Performing Department
Population Health & Reproduction
Non Technical Summary
Feeding waste milk (WM) to preweaned dairy calves has been shown to significantly increase selection for resistance to antimicrobial drugs of great importance to human and veterinary health, such as third generation cephalosporins.previous study evaluating degradation of ceftiofur in water suggested that thecombinedeffectof high pH and heat treatment resulted in rapid degradation of ceftiofur, a drug commonly found in waste milk.This project aims to degrade ceftiofur in milk using approaches that are feasible for on-farm implementation, adn that would allow the use of waste milk as a safe source of nutrients for dairy calves.
Animal Health Component
100%
Research Effort Categories
Basic
20%
Applied
20%
Developmental
60%
Goals / Objectives
Develop a method for degrading ceftiofur in whole milk by evaluating the most effective treatment combination between alkaline pH (8.0 and 10.0) and heat treatment at 63 °C (145°F) for 30 minutes, 72°C (161°F) for 15 seconds, or 92ºC for 20 minutes.
Project Methods
Spiking milk with antimicrobials: saleable whole milk will be spiked using stocks solution of antimicrobials previously prepared by diluting powder ceftiofur sodium in distilled water as previously described1. Final drug concentration in milk will be 0.25 ppm, replicating ceftiofur concentrations identified in WM at dairy farms5.Preparing milk with pH of 8 and 10: sodium bicarbonate will be added to whole milk samples until desired pH is achieved ( 8 and 10)4. This will be done prior to adding ceftiofur so that the time needed for degradation of the drug can more accurately be measured. The choice for the two alkaline pH selected (8 and 10) are based on ceftiofur degradation ability at this pH and potential impacts on milk nutrients, palatability and ability to neutralize milk pH after alkaline pH treatment (which will be conducted in future studies)4,14.Heat treatments: three heat treatment methods will be used, from which the first two are currently used on dairy farms (a & b): a) Low-temperature long-time (LTLT), milk heated to 63 °C (145°F) and held at that temperature for 30 minutes; b) High temperature short time (HTST), milk heated to 72°C (161°F) for 15 seconds; and c) milk heated to 92ºC for 20 minutes (previous data indicates degradation of ceftiofur at this temperature)15.pH and heat treatments: once milk is at the desired pH and ceftiofur is added to the desired final concentration, milk will immediately be treated using the three heat treatments proposed. Samples will be collected for antimicrobial drug quantification prior and after heat treatment of milk. A pH treated samples will not be processed by heat treatments, and will serve to measure the added effect of heat on the degradation of ceftifour.Control samples: A control group with milk with natural pH (~6.5-6.7) will also be spiked with ceftiofur and treated with the three heat treatment protocols. Samples will be collected at the same initial and final time when heat treatment samples are collected from the treatment group. These samples will serve as a control to evaluate the added impacts of pH treatments and heat treatment compared to no treatment. Control milk samples will be maintained at 20ºC, simulating approximate average annual temperatures in California Central Valley16.Sample size: A total of 4 repetitions will be conducted for each pH and heat treatment combination. Based on reported references for heat and pH stability of antimicrobials, we estimate an 80.5% statistical power to identify a significant difference between samples after treatment when compared to the control group (α = 0.05, standard deviation= 0.22, difference to detect= 0.18)4,15,17.Drug quantification: cefiotfur quantification in milk samples will be conducted in using High Performance Liquid Chromatography (HPLC). The assay to be used will convert ceftiofur and its actives metabolite (desfuroylceftiofur and desfuroylceftiofur conjugates), to desfuroylceftiofur acetamide (DAC) for analysis18. DAC is a more representative quantification of ceftiofur and its metabolites, which are microbiologically active19. Sample analysis will be conducted at the Veterinary Drug Residue Laboratory at UC Davis (Dr Lisa Tell), as indicated by a letter of support letter.Statistical Analysis: the combined effect of treatment of milk with alkaline pH and heat treatment on ceftiofur degradation will be evaluated using a mixed linear model accounting for repeated measures (batches), using the Glimmix procedure in SAS. In these models the dependent variables will be the percent degradation and biological half-life (hrs), and the independent variable will be the sampling points. A Tukey honestly significant difference (HDS) test will be performed to compare time points.