injection of this solution onto a reversed-phase LC apparatus, ochratoxin A was measured by fluorescence detection. Eight laboratory samples of low-level naturally contaminated roasted coffee and 2 laboratory samples of blank coffee (< 0.2 ng/g ochratoxin A at the signal-to-noise ratio of 3:1), along with ampules of ochratoxin A calibrant and spiking solutions, were sent to 15 laboratories in 13 different European countries. Test portions of the laboratory samples were spiked at levels of 4 ng/g ochratoxin A, and recoveries ranged from 65 to 97%. Based on results for spiked blank material (blind duplicates) and naturally contaminated material (blind duplicates at 3 levels), the relative standard deviation for repeatability (RSDr) ranged from 2 to 22% and the relative standard deviation for reproducibility (RSDR) ranged from 14 to 26%. The method showed acceptable within- and between-laboratory precision, as evidenced by HORRAT values, at the low level of determination for ochratoxin A in roasted coffee.

scholarly journals Determination of Aflatoxins and Ochratoxin a in Animal Liver Using HPLC-FD Method with Immunoaffinity Column Clean-Up

2021 ◽  
Vol 0 (0) ◽  
Author(s):  
Biljana Stojanovska-Dimzoska ◽  
Elizabeta Dimitrieska-Stojkovic ◽  
Zehra Hajrulai-Musliu ◽  
Risto Uzunov ◽  
Aleksandra Angeleska ◽  
...  
Keyword(s):  
Liquid Chromatography ◽  
Ochratoxin A ◽  
Immunoaffinity Column ◽  
Acceptable Range ◽  
Animal Liver ◽  
Relative Standard ◽  
Validation Parameters ◽  
The Mean ◽  
Concentration Levels

Abstract Analytical methods based on immunoaffinity column clean-up and quantitative determination with liquid chromatography-fluorescence detection were used to determine aflatoxins and ochratoxin A in liver samples. The validation of the procedures was performed. The linearity of the methods was checked, and a good coefficient of correlation was found for all aflatoxins and OTA as well. The LOD and LOQ were acceptable: 0.003 µg/kg and 0.009 µg/kg for AFB1; 0.001 µg/kg and 0.005 µg/kg for AFB2; 0.006 µg/kg and 0.020 µg/kg for AFG1; 0.007 µg/kg and 0.022 µg/kg for AFG2; 0.08 µg/kg and 0.27 µg/kg for OTA. The results for the repeatability estimated by the relative standard deviation (RSDr) were satisfactory and the obtained values were in the acceptable range (1.97–14.41% for all aflatoxins and 3.76-8.31% for OTA) at three proposed concentration levels. RSDR values showed acceptable correlation between two analysts for all four aflatoxins and OTA. The RSDR values were as followed: 2.37% and 5.60% for AFB1, 6.71% and 8.78% for AFB2, 4.40% and 7.00% for AFG1 and 10.30% and 13.91% for AFG2 (for the first and second analyst, respectively). The RSDR values for OTA were 4.91% and 3.15% (1 µg/kg); 3.76% and 4.12% (5 µg/kg) and 8.31% and 8.21% (10 µg/kg). The mean recovery for total aflatoxins and OTA were 78.10% and 93.34%, respectively. All validation parameters were in accordance to European legislation. They indicate that the proposed analytical procedures are suitable and they could be methods of choice for the determination of aflatoxins and OTA in liver samples.

scholarly journals Analysis of ochratoxin A in malt beverage samples using dispersive liquid–liquid microextraction coupled with liquid chromatography-fluorescence detection

2013 ◽  
Vol 31 (No. 5) ◽  
pp. 520-525 ◽  
Author(s):  
M. Maham ◽  
V. Kiarostami ◽  
S. Waqif-Husain ◽  
R. Karami-Osboo ◽  
M. Mirabolfathy
Keyword(s):  
Liquid Chromatography ◽  
Ochratoxin A ◽  
Fluorescence Detection ◽  
High Performance ◽  
Optimum Conditions ◽  
Beverage Samples ◽  
Extraction Solvent ◽  
Relative Standard ◽  
The Mean ◽  
Liquid Microextraction

A simple and economic procedure based on dispersive liquid–liquid microextraction has been applied to extract and pre-concentrate trace levels of ochratoxin A (OTA) in malt beverage prior to analysis using high performance liquid chromatography with fluorescence detection. The method was based on the formation of fine droplets of a water-immiscible extraction solvent in the sample solution using a water-miscible disperser solvent. The influences of various parameters such as the type and volume of extraction and disperser solvents, centrifuging time, sonication time, and salt concentration on the extraction efficiency of ochratoxin A were investigated. Under optimum conditions, the relative standard deviations for five replicates of 2 ng/ml of OTA were 3.4% as within-day and 6.2% as between-day precisions. The detection limit (S/N = 3) was 0.1 ng/ml and the mean recoveries of OTA from malt beverage samples at spiking levels of 0.5, 2, and 4 ng/ml were in the range of 104–108.2%.

Development and inter-laboratory study of a method for quantifying ochratoxin A in pet foods

2017 ◽  
Vol 10 (1) ◽  
pp. 53-61
Author(s):  
T. Wada ◽  
H. Saito ◽  
K. Aoyama ◽  
S. Saito ◽  
M. Shibukawa
Keyword(s):  
High Performance Liquid Chromatography ◽  
Liquid Chromatography ◽  
Ochratoxin A ◽  
Laboratory Study ◽  
High Performance ◽  
Immunoaffinity Column ◽  
Pet Food ◽  
Aqueous Acetonitrile ◽  
Relative Standard ◽  
The Mean

An analytical method for quantifying ochratoxin A (OTA) in pet foods using high-performance liquid chromatography was developed, and an inter-laboratory study was conducted. OTA was extracted from samples with aqueous acetonitrile. The extract was purified by an immunoaffinity column, OCHRAKING, and analysed by high performance liquid chromatography with fluorescence detection. The limits of quantification by this method were 2 µg/kg for dry and semidry pet food and 1 µg/kg for wet type pet food. The calibration curve showed linearity in the range of 0.5-50 ng/ml (equivalent to 1-100 µg/kg for wet type pet food). The mean recoveries of OTA spiked at 1-5 µg/kg were in the range of 83.0-106% and relative standard deviations of the in-house method validation were 2.6-6.8%. The mean recoveries, repeatability, reproducibility and the Horwitz ratios for OTA from the inter-laboratory validation study were 75.6-83.1%, 3.5-6.1%, 5.0-15.0% and 0.23-0.68, respectively.

Variability and distribution among sample test results when sampling unprocessed wheat lots for ochratoxin A

2016 ◽  
Vol 9 (2) ◽  
pp. 163-178 ◽  
Author(s):  
T.B. Whitaker ◽  
A.B. Slate ◽  
T.W. Nowicki ◽  
F.G. Giesbrecht
Keyword(s):  
Liquid Chromatography ◽  
Sample Preparation ◽  
Ochratoxin A ◽  
Test Procedure ◽  
Test Results ◽  
Laboratory Sample ◽  
Sampling Step ◽  
Test Portion ◽  
Sample Test ◽  
Sampling Statistics

In 2008, Health Canada announced it was considering the establishment of maximum levels for ochratoxin A (OTA) in unprocessed wheat, oats, and their products. The Canada Grains Council and Canadian National Millers Association initiated two studies to measure the variability and distribution among sample test results for unprocessed wheat and oats so that scientifically based OTA sampling plans could be designed to meet regulatory and industry requirements. Sampling statistics related to detecting OTA in oats has been published. 54 OTA contaminated wheat lots representing three wheat classes were identified for the sampling study. Each lot was sampled according to a nested experimental protocol where sixteen 2-kg laboratory samples were taken from each lot, multiple 5-g test portions were taken from each comminuted 2-kg laboratory sample, and multiple OTA measurements were made on each test portion using liquid chromatography. The sampling, sample preparation, and analytical variances associated with each step of the OTA test procedure were found to be a function of OTA concentration and regression equations were developed to predict the functional relationships between variance and OTA concentration. When sampling a wheat lot containing 5 µg/kg OTA with an OTA test procedure consisting of a sampling step employing a single 2-kg laboratory sample, sample preparation step employing a single 100-g test portion, and an analytical step that used liquid chromatography to quantify OTA, the sampling step accounted for 95.3% of the total variability. The observed OTA distribution among the 16 OTA sample results was found to be positively skewed and the negative binomial distribution was selected to model the OTA distribution among sample test results. The sampling statistics were incorporated into the FAO Mycotoxin Sampling Tool and the chances of rejecting good lots and accepting bad lots were calculated for various sampling plan designs.

Development of Guidelines for Accurate Measurement of Small Volume Parenteral Products Using Syringes

2019 ◽  
pp. 001857871987386
Author(s):  
Melanie A. Jordan ◽  
Dimpa Choksi ◽  
Kelsey Lombard ◽  
Lynn R. Patton
Keyword(s):  
Standard Deviation ◽  
Small Volume ◽  
Selection Methods ◽  
Accuracy And Precision ◽  
Relative Standard ◽  
Volume Measurements ◽  
The Mean ◽  
Precision And Accuracy ◽  
Parenteral Products ◽  
Sterile Product

Background: Syringes are commonly used in pharmacy compounding for the measurement of small volumes, especially in the preparation of sterile products for injection and infusion. However, there are no current official guidelines for the proper use of syringes in measuring small volumes. Objective: The purpose of this project was to determine the accuracy and precision of commercially available syringes in measuring small volumes during sterile product preparation to make recommendations for syringe size selection. Methods: To assess precision and accuracy of syringes, 3 separate investigators measured 5%, 10%, or 20% (n = 30 each) of the volume of a 1-, 3-, 5-, 10-, or 20-mL syringe with an attached 18G, 1½” needle by drawing sterile water for injection from a vial. Delivered volumes were measured gravimetrically using an electronic balance and converted to volume using the specific gravity of water (1.0). Accuracy is represented as the mean and standard deviation, while precision is represented as percent relative standard deviation. Differences were assessed using a 1-way analysis of variance with Bonferroni adjustments and significance set at P < .05. Results: Precision and accuracy were highly variable and often significantly ( P < .05) different compared to the theoretical volume delivered both within and between investigators. An increased likelihood of unacceptable error (>5%) was observed when less than 20% of the labeled capacity of a syringe was measured. Mean percent error ranged from 1.4% to 18.6%, despite manufacturer specification of ±5% accuracy, suggesting proper technique as a major factor in small-volume measurements. Conclusion: In addition to proper, validated training of syringe users, we recommend that users measure no less than 20% of the indicated volume of the syringe while choosing syringes as close as possible to the desired measurement. When possible, very small volumes should be diluted to meet the minimum volume of the smallest syringe available. Implementation of these recommendations will improve accurate dosing and, ultimately, patient safety.

scholarly journals Determination of Ochratoxin A in Currants, Raisins, Sultanas, Mixed Dried Fruit, and Dried Figs by Immunoaffinity Column Cleanup with Liquid Chromatography: Interlaboratory Study

2003 ◽  
Vol 86 (6) ◽  
pp. 1164-1171 ◽  
Author(s):  
Susan J MacDonald ◽  
Sharron Anderson ◽  
Paul Brereton ◽  
Roger Wood ◽  
G Barrett ◽  
...  
Keyword(s):  
Standard Deviation ◽  
Ochratoxin A ◽  
Relative Standard Deviation ◽  
Reversed Phase ◽  
Interlaboratory Study ◽  
Affinity Column ◽  
Test Portion ◽  
Relative Standard ◽  
Dried Fruit

Abstract An interlaboratory study was performed on behalf of the Food Standards Agency to evaluate the effectiveness of an affinity column cleanup liquid chromatographic (LC) method for the determination of ochratoxin A in a variety of dried fruit at European regulatory limits. To ensure homogeneity before analysis, laboratory samples are normally slurried with water in the ratio of 5 parts fruit to 4 parts water, and test materials in this form were used in the study. The test portion was extracted with acidified methanol. The extract was filtered, diluted with phosphate-buffered saline, and applied to an affinity column. The column was washed and ochratoxin A was eluted with methanol. Ochratoxin A was quantified by reversed-phase LC. The use of post-column pH shift to enhance the fluorescence of ochratoxin A by the addition of 1.1M ammonia solution to the column eluant is optional. Determination was by fluorescence. Currants, sultanas, raisins, figs, and mixed fruit (comprising dried pineapple, papaya, sultanas, prunes, dates, and banana chips), both naturally contaminated and blank (very low level), were sent to 24 collaborators in 7 European countries. Participants were asked to spike test portions of all test samples at a level equivalent to 5 ng/g ochra toxin A. Average recoveries ranged from 69 to 74%. Based on results for 5 naturally contaminated test samples (blind duplicates) the relative standard deviation for repeatability (RSDr) ranged from 4.9 to 8.7%, and the relative standard deviation for reproducibility (RSDR)rangedfrom14to28%. The method showed acceptable within-and be-tween-laboratory precision for all 5 matrixes, as evidenced by HORRAT values &lt;1.3.

scholarly journals Determination of Ochratoxin A in Green Coffee by Immunoaffinity Column Cleanup and Liquid Chomatography: Collaborative Study

2005 ◽  
Vol 88 (3) ◽  
pp. 773-779 ◽  
Author(s):  
Eugênia Azevedo Vargas ◽  
Eliene Alves dos Santos ◽  
Alain Pittet ◽  
T B S Corrêa ◽  
A P P da Rocha ◽  
...  
Keyword(s):  
Standard Deviation ◽  
Ochratoxin A ◽  
Relative Standard Deviation ◽  
Collaborative Study ◽  
Green Coffee ◽  
The European Union ◽  
Immunoaffinity Column ◽  
Test Portion ◽  
Relative Standard

Abstract A collaborative study was conducted to evaluate a method using immunoaffinity column cleanup with liquid chromatography (LC) for the determination of ochratoxin A (OTA) in green coffee at levels that could be included in possible future regulations of the European Union. The test portion was extracted with methanol–3% aqueous sodium hydrogen carbonate solution (50 + 50, v/v). The extract was filtered, and the filtrate was diluted with phosphate-buffered saline and applied to an immunoaffinity column containing antibodies specific for OTA. After washing, the toxin was eluted from the column with methanol and quantified by LC with fluorescence detection. Pairs of 4 homogeneous noncontaminated and naturally contaminated materials (mean levels of &lt;0.12, 2.44, 5.15, and 13.46 ng/g) and blank samples (&lt;0.12 ng/g) for spiking were sent to 20 participant laboratories from 8 countries. The materials were analyzed according to the method description and all difficulties encountered in the analysis were reported. Statistical analysis was carried out according to the Harmonized Protocol of the International Union of Pure and Applied Chemistry. The relative standard deviation for repeatability (RSDr) ranged from 7.42 to 20.94%, and the relative standard deviation for reproducibility (RSDR) ranged from 16.34 to 29.17%. The method showed acceptable within-laboratory and between-laboratories precision for green coffee materials, as evidenced by HorRat values of ≤0.85, at the studied range, for spiked and naturally contaminated materials. The mean recovery was 92.8% for green coffee material spiked with OTA at a level of 4.82 ng/g.

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