A Review of Probiotic Ingredient Safety Supporting Monograph Development Conducted by the United States Pharmacopeia (USP).

The United States Pharmacopeia (USP) is an independent, nonprofit science-based organization whose mission is to improve global health through public standards and related products for medicines, food and dietary supplements. Probiotic-based dietary supplements are increasingly popular in the marketplace and USP has developed fourteen monographs specific to probiotic ingredients, including representatives from the Genera Lactobacillus, Bacillus, Streptococcus, and Bifidobacterium. These monographs include the definition of the article, tests for identification, quantification assays (enumeration in the case of probiotics), limits for contaminants, and other quality parameters when appropriate. In addition to quality, the USP also considers the safety of probiotics for monograph development. This report includes an overview of the USP admission evaluation process for probiotics as well as a tabular summary of the probiotic monographs currently available. Pharmacopeia monographs can guide manufacturers and brand owners and protect consumers through establishment of quality standards.

Probiotic and postbiotic analytical methods: a perspective of available enumeration techniques.

Probiotics are the largest non-herbal/traditional dietary supplements category worldwide. To be effective, a probiotic strain must be delivered viable at an adequate dose proven to deliver a health benefit. The objective of this article is to provide an overview of the various technologies available for probiotic enumeration, including a general description of each technology, their advantages and limitations, and their potential for the future of the probiotics industry. The current "gold standard" for analytical quantification of probiotics in the probiotic industry is the Plate Count method (PC). PC measures the bacterial cell's ability to proliferate into detectable colonies, thus PC relies on cultivability as a measure of viability. Although viability has widely been measured by cultivability, there has been agreement that the definition of viability is not limited to cultivability. For example, bacterial cells may exist in a state known as viable but not culturable (VBNC) where the cells lose cultivability but can maintain some of the characteristics of viable cells as well as probiotic properties. This led to questioning the association between viability and cultivability and the accuracy of PC in enumerating all the viable cells in probiotic products. PC has always been an estimate of the number of viable cells and not a true cell count. Additionally, newer probiotic categories such as Next Generation Probiotics (NGPs) are difficult to culture in routine laboratories as NGPs are often strict anaerobes with extreme sensitivity to atmospheric oxygen. Thus, accurate quantification using culture-based techniques will be complicated. Another emerging category of biotics is postbiotics, which are inanimate microorganisms, also often referred to as tyndallized or heat-killed bacteria. Obviously, culture dependent methods are not suitable for these products, and alternative methods are needed for their quantification. Different methodologies provide a more complete picture of a heterogeneous bacterial population versus PC focusing exclusively on the eventual multiplication of the cells. Alternative culture-independent techniques including real-time PCR, digital PCR and flow cytometry are discussed. These methods can measure viability beyond cultivability (i.e., by measuring cellular enzymatic activity, membrane integrity or membrane potential), and depending on how they are designed they can achieve strain-specific enumeration.

Considerations for determining safety of probiotics: A USP perspective.

A history of safe use is a backbone of safety assessments for many current probiotic species, however, there is no global harmonization regarding requirements for establishing probiotic safety for use in foods and supplements. As probiotic manufacturers are increasingly seeking to use new strains, novel species, and next-generation probiotics, justification based on a significant history of use may be challenged. There are efforts underway by a variety of stakeholders, including the United States Pharmacopeia (USP), to develop best practices guidelines for assessing the quality and safety of probiotics. A current initiative of the USP seeks to provide expert advice specific to safety considerations for probiotics. Toward this goal, this review provides a helpful summary guide to global regulatory guidelines. We question the suitability of traditional animal toxicology studies designed for testing chemicals for relevance in assessing probiotic safety. This includes discussion of the use of excessive dose levels, the length of repeated dose toxicity studies needed, and the most suitable animal species used in toxicology studies. In addition, the importance of proper manufacturing practices with regard to final product safety are also included. Thus, an outline of essential parameters of a comprehensive safety assessment for a probiotic are provided.