Results of a European inter-laboratory comparison study on the determination of EU priority polycyclic aromatic hydrocarbons (PAHs) in edible vegetable oils

A collaborative study on the analysis for 15 + 1 EU priority PAHs in edible oils was organised to investigate the state-of-the-art of respective analytical methods. Three spiked vegetable oils, one contaminated native sunflower oil, and one standard solution were investigated in this study. The results of 52 laboratories using either high-performance liquid chromatography with fluorescence detection or gas chromatography with mass-selective detectors were evaluated by application of robust statistics. About 95% of the laboratories were able to quantify benzo[a]pyrene together with five other PAHs included in the commonly known list of 16 US-EPA PAHs. About 80% of the participants also quantified seven additional PAHs in most samples, two of which were benzo[b]fluoranthene and benzo[k]fluoranthene, which were also known from the EPA list. Only about 50% of the participants quantified cyclopenta[cd]pyrene, benzo[j]fluoranthene, and benzo[c]fluorene. The robust relative standard deviations of the submitted results without discrimination between the methods applied ranged between 100% for 5-methylchrysene in spiked olive oil and 11% for the same analyte in spiked sunflower oil. The results clearly showed that for these analytes the methods of analysis are not yet well established in European laboratories, and more collaborative trials are needed to promote further development and to improve the performances of the respective methods.     

Determination of low-level glucose and fructose in raw and refined crystalline sugar by high-performance anion-exchange chromatography: collaborative study

A method was developed and a collaborative study was performed under the auspices of the International Commission of Uniform Methods for Sugar Analysis. The collaborators used high-performance anion-exchange chromatography (HPAEC) to determine trace amounts of glucose and fructose in 3 raw and 3 refined sugar samples provided as blind duplicates. Fourteen laboratories participated in the study. Although difficulties were experienced by a few analysts, 10 laboratories reported positive results. Average repeatabilities and reproducibilities for glucose and fructose in raw sugar were slightly above 5 and 10%, respectively, and the average Horwitz ratios were well under 2. Average repeatabilities and reproducibilities for glucose and fructose in refined sugar were 10 and 22%, respectively; although the Horwitz ratios were >2, they were marginal (2.8). The HPAEC results agreed with results obtained by gas chromatography in an independent laboratory. Because the method was collaboratively studied according to the protocol of the International Union of Pure and Applied Chemistry for collaborative studies, and the results meet its criteria, it is recommended that the method be adopted Official First Action by AOAC INTERNATIONAL.     

Determination of Nitrilotriacetic Acid in Waters and Waste Waters by Gas-Liquid Chromatography,Differential Pulse Polarography and a Colorimetric Method

Abstract

A comparison of gas-liquid chromatography, differential-pulse polarography and a colorimetric method for the determination of nitrilotriacetic acid in settled sewage, sewage effluent, potable water, soil extracted water and deionised water has been undertaken. Differential-pulse polarography has also been applied to the determination of nitrilotriacetic acid in saline samples. By statistical analysis of replicate determinations, accuracy and precision have been evaluated, and calibration linearity assessed. Interferences were observed for sewage samples when analysed by all three methods. Precision was generally higher for differential-pulse polarography down to 100 μgl^?1, although only gas-liquid chromatography is applicable to concentrations below 25μgl^?1 in non-saline samples. The colorimetric method was not applicable to concentrations below 500 μgl^?1 of nitrilotriacetic acid.     

Micro-determination of total phthalate esters in biological samples by gas-liquid chromatography.

A method was investigated in which all of the phthalate esters in biological samples were determined as phthalic acid by gas-liquid chromatography. The method is based on the separation of phthalate esters from the sample with n-hexane, saponification of the esters with an alkaline ethanolic solution to give phthalic acid, purification of the acid by extraction with diethyl ether and column chromatography using silica gel, and conversion of the acid into bis(2,2,2-trifluoroethyl) phthalate with a 2,2,2-trifluoroethanol solution containing boron trifluoride. The derivative obtained is highly sensitive to an electron-capture detector, giving a sensitivity of 0.1 pg. Biological samples fortified with di(2-ethylhexyl) phthalate at levels of 5-100 ppb were analyzed, with recoveries of 70-100%.     

Determination of dissolved bromate in drinking water by ion chromatography and post column reaction: interlaboratory study

A collaborative study, International Evaluation Measurement Programme-25a, was conducted in accordance with international protocols to determine the performance characteristics of an analytical method for the determination of dissolved bromate in drinking water. The method should fulfill the analytical requirements of Council Directive 98/83/EC (referred to in this work as the Drinking Water Directive; DWD). The new draft standard method under investigation is based on ion chromatography followed by post-column reaction and UV detection. The collaborating laboratories used the Draft International Organization for Standardization (ISO)/Draft International Standard (DIS) 11206 document. The existing standard method (ISO 15061:2001) is based on ion chromatography using suppressed conductivity detection, in which a preconcentration step may be required for the determination of bromate concentrations as low as 3 to 5 microg/L. The new method includes a dilution step that reduces the matrix effects, thus allowing the determination of bromate concentrations down to 0.5 microg/L. Furthermore, the method aims to minimize any potential interference of chlorite ions. The collaborative study investigated different types of drinking water, such as soft, hard, and mineral water. Other types of water, such as raw water (untreated), swimming pool water, a blank (named river water), and a bromate standard solution, were included as test samples. All test matrixes except the swimming pool water were spiked with high-purity potassium bromate to obtain bromate concentrations ranging from 1.67 to 10.0 microg/L. Swimming pool water was not spiked, as this water was incurred with bromate. Test samples were dispatched to 17 laboratories from nine different countries. Sixteen participants reported results. The repeatability RSD (RSD(r)) ranged from 1.2 to 4.1%, while the reproducibility RSD (RSDR) ranged from 2.3 to 5.9%. These precision characteristics compare favorably with those of ISO 15601. A thorough comparison of the performance characteristics is presented in this report. All method performance characteristics obtained in the frame of this collaborative study indicate that the draft ISO/DIS 11206 standard method meets the requirements set down by the DWD. It can, therefore, be considered to fit its intended analytical purpose.     

Determination of glucosamine in raw materials and dietary supplements containing glucosamine sulfate and/or glucosamine hydrochloride by high-performance liquid chromatography with FMOC-Su derivatization: collaborative study

A collaborative study was conducted for determination of glucosamine in raw materials and dietary supplements containing glucosamine sulfate and/or glucosamine hydrochloride by high-performance liquid chromatography (HPLC) with N-(9-fluorenyl-methoxycarbonyloxy) succinimide (FMOC-Su) derivatization. Thirteen blind materials, one pair of which were duplicates, were tested by 12 collaborating laboratories. The test samples consisted of various commercial products, including tablets, capsules, drink mix, and liquids as well as raw materials, blanks, and those for spike recovery analyses. The tests with blank products and products spiked with glucosamine showed good specificity of the method. The average recoveries at spike levels of 100 and 150% of the declared amount were 99.0% with a relative standard deviation (RSD) of 2.1%, and 101% with an RSD of 2.3%, respectively. The test results between laboratories on each commercial product were reproducible with RSD values of no more than 4.0%, and the results were repeatable in the same laboratory with an average RSD of 0.7%. HorRat values ranged from 0.5 to 1.7 on both tests of spike recovery and reproducibility between laboratories on commercial products. The average determination coefficient of the calibration curves from the laboratories was 0.9995 with an RSD of 0.03%. All of the 12 collaborating laboratories succeeded in the study and none of their reported test results were outliers, partly indicating the robustness of the method. It is recommended that the method be accepted by AOAC INTERNATIONAL as Official First Action.     

Separation of fatty acids or methyl esters including positional and geometric isomers by alumina argentation thin-layer chromatography

This paper describes novel and rapid thin-layer chromatography procedures for the analysis of fatty acids and methyl esters using silver-impregnated alumina sheets. These techniques are known in most laboratories, and the equipment is readily available. The fatty acid method allows a separation of petroselinic (C18:1 delta 6c), oleic (C18:1 delta 9c), elaidic (C18:1 delta 9t), erucic (C22:1 delta 13c), and brassidic acids (C22:1 delta 13t), and the methyl ester method gives an excellent resolution with respect to the number, configuration, and position of the unsaturated centers. Sufficient separation for the subsequent ozonolysis and chromatographic quantification of isomeric C18 and C22 fatty acid methyl esters is obtained with both methods.     

In Process Citation

Ethylene oxide is extracted by circulation with distilled water in a closed circuit at 37 degrees . The circulation time, volume and flow of the water are previously determined as a function of the apparatus characteristics. The extract is analysed either by gas-liquid chromatography (direct injection with propylene oxide as internal standard) or by spectrophotometry. The latter method consists in the periodate oxidation of ethylene glycol in two samples of the same extract, with and without acid hydrolysis, followed by determination of formaldehyde with chromotropic acid. The difference between the absorbances of the two samples gives the quantity of ethylene oxide in the extract. A reasonable correlation of the two sets of results is observed, and the choice between these two simple methods depends on the number of samples to be analysed and on the equipment available. The spectrophotometric method is the longer of the two.     

A collaborative study to investigate the retention reproducibility of barbiturates in HPLC with a view to establishing retention databases for drug identification

A collaborative study among 10 laboratories has been undertaken to investigate the interlaboratory reproducibility of retention measurements in a high performance liquid chromatographic (HPLC) system for separating barbiturates. The system involved an ODS-Hypersil column with an eluent of methanol:phosphate buffer (40:60, v/v) at pH 8.5; all laboratories used the same batch of packing material. The conventional methods of recording retention properties (retention times and capacity factors) gave poor interlaboratory reproducibility. Better results were obtained by expressing the retentions relative to a standard barbiturate (quinalbarbitone); relative adjusted retention times proved to be more effective than straightforward relative retention times. Retention indices based on the alkylarylketone scale were not as reproducible as the methods based on a single closely related compound. The best reproducibility was obtained using corrected capacity factors based on the retention of four barbiturates in a standard mixture. The results of the study are discussed with a view to selecting the best methods of recording retention in HPLC when considering the establishment of databases for drug identification.     

Determination of domoic acid toxins in shellfish by biosense ASP ELISA--a direct competitive enzyme-linked immunosorbent assay: collaborative study

A collaborative study was conducted on the Biosense amnesic shellfish poisoning (ASP) enzyme-linked immunosorbent assay (ELISA) for the determination of domoic acid (DA) toxins in shellfish in order to obtain interlaboratory validation data for the method. In addition, a method comparison study was performed to evaluate the ASP ELISA as an alternative to the current liquid chromatography (LC) reference method for DA determination. The study material comprised 16 shellfish samples, including blue mussels, Pacific oysters, and king scallops, spiked with contaminated mussel homogenates to contain 0.1-20 mg DA/kg shellfish flesh. The shellfish samples were extracted with 50% aqueous methanol, and the supernatants were directly analyzed. Sixteen participating laboratories in 10 countries reported data from the ASP ELISA, and 4 of these laboratories also reported data from instrumental LC analysis. The participating laboratories achieved interlaboratory precision estimates for the 8 Youden paired shellfish samples in the range of 10-20% for RSD(r) (mean 14.8 +/- 4%), and 13-29% for RSDR (mean 22.7 +/- 6%). The precision estimates for the ELISA data did not show a strong dependence on the DA concentration in the study samples, and the overall precision achieved was within the acceptable range of the Horwitz guideline with HorRat values ranging from 1.1 to 2.4 (mean HorRat 1.7 +/- 0.5). The analysis of shellfish samples spiked with certified reference material (CRM)-ASP-MUS-b gave recoveries in the range of 88-122%, with an average recovery of 104 +/- 10%. The estimate on method accuracy was supported by a correlation slope of 1.015 (R2 = 0.992) for the determined versus the expected DA values. Furthermore, the correlation of the ASP ELISA results with those for the instrumental LC analyses of the same sample extracts gave a correlation slope of 1.29 (R2 = 0.984). This indicates some overestimation of DA levels in shellfish by the ELISA, but it is also a result of apparent low recoveries for the LC methods. This interlaboratory study demonstrates that the ASP ELISA is suitable for the routine determination and monitoring of DA toxins in shellfish, and that it offers a rapid and cost-effective methodology with high sample throughput.     

Multiresidue gas chromatographic method for determining synthetic pyrethroid pesticides in agricultural products: collaborative study

Fourteen laboratories from 6 countries and regions participated in an international collaborative study to evaluate a multiresidue gas chromatographic (GC) method for determining 8 synthetic pyrethroid pesticides in grains, fruits, and vegetables. The study design was based on Youden's matched-pairs principle for collaborative tests of analytical methods. Each laboratory analyzed 12 collaborative samples of wheat, oranges, and tomatoes as blind samples. Wheat samples were extracted with acetonitrile-water (2 + 1), while orange and tomato samples were extracted with acetone. Residues were partitioned into hexane, evaporated to dryness with a rotary evaporator, and then dissolved in hexane. The hexane extract was partitioned with acetonitrile and cleaned up on a 5% water-deactivated Florisil column with 6% ethyl ether in hexane as eluant. Residue concentrations were determined by GC with electron capture detection with splitless injection by comparison with single-point calibration standards. The appropriate standard concentration was determined by screening sample extracts before analysis. The multiresidue method was tested over the concentration range of 0.095-1.909 mg/kg depending on the 8 different of pesticides and agricultural products analyzed in the collaborative study. Statistical analysis of data from 13 laboratories showed weighted average recoveries for 8 pyrethroids in wheat, oranges, and tomatoes at 0.105-1.909, 0.095-1.909, and 0.105-0.954 mg/kg, respectively, ranging from 91.8 to 100.2%, from 88.1 to 100.6%, and from 88.2 to 101.5%, respectively. Reproducibility relative standard deviation values ranged from 6.46 to 17.74%, from 5.94 to 18.13%, and from 5.59 to 10.48%, respectively. Repeatability relative standard deviation values ranged from 6.34 to 10.84%, from 5.19 to 11.72%, and from 3.20 to 8.09%, respectively. The multiresidue GC method for determining synthetic pyrethroid pesticides in agricultural products has been adopted first action by AOAC INTERNATIONAL.     

Gliadin as a measure of gluten in foods containing wheat, rye, and barley-enzyme immunoassay method based on a specific monoclonal antibody to the potentially celiac toxic amino acid prolamin sequences: collaborative study

The Working Group on Prolamin Analysis and Toxicity (WGPAT) organized a collaborative study to confirm whether the two R5 antibody-based ELISA test kits are able to detect gliadin in the lower mg/kg (ppm) level. Twenty laboratories investigated 12 blind-coded samples, spiked and naturally contaminated, to show the possibility of determining traces of gliadin in heat-treated or nonheat-treated foods by ELISA. It was shown that very small amounts of gliadin (below 100 ppm) could be detected by ELISA with a reproducibility RSD(R) (37%) and a repeatability RSD, (27%) common for ELISA under these conditions. The recovery of gliadin from the spiked samples was between 84 and 109%, based on the results of all laboratories, including those with poor performance. No false positives were found by the method (P < or =0.05), but one negative sample was contaminated during the bakery process. It is recommended that the method be accepted by AOAC as Official First Action.     

Interlaboratory study of analysis of phenoxymethylpenicillin by liquid chromatography

Summary A liquid chromatography method for analysis of phenoxymethylpenicillin has been examined in a collaborative study involving 6 laboratories. The method comprised an isocratic part, which is used in the assay. When the isocratic part is combined with gradient elution, the method is suitable for purity control. Five samples of phenoxymethylpenicillin (potassium salts and acid) were analysed. The main component, the most important side product 4-hydroxyphenoxymethyl-penicillin and other impurities were determined. An analysis of variance proved the absence of consistent laboratory bias. Laboratory-sample interaction was not significant. Estimates of the repeatability and reproducibility of the method, expressed as standard deviations (SD) of the result of the determination of phenoxymethylpenicillin, were 0.50 and 0.63 respectively.     

Determination by glyphosate and aminomethylphosphonic acid in crops by capillary gas chromatography with mass-selective detection: collaborative study.

A collaborative study was conducted to validate a method for the determination of glyphosate and aminomethylphosphonic acid (AMPA) in crops. The analytes are extracted from crops with water, and the crude extracts are then subjected to a cation exchange cleanup. The analytes are derivatized by the direct addition of the aqueous extract into a mixture of heptafluorobutanol and trifluoroacetic anhydride. The derivatized analytes are quantitated by capillary gas chromatography with mass-selective detection (MSD). The collaborative study involved 13 laboratories located in 5 countries; 12 laboratories returned valid data sets. The crops tested were field corn grain, soya forage, and walnut nutmeat at concentrations of 0.050, 0.40, and 2.0 mg/kg. The study used a split-level pair replication scheme with blindly coded laboratory samples. Twelve materials were analyzed, including 1 control and 3 split-level pairs for each matrix, 1 pair at each nominal concentration. For glyphosate, the mean recovery was 91%, the average intralaboratory variance, the repeatability relative standard deviation (RSDr), was 11%, and the interlaboratory variance, the reproducibility relative standard deviation (RSDR), was 16%. For AMPA, the mean recovery was 87%, the RSDr was 16%, and the RSDR was 25% at mg/kg levels.     

Determination of fluoride in wine by fluoride selective ion electrode, standard addition method: collaborative study

The accuracy, precision, and reproducibility of a rapid method for determination of fluoride in wine, using a fluoride selective ion electrode, were established by a collaborative study involving 12 laboratories, 5 in Europe and 7 in the United States. The laboratories assayed 6 Youden pairs of fluoride-fortified, red and white wine samples with fluoride concentrations ranging from 0.2 to 3.0 mg/L. The relative standard deviations of repeatability ranged from 1.94 to 4.88%; relative standard deviations of reproducibility ranged from 4.15 to 18.40%. HORRAT values ranged from 0.30 to 0.97. The average recovery was 99.97%. Based on the statistical results of this collaborative study, the Study Director recommends that this method be adopted First Action.     

Results of collaborative trials concerning the analysis of benzo-a-pyrene in creosote

 The results of five collaborative studies concerning the analysis of benzo-a-pyrene (BaP) in creosote by high-performance liquid chromatography (HPLC) are presented. Six to twelve laboratories participated in these studies over a time period of three years. These studies were carried out in order to evaluate the method which is described in the CEN standard ENV 1014-3. The concentrations of BaP in creosote samples analysed were in the range of 20 ppm to 550 ppm. Whereas the values for repeatability (5–10%) were acceptable in nearly all collaborative trials, those for reproducibility have improved significantly in the last study (23–33%). For the previous four collaborative trials the reasons for the relatively poor reproducibility values, especially for analytes containing low concentrations of BaP, could be detected. They were mainly based on problems with chromatographic separation and with integration. The method described in ENV 1014-3 has been proven to be suitable for the determination of the BaP content in creosote. Received: 21 June 1996/Revised: 21 August 1996/Accepted: 25 August 1996     

Results of collaborative trials concerning the analysis of benzo-a-pyrene in creosote

 The results of five collaborative studies concerning the analysis of benzo-a-pyrene (BaP) in creosote by high-performance liquid chromatography (HPLC) are presented. Six to twelve laboratories participated in these studies over a time period of three years. These studies were carried out in order to evaluate the method which is described in the CEN standard ENV 1014-3. The concentrations of BaP in creosote samples analysed were in the range of 20 ppm to 550 ppm. Whereas the values for repeatability (5–10%) were acceptable in nearly all collaborative trials, those for reproducibility have improved significantly in the last study (23–33%). For the previous four collaborative trials the reasons for the relatively poor reproducibility values, especially for analytes containing low concentrations of BaP, could be detected. They were mainly based on problems with chromatographic separation and with integration. The method described in ENV 1014-3 has been proven to be suitable for the determination of the BaP content in creosote. Received: 21 June 1996/Revised: 21 August 1996/Accepted: 25 August 1996     

Sulfonate group-modified FePtCu nanoparticles as a selective probe for LDI-MS analysis of oligopeptides from a peptide mixture and human serum proteins

The availability of robust methods for the species-specific detection of meat and bone meal (MBM) in compound feedingstuffs is an important prerequisite to enforce current and upcoming European legislation on the use of processed animal proteins in animal nutrition. Among possible methods, those based on DNA turned out to be a reliable tool for this aim, since DNA is a quite thermostable molecule able to resist severe heat treatments applied in the manufacturing of animal meals. The application of such methods by control laboratories implies that the method has been validated including an assessment of its robustness. Successful transferability between laboratories is considered an important robustness criterion of the method. However, corresponding guidelines regarding the design of such a study relevant to this field are missing. Here, we demonstrate the feasibility of an alternative concept that was applied to check for the transferability of a qualitative assay for the detection of banned MBM in feedingstuffs at trace level based on real-time PCR. The concept was based on an experimental nested design applying analysis of variance (ANOVA) that was conducted independently in two laboratories and which allows for establishing major factors influencing the result of analysis. Statistical assessment of the results confirmed the importance of the DNA extraction/purification step utilised, whereas the PCR step turned out to be a minor factor regarding the overall variability of the results. Furthermore, blind samples comprised of compound feed adulterated with MBM at 0.1 % and blank compound feed were correctly classified as "positive" or "negative" samples, thus confirming fitness of purpose for the method. This approach can be of interest for other research groups working in the development of real-time PCR methods and in their use by control laboratories. Nested design of the study to check for the transferability of the real-time PCR method     

Analysis of the anti-coccidial drug, halofunginone, in chicken feed using gas-liquid chromatography and high-performance liquid chromatography

Methods are described for the analysis of the anti-coccidial drug, halofuginone, at concentrations of 3 ppm in chicken feed, using gas-liquid chromatography and high-performance liquid chromatography. Both methods are based on ethyl acetate extraction, partition into hydrochloric acid and purification and concentration using XAD-2 column chromatography. The precision and accuracy of both methods is given.     

A collaborative study for intermethod comparison

A collaborative study for the determination of vinclozolin and captan in lettuce and pears was conducted in the author's laboratory. In two 3-day courses the participants of twelve laboratories analysed the samples by the micro on-line method and two macro methods for intermethod comparison. The pesticides were determined by GC-ECD after clean-up on silica gel deactivated by 10% water. The results of the micro method are well comparable with those of the macro methods. Therefore, the macro extraction methods can be substituted by the micro on-line method.