A highly sensitive HPLC method for determination of nanomolar concentrations of dipicolinic acid, a characteristic constituent of bacterial endospores

Quantification of Endospore-Forming Firmicutes by Quantitative PCR with the Functional Genespo0A

“…For example, due to the heterogeneity under culture conditions, multiple assays have to be carried out to retrieve as many different EFF species as possible. Culture-independent approaches are mainly based on the quantification of dipicolinic acid (DPA), a unique biomarker for endospores (19). Different methods have been developed to extract and quantify DPA from endospores (10,(19)(20)(21)(22), with detection limits ranging from 10 8 endospores/g of sediment (via high-performance liquid chromatography) (23), 10 5 endospores/g of soil (via complexation to terbium and time-resolved fluorometry) (20,23), or 10 3 endospores/ml (via complexation to terbium and direct fluorescence microscopy) (24).…”

“…Despite the relatively high sensitivity of DPA-based methods, a disadvantage of those assays is that they cannot take into account vegetative cells of EFF. Furthermore, terbium fluorescence is easily quenched with substances commonly found in environmental samples (e.g., humic acids [23] or organophosphates [19]), leading to a dramatic decrease in the detection limit.…”

Nanopore back titration analysis of dipicolinic acid

“…DPA, a unique component of bacterial endospores such as Bacillus anthracis that composes approximately 10% of their dry weight , is an important useful marker molecule in the detection of anthrax spores. Thus far, numerous techniques have been developed for the highly sensitive detection of DPA, including gas‐liquid chromatography, surface‐enhanced Raman spectroscopy, lanthanide ion‐based fluorescence, and so on . However, most of these methods require use of instruments relatively expensive and not easily portable.…”

Assessment of spore contamination in pepper by determination of dipicolinic acid with a highly sensitive HPLC approach

“…In contrast, quantification of terbium dipicolinate complexes using a fluorescence detector has an about three orders of magnitude higher sensitivity (Fichtel, Kö ster, Scholz-Bö ttcher et al, 2007) and allowed the detection of spore contaminations in all analysed pepper samples. Fig.…”

“…The approach is based on the determination of dipicolinic acid (DPA, pyridine-2,6-dicarboxylic acid), a universal and specific component of bacterial endospores (Powell, 1953), which makes up 5-14% of endospore dry weight (Murrell, 1969;Murrell & Warth, 1965 shown to be a useful analytical indicator for the presence of bacterial endospores and even permits to estimate total spore numbers, if spore DPA contents are known (Fichtel, Kö ster, Rullkötter, & Sass, 2007). For quantification of endospore-derived DPA in pepper samples we used a sensitive high-performance liquid chromatographic method with indirect fluorescence detection of DPA, which is described in detail by Fichtel, Kö ster, Scholz-Bö ttcher, Sass, and Rullkö tter (2007). The method is based on detection of terbium dipicolinate fluorescence after chromatographic separation and post-column complexation of DPA.…”