Accumulation,Metabolism And Biotransformation Of Perfluorohexyl Sulfonic Acid And Its Precursor Compounds In Rats

Perfluorohexanesulfonic acid（PFHx S）is a per-and polyfluoroalkyl substances（PFAS）containing 6 CF₂ perfluorosulfonic acids（PFSAs）.Because it is stable and not easily degradable in the environment,highly bioaccumulative and toxic,PFHx S and its related compounds were included in the Persistent Organic Pollutants（POPs）list in 2022.There are two main sources of PFAS contamination in the environment:direct use emissions and metabolic production of precursor compounds.PFHx S precursor substances have been detected in various environmental media and wild organisms in succession.However,the distribution,accumulation,transformation and metabolism of precursor substances in the organisms are still unclear.In this study,on the basis of the PFHx S and its precursor compounds qualitative and quantitative analysis method,the PFBS precursor compound NMe FBSE,compared the distribution of PFHx S and its sulfonamide ethanol precursor（N-methyl perfluorohexyl sulfonamide alcohol,NMe FHx SE）and the ammonium-containing precursors（N-3-dimethylamine propyl perfluorohexane sulfonamide,Am Pr-FHx SA）,providing data support for the comprehensive assessment of PFHx S biological occurrence and its origin.The main research contents are described as follows:The chromatography and MS parameters were optimized by HPLC-electrospray-mass spectrometry（HPLC-ESI-MS/MS）and quantitative analysis of PFHx S,NMe FHx SE,Am Pr-FHx SA,NMe FBSE and PFBS.The recoveries of target compounds spiked in liver,blood,kidney,bile,muscle,duodenal contents,as well as urine and fecal samples,and the working curves of liver and urine samples spiked were investigated by using ion-pair liquid-liquid extraction extraction extraction methods combined with SPE cleanup in extraction columns,respectively.The results showed that the recoveries of the spiked samples of each medium ranged from 75%to 120%,and the linear correlation coefficients were all greater than 0.99,which satisfied the analytical requirements for the quantification of the target compounds.The samples were further analyzed by electrostatic field orbitrap mass spectrometry（Q Exactive,QE）in high-resolution full scan and dd MS² modes,and the data were processed by Compounds Discover software in combination with online databases such as mz Cloud and Chemispider for non-target screening of metabolites by mass defects,retention times,secondary mass spectrometry maps and characteristic fragment ions.The Sprague Dawley rats were exposed by gavage at 5 mg/kg _bw PFHx S for 72 hours and sampled at specific time intervals to analysis of PFHx S content in each tissue and excreta.The results showed that PFHx S had the highest concentration in the blood,followed by the liver and kidney.With the change of exposure time,the content of PFHx S increased in each tissue during the first 48 h and decreased at 48–72 h.The blood ratio of PFHx S tissue（TBR）was less than 1,and the liver had the highest hepatic TBR value and the lowest muscle TBR value.Haemodynamic results showed that PFHx S concentrations showed clear bi-peaks over time with reabsorption within 26–72 h of exposure.Urine and fecal kinetics trends were consistent,with the PFHx S concentrations significantly higher in urine than in feces,indicating that PFHx S is more readily excreted through urine.The absorption and elimination half-life of PFHx S in rats were calculated by first-order kinetics,at 0.81 h and 36.9 h,respectively,indicating that PFHx S is easily absorbed into the blood and difficult to clear from the body.SD rats were exposed by gavage with 5 mg/kg_bw precursor compounds（NMe FHx SE,Am Pr-FHx SA and NMe FBSE）for 72 hours,and PFAS species and content in individual tissue and excreta samples were analyzed by target and non-target screening.Six to eight metabolites were screened in the precursor compound exposed group,mainly the corresponding perfluorosulfonamide carboxylic acid,perfluorosulfonamide and perfluorosulfonic acid compounds.Different from PFHx S exposure,the concentration of sulfonamide ethanol precursor compounds and their metabolites in all tissues decreased with the increase of exposure time,while the concentration of quaternary ammonium-containing precursor compounds and their metabolites increased with the exposure time.C6 exposure group metabolites were mainly distributed in blood and liver,and C4 exposure group metabolites were mainly distributed in kidney and bile,with the highest concentrations of sulfonamide carboxylic acid and sulfonamide compounds.Metabolic kinetics results showed that blood Am Pr-FHx SA concentration showed double reabsorption within 12 h of exposure,and unlike the PFHx S exposed group,the PFHx S concentration increased increased over 72 h,indicating that PFHx S is gradually produced within the organism and takes longer time to reach equilibrium.The Am Pr-FHx SA and NMe FHx SE exposure groups were comparable in bioavailability and the peak attainment concentrations were similar in both groups,indicating comparable uptake in the blood of rats at comparable perfluorocarbon chains.The comparison of precursor compound exposed groups with different carbon chain lengths and different terminal functional groups showed that the C4 exposed group was the fastest and mainly excreted through feces and urine;C6 sulfonamide ethanol was more rapidly metabolized than the exposed group with quaternary ammonium precursor,and the total amount was calculated to show that the C6 precursor-exposed group was mainly excreted in the feces.Metabolic pathway analysis showed that the precursor compounds in rats mainly biotransformed through a series of reaction processes,such as hydrolysis,oxidation,decarboxylation and demethylation,successively producing perfluorosulfonamide carboxylic acid,perfluorosulfoprimary amide,perfluorosulfonamide secondary amine and perfluorosulfonamide,and finally PFSAs.