Determination of Novobiocin Residues in Milk, Blood, and Tissues by Liquid Chromatography

A rapid, specific, and sensitive method was developed for determination of tetracyclines (TCs) in egg, milk, and milk powder samples by online molecularly imprinted solid-phase extraction (MISPE) coupled to a reversed-phase high-performance liquid chromatography (HPLC). A molecularly imprinted polymer hybrid composite material (MIP-HCM) was prepared using TC as the template, methacrylic acid as the functional monomer, tetraethoxysilane as inorganic precursor, and methacryloxypropyltrimethoxysilane as the coupling agent. A new technique of online solid-phase extraction (SPE) coupled with HPLC was established. The conditions of online SPE were optimized. Under the optimal conditions, three TCs residues in egg, milk, and milk powder were separated and detected by the proposed method with better clean-up and recoveries. The recoveries of TCs ranged from 85.3% to 98.3%. The limits of detection and quantitation of the proposed method were in the range of 0.76–1.13 µg/kg and 2.53–3.77 µg/kg, respectively. The MIP-HCM was first used as online SPE packing and the hyphenated technique is promising as a general strategy for direct determination of veterinary drug residues in milk and other biological samples.

Seven laboratories participated in a collaborative study of a liquid chromatographic (LC) method for determination of sulfamethazine (SMZ) residues in raw milk that were previously frozen. The milk is extracted with chloroform, the chloroform is evaporated, and the residue is suspended in hexane and extracted with 0.1M KH2PO4 (PDP) solution. The PDP extract is analyzed by LC on a C18 column with methanol-0. 1M PDP (30 + 70) as mobile phase. Individual laboratories were instructed to analyze 5 replicates each of control milk, fortified control milk at 2 levels, and 3 blind samples. Blind samples included raw milk fortified with SMZ at 10 and 20 ppb and 1 sample containing SMZ residue from a dosed cow. For blind fortified samples containing 10 ppb SMZ, average recovery and relative standard deviations for repeatability and reproducibility (RSDr and RSRR) based on the results from 6 of the 7 participating laboratories were 8.21 ppb, 7.16%, and 23.16%, respectively. Similar data, including results from a seventh participant who reported instrumental problems but was not eliminated by the Dixon outlier test, were 9.13 ppb, 8.38%, and 31.94%, respectively. These results demonstrate that the method is suitable for the determination of SMZ residues in milk at 10 ppb and above. The method was adopted first action by AOAC International.

Simultaneous analysis of six antiepileptic drugs and two selected metabolites in human plasma by liquid chromatography after solid-phase extraction

ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTInsecticide Residues, A Procedure for the Microdetermination of 1-Butoxy-2-(2-thiocyanoethoxy)ethane (Lethane 384) with Applications for Determination of Residues in Milk and Animal TissuesC. F. Gordon, L. D. Haines, and A. L. WolfeCite this: J. Agric. Food Chem. 1961, 9, 6, 478–481Publication Date (Print):November 1, 1961Publication History Published online1 May 2002Published inissue 1 November 1961https://doi.org/10.1021/jf60118a018RIGHTS & PERMISSIONSArticle Views19Altmetric-Citations5LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit PDF (453 KB) Get e-Alerts Get e-Alerts

Although human plasma represents an attractive sample for disease biomarker discovery, the extreme complexity and large dynamic range in protein concentrations present significant challenges for characterization, candidate biomarker discovery, and validation. Herein we describe a strategy that combines immunoaffinity subtraction and subsequent chemical fractionation based on cysteinyl peptide and N-glycopeptide captures with two-dimensional LC-MS/MS to increase the dynamic range of analysis for plasma. Application of this "divide-and-conquer" strategy to trauma patient plasma significantly improved the overall dynamic range of detection and resulted in confident identification of 22,267 unique peptides from four different peptide populations (cysteinyl peptides, non-cysteinyl peptides, N-glycopeptides, and non-glycopeptides) that covered 3,654 different proteins with 1,494 proteins identified by multiple peptides. Numerous low abundance proteins were identified, exemplified by 78 "classic" cytokines and cytokine receptors and by 136 human cell differentiation molecules. Additionally a total of 2,910 different N-glycopeptides that correspond to 662 N-glycoproteins and 1,553 N-glycosylation sites were identified. A panel of the proteins identified in this study is known to be involved in inflammation and immune responses. This study established an extensive reference protein database for trauma patients that provides a foundation for future high throughput quantitative plasma proteomic studies designed to elucidate the mechanisms that underlie systemic inflammatory responses.

To meet federal and state regulatory needs, a liquid chromatographic (LC) method with ultraviolet (UV) detection was developed for determination of 7 tetracyclines at 30 ng/ml in milk. Raw milk samples are defatted, acidified, and centrifuged to remove proteins, and tetracyclines are specifically absorbed from the milk by chelation with metal ions bound to small Chelating Sepharose Fast Flow columns. Tetracyclines are removed from these columns with EDTA-containing buffer, and extracts are further cleaned by ultrafiltration. Finally, extracts are concentrated and analyzed simultaneously by using on-line concentration. This method was validated in a collaborative study that involved 11 laboratories, including the authors' laboratory. Each laboratory was asked to prepare and analyze known control and fortified milk samples, as well as 18 coded blind samples. Eight laboratories completed all analyses. Average interlaboratory recoveries for the known fortified samples ranged from 59% (methacycline at 15 ng/ml) to 78% (oxytetracycline at 60 ng/ml). Average recovery for each of 7 residues at 30 ng/ml were between 60 and 110%, meeting single-residue guidelines for accuracy set by the U.S. Food and Drug Administration. Reproducibility relative standard deviation (RSDR) for the known fortified samples varied from 11 to 39%, with 6 of 7 residues at the 30 ng/ml level having RSDR values at or below 20%. Seven of 8 laboratories correctly identified blind control milk samples and all 28 residues present in blind samples. The metal chelate affinity-LC method for determination of multiple tetracycline residues in milk has been adopted first action by AOAC INTERNATIONAL.

A new liquid chromatography/mass spectrometry (LC/MS) method is presented for the determination of chloramphenicol (CAP) residues in milk, eggs, chicken muscle and liver, and beef muscle and kidney. CAP is extracted from the samples with acetonitrile and defatted with hexane. The acetonitrile extracts are then evaporated, and residues are reconstituted in 10mM ammonium acetate--acetonitrile mobile phase and injected into the LC system. CAP is determined by reversed-phase chromatography using an Inertsil ODS-2 column and MS detection with negative ion electrospray ionization. Calibration curves were linear between 0.5-5.0 ng/g for all matrixes studied. The relative standard deviations for measurements by this method were generally <12%, and average recoveries ranged from 80 to 120%, depending on the matrix involved. The method detection limits of CAP ranged from 0.2 to 0.6 ng/g, which are comparable to previously reported results. The proposed method is rapid, simple, and specific, allowing a single analyst to easily prepare over 40 samples in a regular working day.

Determination of gamithromycin residues in eggs, milk and edible tissue of food-producing animals by solid phase extraction combined with ultrahigh-performance liquid chromatography-tandem mass spectrometry

Veterinary treatments, mainly antibiotics, used for therapeutic or prophylactic purposes in dairy farming can be the cause of the presence of antibiotic residues in milk. However, these residues constitute a major health concern for consumers. For the dairy industry, whose objective is to have a raw material suitable for processing, it is necessary to screen for antibiotic residues in milk at each collection. This study is based on two parts, a survey of seventy people (10 practicing veterinarians and 60 milk producers) to describe the main molecules of antibiotics used in dairy cattle farming in the two basins of Tienfala and Kasséla in the Koulikoro region, and a search for residues by Delvotest SP with confirmation of positive samples by liquid chromatography (HPLC). To this end, sixty (60) samples of raw milk were collected in two sampling campaigns per site carried out 15 days apart. All samples were analyzed using the Delvotest SP rapid detection of antibiotic residues in milk kit first before being analyzed by liquid chromatography (HPLC) for positive samples. The results of the surveys revealed the predominance of three pathologies within dairy herds: diarrhoea, mastitis and pulmonary infections with a combination of several antibiotics belonging to five (5) different families of antibiotics. Of the 60 samples submitted for analysis, 38 were found to be positive with Delvotest SP, i.e. a contamination rate of around 63.33%. Of the 38 samples positive to the rapid test, 26 responded positively to confirmation by liquid chromatography, i.e. a rate of 68.42% with mainly four antibiotics belonging to the families of β-lactams and tetracyclines. The results of the study effectively prove the effective presence of antibiotic residues in raw cow's milk from the mini-dairies of Kassela and Tienfala. They thus characterize current practices in the treatment of dairy cattle with antibiotics in the farming areas studied. Compliance with withdrawal periods with the elimination of milk from treated cows must be obtained by raising the awareness of producers by animal health professionals.

Most studies to determine sulfonamide residues in milk samples have used solid-phase extraction as the sample preparation technique. However, the quick, easy, cheap, effective, rugged, and safe (QuEChERS) method, introduced in 2003, has been used in the extraction of various compounds in food matrices. This study aimed to evaluate two sample preparation techniques: solid-phase extraction and QuEChERS, for chromatographic analysis of sulfonamides (sulfathiazole, sulfamethazine, and sulfadimethoxine) in bovine milk. The chromatographic parameters and the QuEChERS extraction procedure were developed by using different experimental designs, obtaining good peak resolution, recovery, precision, accuracy, linearity, selectivity, and limits of detection and quantification. In contrast, using solid-phase extraction, acceptable recoveries and selectivity were not achieved, despite the number of articles published that have applied this sample preparation technique for sulfonamide analysis. As a result of the experiments performed, probably sulfonamides are retained together with other components of the matrix in the sample pretreatment step (prior to its addition in the cartridge containing solid phase), which is an important part of solid-phase extraction with raw whole milk. Therefore, QuEChERS is a better method than solid-phase extraction for the analysis of sulfonamide residues in milk. Validation tests demonstrated that the method is appropriate, within the maximum residue limit (0.1 mg kg−1). Moreover, it was possible to use a lower amount of solvent compared with previously published articles (6 mL against 10 or 15 mL).

Vancomycin and norvancomycin, as potent antibacterial retention drugs, were used illegally on animals bred for food, which directly affected the quality and safety of animal-derived food, and even harmed human health. A fast analysis method, which was adopted to detect residues of vancomycin and norvancomycin in milk, was implemented on a chromatographic system containing online solid-phase extraction (SPE) device that combined with high-resolution mass spectrometer (HRMS). First, the analytes were added to the blank milk sample were extracted with water [containing 0.1% trifluoroacetic acid (TFA)]-acetonitrile (ACN) (8:2, v/v), and then were purified and enriched on a C18-XL column, whereafter eluted from the purification column onto the analytical column (Shiseido Capcell Pak ADME column) for chromatographic separation prior to hybrid quadrupole-Orbitrap (Q-Orbitrap) detection. The results showed that the limit of detection (LOD) for each analyte and the limit of quantitation (LOQ) were 0.15 and 0.5 μg/kg, respectively. The correlation coefficient(s) of vancomycin and norvancomycin ranged from 0 to 200 ng/mL were greater than 0.9983. These validations reflected that it was suitable for the established method to rapidly analyze vancomycin and norvancomycin residues in milk. The method for detecting vancomycin and norvancomycin residues in milk by online SPE combined with LC-HRMS. Online SPE technology realized automation, and the application of HRMS greatly improved the reliability of qualitative and quantitative analyses. The developed method is fast, simple, and reliable; each methodological index can meet requirements of trace analyses of vancomycin and norvancomycin in milk.

Tylosin, an antibiotic developed specifically for agricultural use, and erythromycin are the main macrolide antibiotics used in animal production. Two-dimensional thin layer chromatography has been used for detection of tylosin in poultry meat, eggs, and milk and for erythromycin in poultry meat. Detection limits reported are, for tylosin, 0.1 ppm in poultry meat, 0.05 ppm in egg, and 0.01 ppm in milk, and for erythromycin, 0.25 ppm in poultry meat. Liquid chromatography (LC) has also been used for determination of tylosin in milk, blood, and tissues of animals. Samples (milk, blood serum, or tissue homogenates in water or pH 2.2 buffer) were deproteinized with acetonitrile, tylosin was partitioned into methylene chloride, and the extracts were concentrated and dissolved in acetonitrile. Chromatography was done on a reverse phase end-capped C18 column using 0.002-0.005 M ammonium dihydrogen phosphate-acetonitrile-methanol (10 + 60 + 30-5 + 80 + 15). Solvent composition was varied with the type of sample analyzed. The method will detect 0.1 ppm tylosin in tissues and less in milk and blood serum. The LC method was more sensitive than microbiological assays for detection of tylosin in tissues of treated swine; recoveries of tylosin by the LC method were frequently several-fold higher.

A simple, specific, and sufficiently sensitive liquid chromatography-tandem mass spectrometry (negative-ion electrospray ionization) methodology to determine mevalonic acid (MVA) in human plasma is described, and its application to the analysis of rat plasma MVA levels after rosuvastatin administration is demonstrated. The method was validated over the linearity range of 0.5-50.0 ng/ml (r(2) > 0.99) using deuterated MVA as an internal standard. The lower limit of quantification was 0.5 ng/ml. The assay procedure involved the isolation of MVA from plasma samples using solid-phase extraction. Chromatographic separation was achieved on a HyPurity Advance column with a mobile phase consisting of ammonium formate buffer (10 mM, pH 8.0) and acetonitrile (70:30, v/v). Excellent precision and accuracy were observed. MVA and deuterated mevalonolactone were stable in water and plasma under different storage and processing conditions. The recovery observed was low, which was attributable to a significant matrix effect. A significant decrease (30-40%; P < 0.05) was observed in rat plasma MVA levels after rosuvastatin administration.

A simple liquid chromatographic (LC) method was developed for the determination of fluoroquinolones (ciprofloxacin, difloxacin, enrofloxacin and sarafloxacin) in animal tissues. Isolation of fluoroquinolones from biological matrices was performed with 5% trichloroacetic acid-acetonitrile (7:3) solution. For clean-up, solid-phase extraction with an SDBI (styrene-divinylobenzene) cartridge was used. LC analyses were performed with analytical column (LiChrospher 100 RP-8 5 microns) and mobile phase (0.025 M o-phosphoric acid-acetonitrile 70:30, v/v) in ion-pair mode. The whole procedure was validated in intra- and inter-assay reproducibility and accuracy determination by simultaneously assaying of muscle, liver and kidney samples supplemented with fluoroquinolones at the level of 30 and 60 ng/g, respectively. The statistical evaluation demonstrates high absolute recovery (> 80%) and low coefficient of variation (< 10%) for all analysed samples. The detection limits for fluoroquinolones were 5 ng/g in muscle, liver and kidney samples.

A simple liquid chromatographic (LC) method was developed for the determination of fluoroquinolones (ciprofloxacin, difloxacin, enrofloxacin and sarafloxacin) in animal tissues. Isolation of fluoroquinolones from biological matrices was performed with 5% trichloroacetic acid-acetonitrile (7:3) solution. For clean-up, solid-phase extraction with an SDBI (styrene–divinylobenzene) cartridge was used. LC analyses were performed with analytical column (LiChrospher 100 RP-8 5 µm) and mobile phase (0.025 M O-phosphoric acid–acetonitrile 70:30, v/v) in ion-pair mode. The whole procedure was validated in intra- and inter-assay reproducibility and accuracy determination by simultaneously assaying of muscle, liver and kidney samples supplemented with fluoroquinolones at the level of 30 and 60 ng/g, respectively. The statistical evaluation demonstrates high absolute recovery (>80%) and low coefficient of variation (<10%) for all analysed samples. The detection limits for fluoroquinolones were 5 ng/g in muscle, liver and kidney samples. Copyright © 1999 John Wiley & Sons, Ltd.