Endotoxin Analysis: Correlation Between Biological and Chemical Methods
Journal Title: Biomedical Journal of Scientific & Technical Research (BJSTR) - Year 2018, Vol 19, Issue 4
Abstract
Assessment of occupational exposure to airborne endotoxins was studied for several years but still different procedures are used for sampling and analysis. Among analytical methods LAL test is the most used. A different approach is represented by chemical analyses, based on the detection of the β- hydroxy fatty acids, as markers of lipopolysaccharide (LPS). In this study an analytical method using HPLC-MS/MS was applied to quantify the content of β- hydroxy-dodecanoic (3-OH C12) and β-hydroxytetradecanoic (3-OH C14) acids in thirteen Gram- negative bacterial strains. In order to compare biological and chemical results cell suspensions were analyzed also by Kinetic Chromogenic LAL test. The results obtained by HPLC-MS/MS show that three samples did not contain 3-OH C12 at all, while 3-OH C14 was measured in a concentration range from 11.1 to 4007.5µg/L. Endotoxin concentration measured by LAL assay was in the range of 102 -103 µg/L. Pearson’s test showed that the sum of fatty acids by HPLC-MS/MS correlates positively with the LAL Test (r= 0.79) confirming that it can be considered a reliable marker of endotoxin contamination. Principal Component Analysis (PCA) confirms above results and provides additional information grouping microorganisms related with each other on the basis of their biochemical features, habitat and disease spectra. Endotoxins are unavoidable ubiquitous microbiological contaminants; they may be present in food, environment and clinical products. Their inhalation can poses serious risks to human health, from respiratory and general symptoms such as fever, coughing, irritation and chest congestion up to chronic bronchitis and organic dust toxic syndrome (ODTS) [1,2]. For this reason, assessment of exposure to these components was studied for several years, especially in occupational environments, but a large number of different procedures are still used for sampling and analysis; this makes difficult to compare results and to establish an Occupational Exposure Limit (OEL) for these biological agents [3-5]. Among analytical methods, the most used is the Limulus Amebocyte Lysate (LAL) test, in its kinetic chromogenic version, which measures the relative reactivity of endotoxins with limulus lysate providing rapid and sensitive results. However, several critical aspects of this test are reported in literature: the cross-reactivity with other types of biomolecules such as β-d-glucans and peptidoglycans, the underestimation of the results due to a fraction non-water soluble not detectable with LAL test and the influence of endotoxin content after repeated freezing/thawing of samples [6,7]. Furthermore, this bioassay may underestimate endotoxin exposure because cellbound endotoxins, that may be associated with respiratory disease, are not detected [8].
Authors and Affiliations
Emilia Paba, Alessandra Chiominto, Anna Maria Marcelloni, Francesca Tombolini, Enrico Paci, Giovanna Tranfo
Keywords
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- EP ID EP622416
- DOI 10.26717/BJSTR.2019.19.003339
- Views 181
- Downloads 0
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