Bacillus stearothermophilus Disk Assay for Detection of Residual Penicillins in Milk: Collaborative Study

1979 ◽  
Vol 62 (5) ◽  
pp. 985-988
Author(s):  
Larry A Ouderkirk
Keyword(s):  
Bacillus Stearothermophilus ◽  
Micrococcus Luteus ◽  
Collaborative Study ◽  
Penicillin G ◽  
Plate Method ◽  
Antibiotic Drugs ◽  
Paper Disk ◽  
Whole Milk ◽  
Low Levels ◽  
Disk Method

Abstract A collaborative study was performed on a Bacillus stearothermophilus paper disk method designed to detect residual levels of 4 antibiotic drugs in whole market milk. This method is a modification of an earlier procedure developed for the International Dairy Federation. Whole milk samples spiked at low levels with ampicillin, cephapirin, cloxacillin, and penicillin G were sent frozen to 11 collaborating laboratories with instructions to assay them promptly according to the method provided. Five of the laboratories reported inconclusive results due to technical difficulties encountered with the method. The 6 remaining laboratories all detected levels of 0.005- 0.008 μg or unit/mL for penicillin G, ampicillin, and cephapirin and 0.05-0.08 μg/mL for cloxacillin. The most commonly used official methods, the Sarcina lutea (Micrococcus luteus) cylinder plate method and the Bacillus subtilis paper disk method, can detect levels of 0.01 and 0.05 unit penicillin G/mL, respectively. The B. stearothermophilus method is rapid, simple to perform, and more sensitive than present official methods. The method has been adopted as official first action for the detection of penicillins in milk.

Detection of Residual Penicillins in Milk by Using a Bacillus stearothermophilus Disk Assay

1977 ◽  
Vol 60 (5) ◽  
pp. 1116-1118
Author(s):  
Larry A Ouderkirk
Keyword(s):  
Bacillus Stearothermophilus ◽  
Test Organism ◽  
Spore Suspension ◽  
Penicillin G ◽  
Paper Disk ◽  
Whole Milk ◽  
Disk Method ◽  
International Dairy Federation

Abstract A paper disk method based on a procedure described by the International Dairy Federation is presented for the detection of 4 β-lactam antibiotics (penicillin G, ampicillin, cephapirin, and cloxacillin) in whole milk. Bacillus stearothermophilus var. calidolactis, prepared as a stable spore suspension, was used as the test organism. Levels of 0.005 unit penicillin G/ml, 0.005 μg ampicillin or cephapirin/ml, and 0.05 μg cloxacillin/ml were readily detected in whole milk. Results were produced in 3–4 hr. The method offers several advantages, including greater simplicity, sensitivity, and rapidity, over methods now commonly used to detect residual levels of these drugs in milk.

Qualitative Ampule and Multitest for Beta-Lactam Residues in Fluid Milk Products: Collaborative Study

1982 ◽  
Vol 65 (5) ◽  
pp. 1193-1207 ◽  
Author(s):  
Wesley N Kelley ◽  
◽  
J Andrews ◽  
A W Appelt ◽  
R Barber ◽  
...  
Keyword(s):  
Bacillus Stearothermophilus ◽  
Collaborative Study ◽  
Original Method ◽  
Penicillin G ◽  
Beta Lactam ◽  
Milk Products ◽  
Low Levels ◽  
Positive Results ◽  
Made In ◽  
Fluid Milk

Abstract A collaborative study was performed on a rapid Bacillus stearothermophilus agar diffusion ampule method to detect low levels of penicillin G in 7 types of fluid milk products. A multitest technique for processing a large number of samples simultaneously was also studied. Slight modifications were made in the original method to establish more uniformity and to eliminate doubtful responses by specifying a confirmation procedure. Twenty samples spiked with penicillin G (0.000 to 0.008 IU/mL) and tetracycline hydrochloride were frozen and sent to 20 laboratories in the ampule test, and 16 laboratories in the multitest. Each analyst was asked to do a screening run and a confirmation run. Results were reported by color reaction and also as positive or negative for β-lactam inhibitors. The concentrations (penicillin G) where percent positive results equal 100 or not significantly less than 100 (α = 0.05) ranged from 0.005 to 0.007 IU/mi in the ampule test and from 0.004 to 0.007 IU/mL in the multitest. Both techniques have been adopted official first action.

Evaluation of Two Microbiological Methods for Detecting Residual Antibiotics in Milk

1976 ◽  
Vol 59 (5) ◽  
pp. 1122-1124
Author(s):  
Larry A Ouderkirk
Keyword(s):  
Bacillus Subtilis ◽  
Dairy Industry ◽  
Test Organism ◽  
Plate Method ◽  
Plate Technique ◽  
Whole Milk ◽  
Microbiological Methods ◽  
Aoac Official ◽  
Disk Method ◽  
Official Methods Of Analysis

Abstract Two methods described in the AOAC Official Methods of Analysis for the detection of penicillin residues in whole milk were evaluated to determine the capability of each method to detect residues of 12 antibiotics used in the dairy industry. The first method, a cylinder-plate method that uses Sarcina lutea as the test organism, detected levels of 1 ppm of 8 of the 12 antibiotics tested. The second method, using paper disks with Bacillus subtilis as the test organism, detected approximately 1 ppm of only 4 antibiotics. This disk method was unable to detect <40 ppm of 5 of the antibiotics tested. The data indicate that the S. lutea cylinder-plate technique is more sensitive to more antibiotics than the B. subtilis disk method and is far superior for screening purposes.

Bacillus stearothermophilus Disc Assay for Detection of Inhibitors in Milk: Collaborative Study

1982 ◽  
Vol 65 (5) ◽  
pp. 1208-1214 ◽  
Author(s):  
James W Messer ◽  
James E Leslie ◽  
Gary A Houghtby ◽  
James T Peeler ◽  
Jerald E Barnett ◽  
...  
Keyword(s):  
Bacillus Stearothermophilus ◽  
Collaborative Study ◽  
Raw Milk ◽  
Penicillin G ◽  
Confirmatory Test ◽  
Beta Lactam ◽  
Aoac Method ◽  
Positive Results ◽  
Paper Disc ◽  
Disc Assay

Abstract A 2-part (A and B) collaborative study was conducted on a Bacillus stearothermophilus paper disc (12.7 mm) method to detect residual inhibitors in milk. The 18 participating collaborators assayed raw milk samples spiked with a beta-lactam (penicillin G). Of the 18 collaborators, 14 participated in part A and 16 in part B. Part A demonstrated that either Antibiotic Medium No. 4 or PM Indicator Agar is suitable for use in the assay. The lowest concentration detectable, not significantly different from 100% at the α = 0.05 level, was 0.008 unit/mL with either medium. Part B demonstrated that the sensitivity of the method is equal to that of the current AOAC method (16.131- 16.136). The concentration of beta-lactam detected by 50% of the analysts was 0.003-0.005 unit/mL in this study, compared with 0.005 unit/mL reported in an earlier collaborative study on the current AOAC method. No false positive results were reported in part A or part B. All samples found positive by the confirmatory test in part B were correctly identified as a beta-lactam with commercial Penase discs. The lowest concentration detectable by the method, not significantly different from 100% at the α = 0.05 level, was 0.008 unit/mL. The method was adopted official first action.

scholarly journals Extending Postharvest Longevity and Improving Quality of Strawberry (Fragaria Ananasa Duch Cv. ‘Gaviota’) Fruit by Postharvest Salicylic Acid Treatment

2015 ◽  
Vol 3 (2) ◽  
pp. 17 ◽  
Author(s):  
Mahsa Geransayeh ◽  
Sadegh Sepahvand ◽  
Vahid Abdossi
Keyword(s):  
Weight Loss ◽  
Salicylic Acid ◽  
Vitamin C ◽  
Horticultural Crops ◽  
Postharvest Application ◽  
Spray Method ◽  
Paper Disk ◽  
Fruit Decay ◽  
Activity Decay ◽  
Disk Method

Strawberries are an extremely perishable fruit mainly due to their soft texture and sensitivity to fungal infection. Postharvest application of conventional fungicides to fruits is prohibited. As an alternative to fungicides, salicylic acid has been found to enhance disease resistance of horticultural crops. In order to study the effect of salicylic acid as a phenolic compound on the postharvest durability and quality characteristics of strawberry fruit. ‘Gaviota’ strawberries were treated with SA at different concentrations (0, 25, 50 and 100 µlL-1), then stored for 12 days at 4 ˚C and 75 % RH in darkness. Two different methods were applied (spray SA on fruits and paper disk method). Quality attributes such as weight loss, pH, TA, TSS, vitamin C, anthocyanin, calcium, pectin, CAT, POD, PG activity, decay percentage and sensory analyses evaluated every 3 days during storage. Results showed that, treated fruits with SA had lower weight loss, pH, TSS, POD, PG, decay and higher TA, vitamin C, anthocyanin, calcium, pectin, CAT and fruit quality compared with controls. Between two methods of treatment, paper disk method had higher effect on fruit decay and quality compared to spray method and as a general result, caused longer storability.

Microbial Receptor Assay for Rapi d Detection and Identification of Seven Families of Antimicrobial Drugs in Milk: Collaborative Study

1988 ◽  
Vol 71 (2) ◽  
pp. 304-316 ◽  
Author(s):  
Stanley E Charm ◽  
Ruey Chi
Keyword(s):  
Binding Sites ◽  
Specific Binding ◽  
Scintillation Counter ◽  
Collaborative Study ◽  
False Negative ◽  
Penicillin G ◽  
Specific Binding Sites ◽  
Receptor Assay ◽  
Relative Standard ◽  
Standard Deviations

Abstract A microbial competitive receptor assay for detecting residues of antibiotic families in milk was studied collaboratively by 13 laboratories. The drugs and levels (ppb) tested in this study i nclude penicillin G, 4.8; cephapirin, 5.0; cloxacillin, 100; tetracycline, 2000; chlortetracycline, 2000; oxytetracycline, 2000; erythromycin, 200; lincomycin, 400; clindamycin, 400; sulfamethazine, 75; sulfamethoxazole, 50; sulfisoxazole, 50; streptomycin, 1000; novobiocin, 50; and chloramphenicol, 800. In this method, microbial cells added to a milk sample provide specific binding sites for which 14C or 3H libeled drug competes with drug residues in the sample. The UC or H binding to the specific binding sites is measured in a scintillation counter and compared with a zero standard milk. If the sample is statistically different from the zero standard, it is positive. The assay takes about 15 min. The binding reaction occurs between the receptor site and the drug functional group, so all members of a drug family are detected. In this case, beta-lactams, tetracyclines, macrolides, aminoglycosides, novobiocin, chloramphenicol, and sulfonamides, including/^-aminobenzoic acid (PABA) and its other analogs, are detectable. The incidence of false negative determinations among samples is about 1%; the incidence of false positives is about 3%. For negative cases, the relative standard deviations for repeatability ranged from 0 to 5% and for reproducibility from 0 to 6%. For positive cases, relative standard deviations ranged from 0 to 13% for repeatability and from 0 to 14% for reproducibility. The method has been adopted official first action.

scholarly journals Determination of Fat in Raw and Processed Milks by the Gerber Method: Collaborative Study

2001 ◽  
Vol 84 (5) ◽  
pp. 1499-1508 ◽  
Author(s):  
Dick H Kleyn ◽  
Joanna M Lynch ◽  
David M Barbano ◽  
M Jeffrey Bloom ◽  
Martin W Mitchell ◽  
...  
Keyword(s):  
Collaborative Study ◽  
Reference Method ◽  
Fat Content ◽  
Interlaboratory Study ◽  
Test Portion ◽  
Ether Extraction ◽  
Whole Milk ◽  
Relative Standard ◽  
Extraction Test ◽  
Fat Level

Abstract The Gerber method is used worldwide as a simple and rapid method for determining fat in raw and processed milks. However, the volume of the test portion used in the method has not been internationally agreed upon. A collaborative study was conducted to evaluate performance of the Gerber method using either a weighed test portion (11.13 g) or by a 10.77 mL test portion delivered by pipet. For each method, laboratories received 10 test samples: 5 raw and 5 pasteurized homogenized milks, 2 of which were blind duplicate pairs. Eleven and 10 laboratories participated in the evaluation of aliquot addition by weight and pipet, respectively. Mojonnier ether extraction (Method 989.05) was used as the reference method. Interlaboratory study statistics were similar between methods of test portion addition and between raw and processed materials; therefore, summary interlaboratory study statistics were pooled. The fat content of milk samples ranged from 0.96 to 5.48%. Absolute reproducibility and repeatability were not affected by fat level, and pooled statistical performance (invalid and outlier data removed) was (g fat/100 g milk) sr = 0.026, sR = 0.047, r = 0.074, and R = 0.132. Relative standard deviations increased with decreasing fat content, and were summarized by fat level: 1–2% fat milk, mean = 1.437, RSDr = 1.809%, RSDR = 3.271%; 2–6% fat milk, mean = 4.156, RSDr = 0.626%, RSDR = 1.131%. Compared with ether extraction, test results by the Gerber method were slightly lower (0.02% fat) using a weighed test portion and significantly lower (0.06% fat) using a 10.77 mL volume addition by pipet. A trend toward underestimating fat content at lower fat concentrations (1–2% fat) was observed with the weighed test portion but not when a pipet was used. The Associate Referee recommends that the Gerber method using a weighed test portion be adopted as First Action with applicability limited to whole milk.

Microbiological Determination of Penicillin G, Ampicillin, and Cloxacillin Residues in Milk

1979 ◽  
Vol 62 (6) ◽  
pp. 1247-1250
Author(s):  
Arnost B Vilim ◽  
Steven D Moore ◽  
Lyse Larocque
Keyword(s):  
Quantitative Determination ◽  
Bacillus Stearothermophilus ◽  
Bovine Mastitis ◽  
Penicillin G ◽  
Microbiological Method

Abstract A fast cylinder plate microbiological method was developed for the quantitative determination of penicillin G, ampicillin, and cloxacillin in milk. Agar plates seeded with stable spores of Bacillus stearothermophilus var. calidolactis were used and incubated at 64°C for 4½ hr. Standard curves were obtained for the following ranges of concentration of antibiotics: 0.004-0.064 IU penicillin G/mL, 0.0025-0.04 μg ampicillin/mL, and 0.03-0.48 μg cloxacillin/mL. The method is suitable for detecting penicillin residues in milk and for quantitative milk-out studies of the above antibiotics used in treatment of bovine mastitis.

Determination of Antibiotics in Meat Using Bacillus stearothermophilus Spores

1981 ◽  
Vol 44 (3) ◽  
pp. 194-200 ◽  
Author(s):  
M. BIELECKA ◽  
J. D. BALDOCK ◽  
A. W. KOTULA
Keyword(s):  
Sample Size ◽  
Bacillus Stearothermophilus ◽  
Test Organism ◽  
Optimal Concentration ◽  
Antibiotic Residues ◽  
Plate Method ◽  
Assay Medium ◽  
Organism Growth ◽  
Ph Variations

Ten parameters affecting sensitivity, accuracy and simplicity of the diffusion plate method for determining antibiotic residues in meat were evaluated with spores of Bacillus stearothermophilus as the test organism. Eight antibiotics were studied and included penicillin, bacitracin, tetracycline, chlortetracycline, oxytetracycline, streptomycin, erythromycin and neomycin. Sensitivity of the method was most influenced by concentration of inoculum, quantity of assay medium on the plate and sample size. The optimal concentration of inoculum was established as 2 × 105 spores/ml of medium, quantity of the assay medium on plate/100 mm dia., as 6 ml and quantity of sample poured on disc/12.7 mm dia., as 100 μl. The pH of the assay medium was also important to both antibiotic potency and test organism growth. The activity of streptomycin and erythromycin was the most sensitive to pH variations.

Precision Parameters for AOAC Bacillus stearothermophilus Disc Assay Based on FDA Milk Laboratory Quality Assurance 1982–1986 Samples

1989 ◽  
Vol 52 (12) ◽  
pp. 867-870 ◽  
Author(s):  
J. T. PEELER ◽  
J. W. MESSER ◽  
G. A. HOUGHTBY ◽  
J. E. LESLIE ◽  
J. E. BARNETT
Keyword(s):  
Quality Assurance ◽  
Bacillus Stearothermophilus ◽  
Penicillin G ◽  
Test Results ◽  
Inhibitory Substance ◽  
Milk Samples ◽  
Relative Standard ◽  
Standard Deviations ◽  
Estimate Precision ◽  
Disc Assay

Inhibitory substance (antibiotic) test results from State Split Milk Samples were used to estimate precision parameters and to compare antibiotic medium 4 (A4) and PM indicator (PM) agars. Five inhibitory substances (ampicillin, cephapirin, erythromycin, neomycin, and penicillin-G) were tested. Repeatability relative standard deviations (RSDr) ranged from 1.0 to 4.8%, and the reproducibility relative standard deviations (RSDR) ranged from 4.8 to 10.4%. Zone sizes of erythromycin, neomycin, and penicillin-G were significantly larger on PM agar (α = 0.05) than on A4 agar. The reverse was observed for cephapirin. No difference between agars was noted for ampicillin.

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