Article

A robust method to quantify low molecular weight contaminants in heparin: Detection of tris(2-n-butoxyethyl) phosphate

Departamento de Bioquímica e Biologia Molecular, Universidade Federal do Paraná, Curitiba, PR CEP: 81531-980, Brazil.
The Analyst (Impact Factor: 4.11). 06/2011; 136(11):2330-8. DOI: 10.1039/c0an01010c
Source: PubMed

ABSTRACT

Recently, oversulfated chondroitin sulfate (OSCS) was identified in contaminated heparin preparations, which were linked to several adverse clinical events and deaths. Orthogonal analytical techniques, namely nuclear magnetic resonance (NMR) and capillary electrophoresis (CE), have since been applied by several authors for the evaluation of heparin purity and safety. NMR identification and quantification of residual solvents and non-volatile low molecular contaminants with USP acceptance levels of toxicity was achieved 40-fold faster than the traditional GC-headspace technique, which takes ~120 min against ~3 min to obtain a (1)H NMR spectrum with a signal/noise ratio of at least 1000/1. The procedure allowed detection of Class 1 residual solvents at 2 ppm and quantification was possible above 10 ppm. 2D NMR techniques (edited-HSQC (1)H/(13)C) permitted visualization of otherwise masked EDTA signals at 3.68/59.7 ppm and 3.34/53.5 ppm, which may be overlapping mononuclear heparin signals, or those of ethanol and methanol. Detailed NMR and ESI-MS/MS studies revealed a hitherto unknown contaminant, tris(2-n-butoxyethyl) phosphate (TBEP), which has potential health risks.

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    • "The superfamily of NRs includes 6 subfamilies (ERs, AR, TR, MR, GR, PPAR) (Zhao et al., 2015) and an orphan receptor subfamily that comprise the metazoan transcription factors (Kojima et al., 2013). Recently, several OPEs have been found to induce developmental toxicity and endocrine disruption both in vivo and in vitro (Fu et al., 2013; Han et al., 2014; Kojima et al., 2013; Liu et al., 2013, 2012b; McGee et al., 2012; Porter et al., 2014; Sassaki et al., 2011; Wang et al., 2013a). "

    Full-text · Dataset · Dec 2015
  • Source
    • "The superfamily of NRs includes 6 subfamilies (ERs, AR, TR, MR, GR, PPAR) (Zhao et al., 2015) and an orphan receptor subfamily that comprise the metazoan transcription factors (Kojima et al., 2013). Recently, several OPEs have been found to induce developmental toxicity and endocrine disruption both in vivo and in vitro (Fu et al., 2013; Han et al., 2014; Kojima et al., 2013; Liu et al., 2013, 2012b; McGee et al., 2012; Porter et al., 2014; Sassaki et al., 2011; Wang et al., 2013a). "

    Full-text · Dataset · Dec 2015
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    • "The superfamily of NRs includes 6 subfamilies (ERs, AR, TRα, MR, GR, PPARα) (Zhao et al., 2015) and an orphan receptor subfamily that comprise the metazoan transcription factors (Kojima et al., 2013). Recently several OPEs have been found to induce developmental toxicity and endocrine disruption both in vivo and in vitro (Fu et al., 2013; Han et al., 2014; Kojima et al., 2013; Liu et al., 2013; Liu et al., 2012b; McGee et al., 2012; Porter et al., 2014; Sassaki et al., 2011; Wang et al., 2013a). We had previously demonstrated that TBOEP could interfere with endocrine axes, including hypothalamus-pituitary-thyroidal (HPT), hypothalamuspituitary-adrenal (HPA) and hypothalamus-pituitary-gonadal (HPG) axes in early-stage zebrafish. "
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    ABSTRACT: As one substitute for phased-out brominated flame retardants (BFRs), tris(2-butoxyethyl) phosphate (TBOEP) is frequently detected in aquatic organisms. However, knowledge about endocrine disrupting mechanisms associated with nuclear receptors caused by TBOEP remained restricted to results from in vitro studies with mammalian cells. In the study, results of which are presented here, embryos/larvae of zebrafish (Danio rerio) were exposed to 0.02, 0.1 or 0.5μM TBOEP to investigate expression of genes under control of several nuclear hormone receptors (estrogen receptors (ERs), androgen receptor (AR), thyroid hormone receptor alpha (TRα), mineralocorticoid receptor (MR), glucocorticoid receptor (GR), aryl hydrocarbon (AhR), peroxisome proliferator-activated receptor alpha (PPARα), and pregnane×receptor (P×R)) pathways at 120hpf. Exposure to 0.5μM TBOEP significantly (p<0.05, one-way analysis of variance) up-regulated expression of estrogen receptors (ERs, er1, er2a, and er2b) genes and ER-associated genes (vtg4, vtg5, pgr, ncor, and ncoa3), indicating TBOEP modulates the ER pathway. In contrast, expression of most genes (mr, 11βhsd, ube2i,and adrb2b) associated with the mineralocorticoid receptor (MR) pathway were significantly down-regulated. Furthermore, in vitro mammalian cell-based (MDA-kb2 and H4IIE-luc) receptor transactivation assays, were also conducted to investigate possible agonistic or antagonistic effects on AR- and AhR-mediated pathways. In mammalian cells, none of these pathways were affected by TBOEP at the concentrations studied. Receptor-mediated responses (in vivo) and mammalian cell lines receptor binding assay (in vitro) combined with published information suggest that TBOEP can modulate receptor-mediated, endocrine process (in vivo/in vitro), particularly ER and MR.
    Full-text · Article · Nov 2015 · Aquatic toxicology (Amsterdam, Netherlands)
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