Inhibitor Report for: Tricresyl-phosphate

Over 1 billion pounds of organophosphorus (OP) chemicals are manufactured worldwide each year, including 70 million pounds of pesticides sprayed in the US. Current methods to monitor environmental and occupational exposures to OPs such as chlorpyrifos (CPS) have limitations, including low specificity and sensitivity, and short time windows for detection. Biomarkers for the OP tricresyl phosphate (TCP), which can contaminate bleed air from jet engines and cause an occupational exposure of commercial airline pilots, crewmembers and passengers, have not been identified. The aim of our work has been to identify, purify, and characterize new biomarkers of OP exposure. Butyrylcholinesterase (BChE) inhibition has been a standard for monitoring OP exposure. By identifying and characterizing molecular biomarkers with longer half-lives, we should be able to clinically detect TCP and OP insecticide exposure after longer durations of time than are currently possible. Acylpeptide hydrolase (APH) is a red blood cell (RBC) cytosolic serine proteinase that removes N-acetylated amino acids from peptides and cleaves oxidized proteins. Due to its properties, it is an excellent candidate for a biomarker of exposure. We have been able to purify APH and detect inhibition by both CPS and metabolites of TCP. The 120-day lifetime of the RBC offers a much longer window for detecting exposure. The OP-modified serine conjugate in the active site tryptic peptide has been characterized by mass spectrometry. This research uses functional proteomics and enzyme activities to identify and characterize useful biomarkers of neurotoxic environmental and occupational OP exposures.

The effect of tricresylphosphate (TCP) was studied in vitro and in vivo on the rat liver and brain enzymes acetylcholinesterase (ACC), butyrylcholinesterase (CHE), arylesterase (ARE), aliesterase (ALI), and the microsomal nicotinamide-adenine dinucleotide phosphate oxidase (NADPH2-oxidase) system. The results show that, in the male rat, TCP given intraperitoneally induces an increase in liver microsomal ARE AND NADPH2-oxidase and a decrease in ALI and cholinesterase; no activation of ARE and NADPH2-oxidase is observed in female rats.

INTRODUCTION: Aircraft maintenance workers may be exposed to organophosphates in hydraulic fluid and engine oil. Previous research has indicated that inhalation may not be the primary exposure route. This study sought to measure dermal contact and inhalation in conjunction with cholinesterase inhibition and determine if Air Force Specialty Code serves as an exposure predictor.METHODS: Aircraft maintenance workers were sampled for changes in acetylcholinesterase and butyrylcholinesterase. Dermal contact was measured using wrist-worn silicone passive dosimeters and inhalation exposure was measured using thermal desorption tube air sampling.RESULTS: Overall prevalence of any cholinesterase inhibition in the study population was 25.33%. Prevalence of inhibition of acetylcholinesterase and butyrylcholinesterase was 18.67% and 6.67%, respectively. The mean tributyl phosphate result was 1.71 ng of tributyl phosphate per gram of wristband (ng g(1)) [95% confidence interval (CI): 5.63, 9.05]. Triphenyl phosphate was more prevalent, with only one sample below the limit of detection (mean 1386.26 ng g(1); 95% CI: 7297.78, 10,070.31), and tricresyl phosphate was found in every sample (mean 4311.65 ng g(1); 95% CI: 8890.24, 17,512.31). No organophosphates were detected via air sampling.DISCUSSION: Workers experienced organophosphate exposure and cholinesterase inhibition, but the study was not large enough to establish a statistically significant association between exposure and disease. Exposure to organophosphate esters is more likely to occur through contact and absorption of chemicals through the skin than through inhalation of oil mists. Air Force Specialty Code does not appear to be a good predictor of exposure to organophosphates. Future studies should consider using a larger sample size.

Organophosphate esters (OPEs) draw growing concern about characterizing the potential risk on environmental health due to its wide usage and distribution. Two typical types of organophosphate esters (OPEs): tris (2-chloroethyl) phosphate (TCEP) and tricresyl phosphate (TCP) were selected to evaluate toxicity of OPEs to the soil organism like earthworm (Eisenia fetida). Histopathological examination (H&E), oxidative stress, DNA damage and RT-qPCR was used to identify the effects and potential mechanism of their toxicity. Hameatoxylin and eosin (H&E) demonstrated that intestinal cells suffered serious damage, and the observed up-regulation of chitinase and cathepsin L in mRNA levels confirmed it. Both TCEP and TCP significantly increased the DNA damage when the concentrations exceeded 1mg/kg (p<0.01), and a dose-response relationship was observed. In addition, TCEP and TCP also changed the acetylcholinesterase (AChE) activity and expression of genes associated with neurotoxic effects in earthworms even under exposure to low OPEs concentration (0.1mg/kg). Moreover, genes associated with nicotinic acetylcholine receptors (nAChR) and carrier protein further demonstrated that highest concentration of TCEP (10mg/kg) may have an overloading impact on the cholinergic system of E. fetida. Integrated Biological Response index (IBRv2) showed that TCEP exerted stronger toxicity than TCP under the same concentrations. We deduced that the observed intestinal damage, oxidative stress and neurotoxic effect might be the primary mechanisms of TCEP and TCP toxicity. This study provides insight into the toxicological effects of OPEs on earthworm model, and may be useful for risk assessment of OPEs on soil ecosystems.

Aerotoxic syndrome is assumed to be caused by exposure to tricresyl phosphate (TCP), an antiwear additive in jet engine lubricants and hydraulic fluid. CBDP (2-(ortho-cresyl)-4H-1,2,3-benzodioxaphosphoran-2-one) is the toxic metabolite of triortho-cresylphosphate, a component of TCP. Human butyrylcholinesterase (BChE; EC 3.1.1.8) and human acetylcholinesterase (AChE; EC 3.1.1.7) are irreversibly inhibited by CBDP. The bimolecular rate constants of inhibition (k(i)), determined under pseudo-first-order conditions, displayed a biphasic time course of inhibition with k(i) of 1.6 x 10(8) M(-1) min(-1) and 2.7 x 10(7) M(-1) min(-1) for E and E' forms of BChE. The inhibition constants for AChE were 1 to 2 orders of magnitude slower than those for BChE. CBDP-phosphorylated cholinesterases are nonreactivatable due to ultra fast aging. Mass spectrometry analysis showed an initial BChE adduct with an added mass of 170 Da from cresylphosphate, followed by dealkylation to a structure with an added mass of 80 Da. Mass spectrometry in (18)O-water showed that (18)O was incorporated only during the final aging step to form phospho-serine as the final aged BChE adduct. The crystal structure of CBDP-inhibited BChE confirmed that the phosphate adduct is the ultimate aging product. CBDP is the first organophosphorus agent that leads to a fully dealkylated phospho-serine BChE adduct.

The restoration of plasma acetylcholinesterase activity in mice following inhibition by organophosphorus pesticides and nerve agents has been attributed to synthesis of new enzyme. It is generally assumed that activity levels return to normal, are stable and do not exceed the normal level. We have observed over the past 10 years that recovery of acetylcholinesterase activity levels in mice treated with organophosphorus agents (OP) exceeds pretreatment levels and remains elevated for up to 2 months. The most dramatic case was in mice treated with tri-cresyl phosphate and tri-ortho-cresyl phosphate, where plasma acetylcholinesterase activity rebounded to a level 250% higher than the pretreatment activity. The present report summarizes our observations on plasma acetylcholinesterase activity in mice treated with chlorpyrifos, chlorpyrifos oxon, diazinon, tri-ortho-cresyl phosphate, tri-cresyl phosphate, tabun thiocholine, parathion, dichlorvos, and diisopropylfluorophosphate. We have developed a hypothesis to explain the excess acetylcholinesterase activity, based on published observations. We hypothesize that acetylcholinesterase activity is induced when cells undergo apoptosis and that consequently there is a rise in the level of plasma acetylcholinesterase.

The leakage of tricresyl phosphate-containing engine lubricants into aircraft cabin air, either from worn or defective engine seals or under normal operating conditions, is a serious concern for both the health and safety of the cabin occupants, since the oil contains one to five percent tricresyl phosphate (TCP) esters, known neurotoxins. The exposure of pilots is a particular concern since their impairment can affect their safe operation of the aircraft. Mass spectrometric (MS)-based protocols for documenting exposures of individuals are described that entail a rapid purification of the TCP-modified plasma enzyme butyrylcholinesterase (BChE). Following protease digestion of BChE, the modified active site peptide is characterized by MS analysis. Approaches for identifying safer engine oil additives are also described. Some general comments regarding the necessity of improving the quality and safety of the cabin air supply are presented.

Over 1 billion pounds of organophosphorus (OP) chemicals are manufactured worldwide each year, including 70 million pounds of pesticides sprayed in the US. Current methods to monitor environmental and occupational exposures to OPs such as chlorpyrifos (CPS) have limitations, including low specificity and sensitivity, and short time windows for detection. Biomarkers for the OP tricresyl phosphate (TCP), which can contaminate bleed air from jet engines and cause an occupational exposure of commercial airline pilots, crewmembers and passengers, have not been identified. The aim of our work has been to identify, purify, and characterize new biomarkers of OP exposure. Butyrylcholinesterase (BChE) inhibition has been a standard for monitoring OP exposure. By identifying and characterizing molecular biomarkers with longer half-lives, we should be able to clinically detect TCP and OP insecticide exposure after longer durations of time than are currently possible. Acylpeptide hydrolase (APH) is a red blood cell (RBC) cytosolic serine proteinase that removes N-acetylated amino acids from peptides and cleaves oxidized proteins. Due to its properties, it is an excellent candidate for a biomarker of exposure. We have been able to purify APH and detect inhibition by both CPS and metabolites of TCP. The 120-day lifetime of the RBC offers a much longer window for detecting exposure. The OP-modified serine conjugate in the active site tryptic peptide has been characterized by mass spectrometry. This research uses functional proteomics and enzyme activities to identify and characterize useful biomarkers of neurotoxic environmental and occupational OP exposures.

Aerotoxic syndrome is assumed to be caused by exposure to tricresyl phosphate, an additive in engine lubricants and hydraulic fluids that is activated to the toxic 2-(ortho-cresyl)-4H-1,3,2-benzodioxaphosphoran-2-one (CBDP). Currently, there is no laboratory evidence to support intoxication of airline crew members by CBDP. Our goal was to develop methods for testing in vivo exposure by identifying and characterizing biomarkers. Mass spectrometry was used to study the reaction of CBDP with human albumin, free tyrosine, and human butyrylcholinesterase. Human albumin made a covalent bond with CBDP, adding a mass of 170amu to Tyr411 to yield the o-cresyl phosphotyrosine derivative. Human butyrylcholinesterase made a covalent bond with CBDP on Ser198 to yield five adducts with added masses of 80, 108, 156, 170, and 186amu. The most abundant adduct had an added mass of 80amu from phosphate (HPO(3)), a surprising result given that no pesticide or nerve agent is known to yield phosphorylated serine with an added mass of 80amu. The next most abundant adduct had an added mass of 170amu to form o-cresyl phosphoserine. It is concluded that toxic gases or oil mists in cabin air may form adducts on plasma butyrylcholinesterase and albumin, detectable by mass spectrometry.

The effect of tricresylphosphate (TCP) was studied in vitro and in vivo on the rat liver and brain enzymes acetylcholinesterase (ACC), butyrylcholinesterase (CHE), arylesterase (ARE), aliesterase (ALI), and the microsomal nicotinamide-adenine dinucleotide phosphate oxidase (NADPH2-oxidase) system. The results show that, in the male rat, TCP given intraperitoneally induces an increase in liver microsomal ARE AND NADPH2-oxidase and a decrease in ALI and cholinesterase; no activation of ARE and NADPH2-oxidase is observed in female rats.

This study describes an outbreak of polyneuropathies in an artificial leather shoe factory. Thirty-seven females employed in the factory were examined; clinical findings and hematochemical data suggest a possible role of tricresylphosphate (TCP) in the genesis of these polyneuropathies. A high percentage of subjects evidenced definite signs of upper-motor neuron involvement, which appeared at an early or late stage of the disease. A statistically significant (P less than .001) reduction of red-blood-cell acetylcholinesterase (AChE) was found in the subjects. The ascertained upper-motor neuron involvement suggests a connection with previously described intoxications by TCP- by ingestion or occupational poisoning. The reduction of AChE seems attributable to TCP, and not to n-hexane, whose anti-AChE activity has never been demonstrated.