Saccharomyces cerevisiae Cell Wall-Based Adsorbent Reduces Aflatoxin B1 Absorption in Rats.

Mycotoxins are naturally occurring toxins that can affect livestock health and performance upon consumption of contaminated feedstuffs. To mitigate the negative effects of mycotoxins, sequestering agents, adsorbents, or binders can be included to feed to interact with toxins, aiding their passage through the gastrointestinal tract (GI) and reducing their bioavailability. The parietal cell wall components of Saccharomyces cerevisiae have been found to interact in vitro with mycotoxins, such as, but not limited to, aflatoxin B1 (AFB1), and to improve animal performance when added to contaminated diets in vivo. The present study aimed to examine the pharmacokinetics of the absorption of radiolabeled AFB1 in rats in the presence of a yeast cell wall-based adsorbent (YCW) compared with that in the presence of the clay-based binder hydrated sodium calcium aluminosilicate (HSCAS). The results of the initial pharmacokinetic analysis showed that the absorption process across the GI tract was relatively slow, occurring over a matter of hours rather than minutes. The inclusion of mycotoxin binders increased the recovery of radiolabeled AFB1 in the small intestine, cecum, and colon at 5 and 10 h, revealing that they prevented AFB1 absorption compared with a control diet. Additionally, the accumulation of radiolabeled AFB1 was more significant in the blood plasma, kidney, and liver of animals fed the control diet, again showing the ability of the binders to reduce the assimilation of AFB1 into the body. The results showed the potential of YCW in reducing the absorption of AFB1 in vivo, and in protecting against the damaging effects of AFB1 contamination.

The Analysis of Docosahexaenoic Acid (DHA) in Dried Dog Food Enriched with an Aurantiochytrium limacinum Biomass: Matrix Extension Validation and Verification of AOAC Method 996.06.

BACKGROUND: Docosahexaenoic acid (DHA) plays an important role in brain and retinal development in dogs. However, supranutritional dietary supplementation can result in health issues, including gastrointestinal bleeding, making the accurate analysis of DHA in dog food important for nutritional and welfare regulatory compliance. OBJECTIVE: The aim of this study was to conduct a validation and verification of the AOAC 996.06 method, and hence establish its fitness for purpose, for the analysis of DHA in dried dog food supplemented with a heterotrophically grown unextracted DHA-rich Aurantiochytrium limacinum biomass. METHODS: The AOAC 996.06 method, which involves the use of gas chromatography coupled to flame ionization detection (GC-FID), was used to conduct a validation of the analysis of DHA in dried dog food and the results were verified in a second laboratory. RESULTS: The method was found to be linear over the ranges analyzed and results were found to be within the acceptance criteria for precision and accuracy, verifying the applicability for this matrix. The selectivity and sensitivity of the method were also determined. CONCLUSIONS: The AOAC 996.06 method is fit for purpose for the analysis of DHA in dry dog food kibble. HIGHLIGHTS: The method can be applied to various dog food samples, supplemented with an unextracted Aurantiochytrium limacinum biomass, using alternative manufacturing methods, i.e. pelleted and extruded with no significant matrix effects being observed.

A comparison of analytical methods for the determination of urea in yeast-based food and feed ingredients.

Urea is a non-protein nitrogen compound, authorised in the EU as a nutritional source of nitrogen in animal feed intended for ruminants with a functional rumen. The Rapid Alert System for Food and Feed (RASFF) is the EU online platform through which food and feed safety risks are reported. During 2017, several rapid alerts were raised in the EU by individual member states regarding the presence of unlabelled urea in feed-grade yeast. The presence of urea in animal feed is analysed with a spectro-colorimetric method, which is the EU official control method. Other urea analytical methods include enzymatic and LC-MS methods, which are also investigated in this study. The main objectives of this study were to analyse yeast feed-grade and food-grade products with independent laboratories using the EU official control method, as well as enzymatic and LC-MS methods. Comparison of results from the spectro-colorimetric, enzymatic and LC-MS methods for reliability, consistency and accuracy determined the most reliable method for the quantification of urea in animal feed. Yeast food and feed samples were tested using four independent laboratories. Results demonstrated a lack of consistency both within the same laboratory and between separate laboratories for urea quantification using the EU official control method. Urea quantification results for the AOAC 941.04 enzymatic method were found to be much higher than anticipated. The LC-MS method yielded the most consistent and reliable results. In conclusion, urea quantification by LC-MS analysis has the potential to be a viable method for urea analysis of yeast samples.

Fitness for purpose and stability assessment of long-chain polyunsaturated fatty acids in chicken tissues.

The global popularity of chicken in human diets make it an obvious choice for enrichment with DHA and LC-PUFA. There is presently a need for a robust method for the analysis of chicken tissues and where the fitness for purpose of the method has been demonstrated. The purpose of this paper is to present the validation of the AOAC method 996.06 on five different chicken tissues; breast, thigh, skin, kidney and liver. The parameters investigated as part of the validation study included; linearity and range, the limit of detection and limit of quantification, accuracy, repeatability, inter-analyst reproducibility, and specificity. The method was further applied to assess the stability of DHA and other specific LC-PUFA in chicken tissues over short and long timepoints. The disclosure of this information is relevant for researchers concerned with the analysis of LC-PUFA in regulatory and efficacy studies.

The validation & verification of an LC/MS method for the determination of total docosahexaenoic acid concentrations in canine blood serum.

Docosahexaenoic acid (DHA), is an omega 3 fatty acid (n-3 FA) that has been shown to play a role in canine growth and physiological integrity and improvements in skin and coat condition. However, potential adverse effects of n-3 FA specifically, impaired cellular immunity has been observed in dogs fed diets with elevated levels of n-3 FA. As such, a safe upper limit (SUL) for total n-3 FAs (DHA and EPA) in dogs has been established. Considering this SUL, sensitive methods detecting DHA in blood serum as a biomarker when conducting n-3 FA supplementation trials involving dogs are required. In this study, an LC-ESI-MS/MS method of DHA detection in dog serum was validated and verified. Recovery of DHA was optimized and parallelism tests were conducted with spiked samples demonstrating that the serum matrix did not interfere with quantitation. The stability of DHA in serum was also investigated, with -80 °C considered suitable when storing samples for up to six months. The method was linear over a calibration range of 1-500 µg/mL and precision and accuracy were found to meet the requirements for validation. This method was verified in an alternative laboratory using a different analytical system and operator, with the results meeting the criteria for verification.

Validation and Verification of a Liquid Chromatography-Mass Spectrometry Method for the Determination of Total Docosahexaenoic Acid in Pig Serum.

The paper presents the validation and verification of an analytical method for the determination of total docosahexaenoic acid (DHA) in pig serum by liquid chromatography-electrospray ionization-tandem mass spectrometry. The characteristics studied during the validation included precision and accuracy, limit of quantitation (LOQ), selectivity, calibration range and linearity, parallelism, and stability. A separate verification study was also performed. The method was linear over the range. Precision and accuracy met acceptance criteria at all levels, and the LOQ was determined as 1 µg/mL. Parallelism experiments were conducted to show that there was no bias introduced in using a surrogate matrix to quantify DHA. Recoveries of free DHA were obtained for quality control samples, and stability studies were conducted over 1, 7, 31, and 180 days. The results of the verification study were in line with the validation study, and in conclusion, the method was deemed fit for purpose for measuring total DHA in pig serum.

An Accurate Assessment of Docosahexaenoic Acid in Laying Hen Serum for Regulatory Studies.

Diets rich in omega-3 fatty acids (n-3 FA) have been associated with several health benefits. With the increased interest in n-3 FA both scientifically and societally, the accurate detection of such analytes has become increasingly important. Recently, tandem mass spectrometry (MS/MS) with electrospray ionization interface (ESI), hyphenated to both gas chromatography (GC) and liquid chromatography (LC), has become a valuable tool in the detection of docosahexaenoic acid (DHA). Liquid chromatography-electrospray ionization interface-tandem mass spectrometry methods have been developed for the determination of DHA in canine and poultry species. The objective of this article is to investigate whether LC-ESI-MS/MS is fit for purpose for the determination of DHA in laying hen serum. The disclosure of this work will be beneficial for researchers investigating poultry enrichment for regulatory and toxicological studies. The method was found to be linear over the range. Precision and accuracy results met acceptance criteria and the Limit of Quantitation (LOQ) was established as 1 µg/mL. Recoveries of DHA were obtained for quality control samples and stability studies were performed. The results of the verification study complimented those of the validation study. In summation, the method was established as fit for purpose for measuring total DHA in laying hen serum.