Improvement of a commercial probiotic product through metabolite profiling and quality assurance system development

Abstract

In recent years, use of probiotic formulation for animal health and nutrition has gained popularity to produce high quality animal based food products i.e. milk and meat. As a result, number of probiotic products in the market has increased. However, there is an increasing concern about the quality and safety of these products and the importance of ensuring quality control of the probiotics has widely been accepted. BioBrew Ltd produces liquid animal probiotics widely administered to the farm animals in New Zealand. In this study more than 300 samples at various stages of production were analysed. Sampling across New Zealand was carried out for a period of eight months (August 2013 to March 2014). BioBrew Animal Nutritional supplement (BBAN) is molasses fermented liquid formulation having multi-species of Lactic Acid Bacteria (LAB) and yeast cells. Viability study was undertaken to enumerate LAB and yeast cell counts in BBAN using traditional plate count methods. Production involved three stages fermentation and viable LAB cell counts in stage 1, 2 and 3 differed significantly (P < 0.05). However, viable counts in stage 3 and final product with different storage days did not differ significantly (P < 0.05) and cell densities on average were found to be more than 109 CFU/ml in the product, which was higher than Minimum Suggested Level (MSL). On contrary, average yeast cell densities reached maximum in stage 1 and 2 (9.97 and 10.24 log CFU/ml respectively) dropped to 5.47 log CFU/ml at the end of storage period. On average, yeast cell counts in the product were just below the MSL, irrespective of product from different production sites. Analysis confirmed that the product contained considerable amount of lactic acid and ethanol in BBAN. Headspace Solid-Phase Micro-Extraction Gas Chromatograph Mass Spectrometry (HSSPME/GCMS) analysis was used to detect the semi-volatile and volatile metabolites in this probiotic formulation. Lactic acid, ethanol and butyric acid were primary metabolites detected in higher quantities in BBAN formulation. Lactic acid, acetic acid, ethanol, butyric acid, diacetyl were the important metabolites identified with antimicrobial properties. Products were screened for potential foodborne pathogens and confirmed BBAN formulation were not detected positive for the presence of any bacterial pathogens over the period of study. However, contamination with yeast (Rhodotorula rubra) observed in 2 batches during the period of study. Favourable conditions for this detected yeast contamination were identified and were effectively controlled through measures suggested in this study. Based on our study, model quality assurance (QA) system was proposed to produce consistent, high quality and safe BBAN. Model QA system includes products testing for pH, temperature, viable counts, safety assessment and presence of metabolites at different stages as well as during various storage days. In summary, it was confirmed that BBAN met the quality control parameters. BBAN has lactic acid, butyric acid, ethanol, acetic acid and diacetyl as major bioactive compounds. BBAN batches tested were free from pathogenic contamination. This study suggested a model QA system to enhance quality and safety of the product.