Uptake, translocation, accumulation, and metabolism of fluroxypyr-meptyl and oxidative stress induction in rice seedling.

Fluroxypyr-meptyl (FLUME) is heterocyclic herbicide with internal absorption and transmission characteristics. Owing to its low cost and rapid efficacy, it has been widely used to control broad-leaved weeds in wheat, corn, and rice fields. However, the uptake, translocation, accumulation, and metabolism of FLUME in rice seedlings and the extent of oxidative stress induced by it remain largely unknown, which consequently restricts the comprehensive risk assessment of FLUME residues in the environment during rice production. Hence, we systematically investigated the growth and physiological responses of rice to FLUME and analyzed its uptake, translocation, accumulation, and metabolism in rice seedlings. The results indicated that under 0-0.12 mg/L FLUME treatment, only a small proportion of FLUME was translocated upward and accumulated in rice shoots following absorption via roots, with all the translocation factor values being < 1. Moreover, the distribution and enrichment ability of FLUME in rice seedlings were greater in roots than in shoots. Furthermore, we revealed that FLUME accumulation in rice seedlings evidently inhibited their growth and activated the defense system against oxidative stress, with an increase in the activity of antioxidant and detoxifying enzymes. In addition, multiple metabolic reactions of FLUME were observed in rice seedlings, including dehalogenation, hydroxylation, glycosylation, acetylation, and malonylation. Our study provides systematic insights into the uptake, translocation, accumulation, and metabolism of FLUME in rice seedlings as well as the oxidative stress induced by FLUME accumulation, which can help improve FLUME applications and environmental risk assessments in crops.

Multiple strategies for metabolic engineering of Escherichia coli for efficient production of glycolate.

Glycolate is a bulk chemical with wide applications in the textile, food processing, and pharmaceutical industries. Glycolate can be produced from glucose via the glycolysis and glyoxylate shunt pathways, followed by reduction to glycolate. However, two problems limit the productivity and yield of glycolate when using glucose as the sole carbon source. The first is a cofactor imbalance in the production of glycolate from glucose via the glycolysis pathway, since NADPH is required for glycolate production, while glycolysis generates NADH. To rectify this imbalance, the NADP+ -dependent glyceraldehyde 3-phosphate dehydrogenase GapC from Clostridium acetobutylicum was introduced to generate NADPH instead of NADH in the oxidation of glyceraldehyde 3-phosphate during glycolysis. The soluble transhydrogenase SthA was further eliminated to conserve NADPH by blocking its conversion into NADH. The second problem is an unfavorable carbon flux distribution between the tricarboxylic acid cycle and the glyoxylate shunt. To solve this problem, isocitrate dehydrogenase (ICDH) was eliminated to increase the carbon flux of glyoxylate and thereby improve the glycolate titer. After engineering through the integration of gapC, combined with the inactivation of ICDH, SthA, and by-product pathways, as well as the upregulation of the two key enzymes isocitrate lyase (encoding by aceA), and glyoxylate reductase (encoding by ycdW), the glycolate titer increased to 5.3 g/L with a yield of 1.89 mol/mol glucose. Moreover, an optimized fed-batch fermentation reached a titer of 41 g/L with a yield of 1.87 mol/mol glucose after 60 h.

GHB related acids are useful in routine casework of suspected GHB intoxication cases.

GHB related acids (3,4-dihydroxy butyric acid, 2,4-dihydroxy butyric acid and glycolic acid) are produced through oxidative GHB metabolism. These analytes could be potential biomarkers to ensure the diagnosis of a GHB intoxication and even prolong the detection window. Within this study, forensic routine cases were measured to consider the potential of additional gas chromatographic mass spectrometric analysis on these acids. 17 GHB positive real cases (10 serum samples and 7 urine samples) and 40 cases with suspicion of drugging in DFC cases and negative GHB results (21 serum samples and 19 urine samples) were evaluated. Increased GHB related acid concentrations were detected in all serum and most urine samples positive on GHB. In some GHB negative cases, especially in serum samples, concentrations of GHB related acids gave hints that GHB actually was taken. We recommend to use the following cut-offs for a more reliable interpretation of potential GHB intoxication cases: 3,4-OH-BA:>3 mg/L in serum and>50 mg/L in urine; 2,4-OH-BA:>2 mg/L in serum and>25 mg/L in urine; GA:>5 mg/L in serum and>400 mg/L in urine.

A case of fatal intoxication by ingestion of an herbicide formulation containing fluroxypyr-meptyl and triclopyr.

Fluroxypyr-meptyl and triclopyr are synthetic auxin-like herbicides that are used to control woody and broadleaf weeds. Herein, we report a case of fatal intoxication involving fluroxypyr-meptyl and triclopyr. A 61-year-old man was found dead at his farm with several suicide notes, and a white plastic bottle and a plastic cup with traces of white emulsion were found next to him. The plastic bottle was labeled as an herbicide formulation containing fluroxypyr-meptyl and triclopyr. Forensic toxicological screening of the stomach contents revealed the presence of fluroxypyr-meptyl, fluroxypyr and triclopyr. However, no fluroxypyr-meptyl was detected in blood owing to its rapid hydrolysis to fluroxypyr. In this study, fluroxypyr and triclopyr in blood were extracted using solid-phase extraction, and analyzed by liquid chromatography-tandem mass spectrometry. The analytical method was validated in terms of linearity, precision, accuracy, recovery and matrix effect, and the acceptable criteria were satisfied. Toxicological analysis showed that fluroxypyr and triclopyr concentrations were 19.7 µg/mL and 137.4 µg/mL in peripheral blood and 16.5 µg/mL and 147.8 µg/mL in heart blood, respectively. Based on these toxicological results and autopsy findings, the cause of death was determined to be acute fatal intoxication by ingestion of the pesticide containing fluroxypyr-meptyl and triclopyr. This is the first report of the determination of fluroxypyr and triclopyr in a fatal intoxication case.

GHB related acids (dihydroxy butyric acids, glycolic acid) can help in the interpretation of post mortem GHB results.

Post mortem gamma hydroxy butyric acid (GHB) concentrations should be interpreted with caution since GHB concentrations can increase after death. Post mortem concentrations after the intake of GHB ante mortem do overlap with concentration ranges in cases without known exposure to GHB and make an interpretation challenging. GHB is known to undergo intensive metabolism to related acids (glycolic acid (GA), succinic acid (SA), 2,4- and 3,4-dihydroxy butyric acid (2,4-OH-BA and 3,4-OH-BA)). GHB and these related acids were analyzed using a validated gas chromatographic mass spectrometric (GC-MS) method after liquid liquid extraction and trimethylsilylation. SA concentrations were not usable post mortem due to instability. Concentrations in cases without known exposure to GHB (urine: n = 80; femoral blood: n = 103) were: for GA 4.6-121 mg/L in urine and 1.6-11.2 mg/L in blood, for 2,4-OH-BA < LoD-25,3 mg/L in urine and < LoD-3.7 mg/L in blood and for 3,4-OH-BA < LoD-54,3 mg/L in urine and < LoD-5.3 mg/L in blood. In death cases involving GHB (n = 11) concentrations of GHB related acids were increased compared to these levels (for GA in 7/10 cases and up to 391 mg/L in urine, in 6/11 cases and up to 34 mg/L in blood; for 2,4-OH-BA in 9/10 cases and up to 144 mg/L in urine, in 11/11 cases and up to 9.1 mg/L in blood; for 3,4-OH-BA in 7/10 cases and up to 665 mg/L in urine, in 11/11 cases and up to 19 mg/L in blood). Therefore, the concentrations of these GHB related acids can aid in a more reliable differentiation of GHB exposure in post mortem toxicology. We recommend to add the analysis of 2,4-OH-BA, 3,4-OH-BA and GA in femoral blood for the diagnosis of a GHB intake post mortem. Post mortem femoral blood concentrations > 4 mg/L for 2,4-OH-BA, > 5 mg/L for 3,4-OH-BA and > 12 mg/L for GA give hints for a GHB intake.

Use of Index-Based Screening Models to Evaluate the Leaching of Triclopyr and Fluroxypyr Through a Loam Soil Amended with Vermicompost.

The effect of vermicompost added to a loam soil on the leaching behaviour of two herbicides (triclopyr and fluroxypyr) was examined. Mobility of the herbicides was assessed using disturbed soil columns under laboratory conditions. In both cases, the addition of vermicompost significantly increased the sorption of the compounds. For both, DT50 values were slightly higher in the amended soil, due to the increased adsorption. Rate constants (k) calculated according to pseudo-first order model were significantly lower in the case of triclopyr (very persistent), which led to a much lower degradation rate compared to fluroxypyr (persistent) in both unamended and amended soils. Values calculated for the experimental leaching index (ELI) in unamended and amended soils showed medium and high leachability for fluroxypyr (0.31 and 0.29) and triclopyr (0.72 and 0.70), respectively. Other index-based screening models (GUS, RLPI, LIX) also catalogue both herbicides as potential leachers. Results confirm that triclopyr and fluroxypyr may contaminate groundwater resources.

HelixComplex snail mucus exhibits pro-survival, proliferative and pro-migration effects on mammalian fibroblasts.

Snail mucus is a mixture of active substances commonly thought to have healthy properties for the treatment of skin disorders. Although snail mucus is an ingredient of several cosmetic and para-pharmaceutic products, a comprehensive characterization of chemical composition and biological effects is still missing. Crude purified extracts from Helix aspersa muller mucus (HelixComplex) were prepared and, after chemical characterization, tested on in vitro experimental models. Differently from what expected, HelixComplex was characterized by the presence of small amounts of glycolic acid and allantoin. By using different in vitro assays on fibroblast cultures, we found that HelixComplex lacked of cytotoxicity, protected cells from apoptosis (p < 0.05) and, importantly, was able to significantly induce cell proliferation and migration through direct and indirect mechanisms. These effects were associated to morphological changes, cytoskeleton re-organization and release of cytokines. In conclusion, our findings suggest that snail mucus biological effects are attributable to cell proliferation and migration, and pave the way for further investigating snail mucus potential as therapeutic agent.

Analysis of diglycolic acid in food packaging, over the counter products, direct additive carboxymethyl cellulose, and retail foods.

Carboxymethyl cellulose (i.e. CMC or cellulose gum) is used as a direct additive for foods and drugs to change texture and act as a binder. CMC can also be a fluid absorbent used in food packaging and food contact materials. CMC and other carboxymethyl starches are synthesised by condensing glycolic acid with monochloroacetic acid. Diglycolic acid (DGA) is a byproduct produced by this condensation which cannot be completely removed. Currently, there are no analytical methods to accurately detect and quantify DGA in foods and food packaging materials. A method using a methanol/water extraction coupled with weak anion exchange solid phase extraction cleanup for more complex matrices was developed. A novel LC-MS/MS method was used to determine the DGA concentration in food contact materials, food grade direct additive CMC, and foods containing CMC. This paper will discuss the development of a new method for the preparation and cleanup of various food matrices and LC-MS/MS analysis for the presence of DGA.

N-doped carbon nanotubes-reinforced hollow fiber solid-phase microextraction coupled with high performance liquid chromatography for the determination of phytohormones in tomatoes.

A N-doped carbon nanotubes-reinforced hollow fiber solid-phase microextraction (N-doped CNTs-HF-SPME) method was developed for determination of two naphthalene-derived phytohormones, 1-naphthalene acetic acid (NAA) and 2-naphthoxyacetic acid (2-NOA), at trace levels in tomatoes. N-doped CNTs were dispersed in ultrapure water with the assistance of surfactant, and then immobilized into the pores of hollow fiber by capillary forces and sonification. The resultant N-doped CNTs-HF was wetted with 1-octanol, subsequently immersed into the tomato samples to extract the target analytes under a magnetic stirring, and then desorbed with methanol by sonication prior to chromatographic analysis. Compared with CNTs, the surface hydrophilicity of N-doped CNTs was improved owing to the doping of nitrogen atoms, and a uniform dispersion was formed, thus greatly simplifying the preparation process and reducing waste of materials. In addition, N-doped CNTs-HF exhibits a more effective extraction performance for NAA and 2-NOA on account of the introduction of Lewis-basic nitrogen. It is worth to mention that owing to the clean-up function of HF, there are not any complicated sample pretreatment procedures prior to the microextraction. To achieve the highest extraction efficiency, important microextraction parameters including the length and the concentration level of N-doped CNTs in surfactant solution, extraction time, desorption conditions such as the type and volume of solvents, pH value, stirring rate and volume of the donor phase were thoroughly investigated and optimized. Under the optimal conditions, the method showed 165- and 123-fold enrichment factors of NAA and 2-NOA, good inter-fiber repeatability and batch-to-batch reproducibility, good linearity with correlation coefficients higher than 0.9990, low limits of detection and quantification (at ng g-1 levels), and satisfactory recoveries in the range of 83.10-108.32% at three spiked levels. The proposed method taking advantages of both excellent adsorption performance of N-doped CNTs and the clean-up function of HF, was a simple, green, efficient and cost-effective enrichment procedure for the determination of trace NAA and 2-NOA in tomatoes.

Residue determination of triclopyr and aminopyralid in pastures and soil by gas chromatography-electron capture detector: Dissipation pattern under open field conditions.

In this study, a new method for the simultaneous quantitative determination of triclopyr and aminopyralid in forage grass, hay, and soil was developed and validated using gas chromatography coupled with electron capture detector (GC-ECD). In this method, a simple and maneuverable esterification reaction was applied to convert the two acidic herbicides into their ester form with methanol. The target compounds were extracted with 1% hydrochloric acid-acetonitrile, esterified, purified by florisil solid-phase extraction cartridge, and detected in a single run by the GC-ECD. The average recoveries using this method, at different fortified levels, ranged from 80% to 104% with intra-day and inter-day RSDs in the range of 1.2-10.8% and 3.3-10.3% for both the herbicides, respectively. The LODs were below 0.02 mg/kg while the LOQs were below 0.05 mg/kg, both of which were much lower than the maximum residue limits (MRLs) of 25-700 mg/kg in pastures, as established by the USA (the code of federal regulations). The open field dissipation and residual analysis in pastures and soil were conducted with the commercial formulation at two locations. With time, both triclopyr and aminopyralid dissipated via first-order kinetics. In forage grass, both compounds degraded rapidly over the first 14- or 21-d period and at a slow rate over the remainder of experimental days. In soil, they degraded at a relatively slow rate, and dissipated steadily to below or close to the LOQ by 60-d post application. The half-lives of triclopyr were 1.4-1.8 d and 6.2-9.0 d and aminopyralid were 1.7-2.1 d and 8.2-10.6 d in terms of forage grass and soil, respectively. The terminal residue results indicated that on 7 d after the treatment, the residues of aminopyralid and triclopyr in forage grass and hay were lower than the MRLs set by the USA. This work can provide guidance on the reasonable use of these herbicides and also provide an analytical method for the determination of triclopyr and aminopyralid in pasture and soil.

Runoff of the herbicides triclopyr and glufosinate ammonium from oil palm plantation soil.

This study focused on the residue detection of the herbicides triclopyr and glufosinate ammonium in the runoff losses from the Tasik Chini oil palm plantation area and the Tasik Chini Lake under natural rainfall conditions in the Malaysian tropical environment. Triclopyr and glufosinate ammonium are post-emergence herbicides. Both herbicides were foliar-sprayed on 0.5 ha of oil palm plantation plots, which were individualized by an uneven slope of 10-15%. Samples were collected at 1, 3, 7, 15, 30, 45, 60, 90, and 120 days after treatment. The concentrations of both herbicides quickly diminished from those in the analyzed sample by the time of collection. The highest residue levels found in the field surface leachate were 0.031 (single dosage, triclopyr), 0.041 (single dosage, glufosinate ammonium), 0.017 (double dosage, triclopyr), and 0.037 µg/kg (double dosage, glufosinate ammonium). The chromatographic peaks were observed at "0" day treatment (2 h after herbicide application). From the applied active ingredients, the triclopyr and glufosinate losses were 0.025 and 0.055%, respectively. The experimental results showed that both herbicides are less potent than other herbicides in polluting water systems because of their short persistence and strong adsorption onto soil clay particles.

Winery biomass waste degradation by sequential sonication and mixed fungal enzyme treatments.

To increase the efficiency of winery-derived biomass biodegradation, grape pomace was ultrasonicated for 20min in the presence of 0.25M, 0.5Mand1.0MKOH and 1.0MNaOH. This was followed by treatment with a 1:1 (v/v) mix of crude enzyme preparation derived from Phanerochaete chrysosporium and Trametes versicolor for 18h and a further 18h treatment with a 60:14:4:2 percent ratio combination of enzymes derived from Aspergillus niger: Penicillium chrysogenum: Trichoderma harzianum: P. citrinum, repsectively. Process efficiency was evaluated by its comparison to biological only mixed fungal degradation over 16days. Ultrasonication treatment with 0.5MKOH followed by mixed enzyme treatment yielded the highest lignin degradation of about 13%. Cellulase, ß-glucosidase, xylanase, laccase and lignin peroxidase activities of 77.9, 476, 5,390.5, 66.7 and 29,230.7U/mL, respectively, were observed during biomass degradation. Gas chromatography-mass spectrometry (GC-MS) analysis of the degraded material identified commercially important compounds such as gallic acid, lithocholic acid, glycolic acid and lactic acid which were generated in considerable quantities. Thus, the combination of sonication pre-treatment and enzymatic degradation has the potential to considerably improve the breakdown of agricultural biomass and produce commercially useful compounds in markedly less time (<40h) with respect to biological only degradation (16days).

Weather dependent dynamics of the herbicides florasulam, carfentrazone-ethyl, fluroxypyr-meptyl and fluroxypyr in wheat fields through field studies and computational simulation.

A dynamic model of dynamiCROP was applied to study environmental fate and behavior of four herbicides in wheat including florasulam, carfentrazone-ethyl, fluroxypyr-meptyl, and fluroxypyr. Meantime, their residue in wheat and dissipation half-lives in plant determined by field trials using QuEChERS liquid chromatography tandem mass spectrometry were used to verify modelling results. The combination of experimental verification and modelling prediction deciphered the fate of four pesticides in wheat field ecosystem. Besides, temperature difference of 3 °C only resulted in lower than 15% half-life difference. By quantifying the contribution of temperature, the predominant role of rain on pesticide dissipation was highlighted for the first time, namely higher precipitation leaded to faster degradation and vice versa.

Magnetic reduced graphene oxide functionalized with ß-cyclodextrin as magnetic solid-phase extraction adsorbents for the determination of phytohormones in tomatoes coupled with high performance liquid chromatography.

A ß-cyclodextrin (ß-CD) functionalized magnetic reduced graphene oxide composite (Fe3O4/RGO@ß-CD) has been prepared and its application as a selective adsorbent for the determination of the two naphthalene-derived phytohormones (1-naphthalene acetic acid (NAA) and 2-naphthoxyacetic acid (2-NOA)) has been investigated. Magnetic reduced graphene oxide composite (Fe3O4/RGO) was first synthesized via in situ chemical precipitation method and then ß-CD was applied to further functionalize the resultant Fe3O4/RGO composite. The as-prepared Fe3O4/RGO@ß-CD was characterized by Fourier transform infrared spectrometry (FT-IR), X-ray diffraction (XRD) and vibrating sample magnetometer (VSM). Compared with Fe3O4/RGO, the as-prepared Fe3O4/RGO@ß-CD showed better molecular selectivity and higher extraction efficiency for NAA and 2-NOA by dint of the size complementarity brought by the introduction of ß-CD. Response surface methodology (RSM), a multivariate experimental design technique, was used to optimize experimental parameters affecting the extraction efficiency in detail. Under the optimal conditions, good performance data was obtained. The calibration curves were linear over the concentration ranging from 2 to 600 ngg(-1) with correlation coefficients (R(2)) between 0.9995 and 0.9997 for all the analytes. The limits of detection (LODs) were 0.67 ngg(-1) for both NAA and 2-NOA. The intra- and inter-day relative standard deviations (RSDs) were less than 6.02% and 7.34%, respectively. The recoveries ranged from 91.45% to 95.89%. Taken together, the proposed method was an efficient pretreatment and enrichment procedure and could be successfully applied for selective extraction and determination of naphthalene-derived phytormones in complex matrices.

Comparison of chlorination and chloramination in carbonaceous and nitrogenous disinfection byproduct formation potentials with prolonged contact time.

Due to decreasing water demands in Japan, hydraulic retention times of water in piped supply systems has been extended, resulting in a longer contact time with disinfectants. However, the effects of extended contact time on the formation of various disinfection byproducts (DBPs), including carbonaceous DBPs such as trihalomethane (THM) and haloacetic acid (HAA), and nitrogenous DBPs such as nitrosodimethylamine (NDMA) and nitrosomorpholine (NMor), have not yet been investigated in detail. Herein, we compared the formation of these DBPs by chlorination and chloramination for five water samples collected from rivers and a dam in Japan, all of which represent municipal water supply sources. Water samples were treated by either filtration or a combination of coagulation and filtration. Treated samples were subjected to a DBP formation potential test by either chlorine or chloramine for contact times of 1 day or 4 days. Four THM species, nine HAA species, NDMA, and NMor were measured by GC-ECD or UPLC-MS/MS. Lifetime cancer risk was calculated based on the Integrated Risk Information System unit risk information. The experiment and analysis focused on (i) prolonged contact time from 1 day to 4 days, (ii) reduction efficiency by conventional treatment, (iii) correlations between DBP formation potentials and water quality parameters, and (iv) the contribution of each species to total risk. With an increased contact time from 1 day to 4 days, THM formation increased to 420% by chloramination. Coagulation-filtration treatment showed that brominated species in THMs are less likely to be reduced. With the highest unit risk among THM species, dibromochloromethane (DBCM) showed a high correlation with bromine, but not with organic matter parameters. NDMA contributed to lifetime cancer risk. The THM formation pathway should be revisited in terms of chloramination and bromine incorporation. It is also recommended to investigate nitrosamine formation potential by chloramination.

Solid-state UV-MALDI mass spectrometric quantitation of fluroxypyr and triclopyr in soil.

The work presented here refers firstly to solid-state UV-MALDI-Orbitrap-mass spectrometric analysis of fluroxypyr (A) and triclopyr (B) in soils under laboratory conditions. The experimental design has involved the following: (a) determination of analytes A and B in polycrystalline composites of organic materials 1-7, based on 2-piperidine (pyrrolidine or piperazine)-1-yl-ethyl ammonium salts in order to determine the effect of sample preparation techniques on method performance using commercial herbicide formulations and (b) analysis of non-(X j,k,l (i) ) and sterilized (Y j,k,l (i) ) soil samples (i-fold rate 1, 10, 100, or 1,000; j-pesticide type A or B; k-time (0, 5, 10, 20, and 50 days) and l = 1-3 replicated samples) having clay content ∈ 5.0-12.0 %, silt ∈ 23.0-51.1 %, sand ∈ 7.2-72.0 %, and pH ∈ 4.0-8.1. In order to obtain a high representativeness of the data toward real-field experiments, the pollution scheme has involved 1-, 10-, 100-, and 1,000-fold rates. The firstfold rate has concentration of pollutant A of 2.639 × 10(-4) g in 625 cm(2) soil horizon of 0-25 cm(2) (5 cm depth) according to registration report (PSM-Zulassungbericht) of German Federal Office of Consumer Protection and Food Safety (Bundesamt für Verbraucherschutz und Lebensmittelsicherheit) 6337/26.10.2009. The experimental design has involved quincunx systematic statistical approach for collection of soil samples. The performance has been compared with the corresponding statistical variable obtained, using an independent HPLC-ESI-(APCI-)-MS/MS analysis.

Tracking the processing of damaged DNA double-strand break ends by ligation-mediated PCR: increased persistence of 3'-phosphoglycolate termini in SCAN1 cells.

To track the processing of damaged DNA double-strand break (DSB) ends in vivo, a method was devised for quantitative measurement of 3'-phosphoglycolate (PG) termini on DSBs induced by the non-protein chromophore of neocarzinostatin (NCS-C) in the human Alu repeat. Following exposure of cells to NCS-C, DNA was isolated, and labile lesions were chemically stabilized. All 3'-phosphate and 3'-hydroxyl ends were enzymatically capped with dideoxy termini, whereas 3'-PG ends were rendered ligatable, linked to an anchor, and quantified by real-time Taqman polymerase chain reaction. Using this assay and variations thereof, 3'-PG and 3'-phosphate termini on 1-base 3' overhangs of NCS-C-induced DSBs were readily detected in DNA from the treated lymphoblastoid cells, and both were largely eliminated from cellular DNA within 1 h. However, the 3'-PG termini were processed more slowly than 3'-phosphate termini, and were more persistent in tyrosyl-DNA phosphodiesterase 1-mutant SCAN1 than in normal cells, suggesting a significant role for tyrosyl-DNA phosphodiesterase 1 in removing 3'-PG blocking groups for DSB repair. DSBs with 3'-hydroxyl termini, which are not directly induced by NCS-C, were formed rapidly in cells, and largely eliminated by further processing within 1 h, both in Alu repeats and in heterochromatic α-satellite DNA. Moreover, absence of DNA-PK in M059J cells appeared to accelerate resolution of 3'-PG ends.

Simultaneous determination of allantoin and glycolic acid in snail mucus and cosmetic creams with high performance liquid chromatography and ultraviolet detection.

A new methodology for simultaneous quantitative analysis of allantoin and glycolic acid in snail mucus and cosmetic creams was developed. HPLC separation was achieved a Synergi-Hydro RP column within 7min using isocratic elution with potassium phosphate (pH 2.7; 10mM) at a flow rate of 0.7mL/min at 30°C. Sample pretreatment was performed by dilution of mucus or cosmetic cream in the elution buffer, heating at 60°C for 20min, adjusting the pH to 2.9 and purification with hexane extraction. Linearity was determined with spiked samples and the LLOQ values of 0.0125 and 0.2500mg/mL were determined for allantoin and glycolic acid, respectively. Accuracy and intra- and inter-day repeatability were studied at three levels of concentrations (0.04, 0.08 and 0.16mg/mL for allantoin and 0.1, 1.5 and 4.0mg/mL for glycolic acid) using spiked mucus and cream base samples; mean values of recovery were in the range of 96.81-102.42% in all matrices tested, whereas the respective RSDs (%Relative Standard Deviation) were less than 3.04% in all cases. Spiked mucus and cream samples were stable (RSD<4.16 and relative error<4.34%) at room temperature and at 4°C for 1 week and at -18°C for 6 months; samples were also stable after three freeze-thaw cycles. The method was applied to the analysis of different lots of snail mucus, and of three commercial creams containing snail mucus.

Identification and analysis of an impurity inducing clinical adverse effect in anti-adhesion carboxymethyl chitosan products.

Controlling and minimizing the adverse effects of drugs are the key issues in ensuring the safety of drug therapy. Carboxymethyl chitosan has been widely used as an anti-adhesion material. However, recently in China there have been several reported instances of conjunctival hyperemia associated with the use of carboxymethyl chitosan containing products. Through MS, FTIR, and GC analysis, an impurity, diglycolic acid, was discovered in carboxylmethyl chitosan products. Pharmacological tests further indicated diglycolic acid has antithrombogenicity properties and induces vasodilation, both of which can cause conjunctival hyperemia. Thus, through these tests it was ascertained that diglycolic acid was the culprit responsible for the adverse clinical effects. Next, emphasis shifted to trying to discover the mechanism responsible for generating the diglycolic acid. Under strong basic conditions, chloroacetic acid can generate glycolic acid, which, upon etherification, can become diglycolic acid. In order to avoid future adverse effects, we have established an HPLC method to detect and determine diglycolic acid in carboxymethyl chitosan products. This method is specific, accurate, and precise, and can be easily implemented into routine monitoring practice. Concurrently, a refined method has also been established in order to eliminate diglycolic acid from carboxymethyl chitosan.

Simultaneous determination of 2-naphthoxyacetic acid and indole-3-acetic acid by first derivation synchronous fluorescence spectroscopy.

A simple, rapid, sensitive and selective method for simultaneously determining 2-naphthoxyacetic acid (BNOA) and Indole-3-Acetic Acid (IAA) in mixtures has been developed using derivation synchronous fluorescence spectroscopy based on their synchronous fluorescence. The synchronous fluorescence spectra were obtained with Δλ=100 nm in a pH 8.5 NaH2PO4-NaOH buffer solution, and the detected wavelengths of quantitative analysis were set at 239 nm for BNOA and 293 nm for IAA respectively. The over lapped fluorescence spectra were well separated by the synchronous derivative method. Under optimized conditions, the limits of detection (LOD) were 0.003 µg/mL for BNOA and 0.012 µg/mL for IAA. This method is simple and expeditious, and it has been successfully applied to the determination of 2-naphthoxyacetic acid and indole-3-acetic acid in fruit juice samples with satisfactory results. The samples were only filtrated through a 0.45 µm membrane filter, which was free from the tedious separation procedures. The obtaining recoveries were in the range of 83.88-87.43% for BNOA and 80.76-86.68% for IAA, and the relative standard deviations were all less than 5.0%. Statistical comparison of the results with high performance liquid chromatography Mass Spectrometry (HPLC-MS) method revealed good agreement and proved that there were no significant difference in the accuracy and precision between these two methods.