Deep Learning-Based Spectral Extraction for Improving the Performance of Surface-Enhanced Raman Spectroscopy Analysis on Multiplexed Identification and Quantitation.

Surface-enhanced Raman spectroscopy (SERS) has been recognized as a promising analytical technique for its capability of providing molecular fingerprint information and avoiding interference of water. Nevertheless, direct SERS detection of complicated samples without pretreatment to achieve the high-efficiency identification and quantitation in a multiplexed way is still a challenge. In this study, a novel spectral extraction neural network (SENN) model was proposed for synchronous SERS detection of each component in mixed solutions using a demonstration sample containing diquat dibromide (DDM), methyl viologen dichloride (MVD), and tetramethylthiuram disulfide (TMTD). A SERS spectra dataset including 3600 spectra of DDM, MVD, TMTD, and their mixtures was first constructed to train the SENN model. After the training step, the cosine similarity of the SENN model can achieve 0.999, 0.997, and 0.994 for DDM, MVD, and TMTD, respectively, which means that the spectra extracted from the mixture are highly consistent with those collected by the SERS experiment of the corresponding pure samples. Furthermore, a convolutional neural network model for quantitative analysis is combined with the SENN, which can simultaneously and rapidly realize the qualitative and quantitative SERS analysis of mixture solutions with lower than 8.8% relative standard deviation. The result demonstrates that the proposed strategy has great potential in improving SERS analysis in environmental monitoring, food safety, and so on.

Inhibition of the LOX enzyme family members with old and new ligands. Selectivity analysis revisited.

Lysyl oxidase (LOX) enzymes as potential drug targets maintain constant attention in the therapy of fibrosis, cancer and metastasis. In order to measure the inhibitory activity of small molecules on the LOX enzyme family members a fluorometric activity screening method was developed. During assay validation, previously reported non-selective small inhibitor molecules (BAPN, MCP-1, thiram, disulfiram) were investigated on all of the major LOX enzymes. We confirmed that MCP-1, thiram, disulfiram are in fact pan-inhibitors, while BAPN inhibits only LOX-like enzymes (preferably LOX-like-protein-2, LOXL2) in contrast to the previous reports. We measured the LOX inhibitory profile of a small targeted library generated by 2D ligand-based chemoinformatics methods. Ten hits (10.4% hit rate) were identified, and the compounds showed distinct activity profiles. Potential inhibitors were also identified for LOX-like-protein-3 (LOXL3) and LOX-like-protein-4 (LOXL4), that are considered as emerging drug targets in the therapy of melanoma and gastric cancer.

Gecko-Inspired Nanotentacle Surface-Enhanced Raman Spectroscopy Substrate for Sampling and Reliable Detection of Pesticide Residues in Fruits and Vegetables.

Rapid sampling and multicomponent detection are crucial for monitoring of pesticide residues analysis. Here, a gecko-inspired nanotentacle surface-enhanced Raman spectroscopy (G-SERS) platform is proposed for the first time for the simultaneous detection of three kinds of pesticides via a simple and intuitive "press and peeled-off" approach. The G-SERS platform obtained from seeding deposition of silver nanoparticles (Ag NPs) on 3D PDMS nanotentacle array is flexible and free-standing. Compared with other substrates, this G-SERS substrate can simultaneously provide outstanding SERS activity (enhancement factor = 1.2 × 107), superior reproducibility (RSD = 5.8%) and countless flexible nanoscale "tentacles" (∼6.7 × 108/cm2). Moreover, the high density of "tentacles" can freely approach the microarea and enable efficient target collection, which were confirmed by SEM and HPLC. By direct sampling from cucumber, apple, and grape surfaces, thiram (TMTD), methyl parathion (MPT), malachite green (MG), and their multiple components have been rapidly and reliably determined. For example, under the optimal conditions, a sensitivity of 1.6 ng/cm2 (S/N = 3) for TMTD was obtained on apple peels with a correlation coefficient (R) of 0.99. Therefore, the G-SERS substrate could offer a great practical potential for on-spot identification of various pesticide residues on real samples.

Mechanistic understanding of nanoparticle interactions to achieve highly-ordered arrays through self-assembly for sensitive surface-enhanced Raman scattering detection of trace thiram.

Over the last few decades, there is increasing worldwide concern over human health risks associated with extensive use of pesticides in agriculture. Developing excellent SERS substrate materials to achieve highly sensitive detection of pesticide residues in the food is very necessary owing to their serious threat to human health through food chains. Self-assembled metallic nanoparticles have been demonstrated to be excellent SERS substrate materials. Hence, alkanethiols-protected gold nanoparticles have been successfully prepared for forming larger-scale two-dimensional monolayer films. These films can be disassembled into a fluid state and re-assembled back to crystallized structure by controlling surface pressure. Further investigations reveal that their self-assembled structures are mainly dependent on the diameter of gold nanoparticles and ligand length. These results suggest that the size ratio of nanoparticle diameter/ligand length within the range of 4.45-2.35 facilitates the formation of highly ordered 2D arrays. Furthermore, these arrays present excellent Surface-Enhanced Raman Scattering performances in the detection of trace thiram, which can cause environmental toxicity to the soil, water, animals and result in severe damage to human health. Therefore, the current study provides an effective way for preparing monodispersed hydrophobic gold nanoparticles and forming highly ordered 2D close-packed SERS substrate materials via self-assembly to detect pesticide residues in food. We believe that, our research provides not only advanced SERS substrate materials for excellent detection performance of thiram in food, but also novel fundamental understandings of self-assembly, manipulation of nanoparticle interactions, and controllable synthesis.

Carbon dots-based stimuli-responsive hydrogel for in-situ detection of thiram on fruits and vegetables.

Stimuli-responsive hydrogel possesses a strong loading capacity to embed luminescent indicators for constructing food safety sensors, which are suitable for field application. In this work, a fluorescent hydrogel sensor was fabricated by incorporating Ag+-modified carbon dots (CDs-Ag+) into a sodium alginate (SA) hydrogel for in-situ detection of thiram. The fluorescence of CDs was quenched due to the combined effects of electrostatic adsorption and electron transfer between Ag+ and CDs. The formation of an AgS bond between thiram and Ag+ facilitates the release of CDs, causing subsequently fluorescence recovery. Combined with smartphone and analysis software, the fluorescence color change of the hydrogel sensor was converted into data information for quantitative detection of thiram. Such a sample-to-result step is completed within 10 min. Notably, the in-situ detection experiment of thiram in fruit and vegetable samples confirmed the practical application of the hydrogel sensor. Therefore, the hydrogel sensor provides a new research direction for the in-situ detection of pesticide residues in the monitoring of food safety.

Helical au nanostructure for SERS detection of hazardous molecular and chiral enantiomers.

In recent years, incidents of pesticide pollution and abuse of feed additives have occurred frequently, which pose a great threat to human health. Raman spectroscopy has become an important method in the field of food safety due to its rapidity, simplicity and sensitivity. It is important to obtain complex structure to promote surface-enhanced Raman scattering (SERS) effect. In this study, gold helical nanoparticles with rich surface structure were synthesized using cysteine as induce agent. Notably, the complex helical structure and tip led to an excellent electromagnetic enhancement property. The helical structure showed ultra-sensitive detection of hazardous molecular, such as thiram and ractopamine. Interestingly, the D/L-Au structure had significant chiral optical activity and could be used as an unlabeled SERS platform for enantiomer identification. This study provided an effective strategy for the detection of pesticides and feed additives, which could be applied in other aspects of food safety in the future.

Glutathione­iron hybrid nanozyme-based colorimetric sensor for specific and stable detection of thiram pesticide on fruit juices.

Given the potential dangers of thiram to food safety, constructing a facile sensor is significantly critical. Herein, we presented a colorimetric sensor based on glutathione­iron hybrid (GSH-Fe) nanozyme for specific and stable detection of thiram. The GSH-Fe nanozyme exhibits good peroxidase-mimicking activity with comparable Michaelis constant (Km = 0.551 mM) to the natural enzyme. Thiram pesticides can specifically limit the catalytic activity of GSH-Fe nanozyme via surface passivation, causing the change of colorimetric signal. It is worth mentioning that the platform was used to prepare a portable hydrogel kit for rapid qualitative monitoring of thiram. Coupling with an image-processing algorithm, the colorimetric image of the hydrogel reactor is converted into the data information for accurate quantification of thiram with a detection limit of 0.3 µg mL-1. The sensing system has good selectivity and high stability, with recovery rates in fruit juice samples ranging from 92.4% to 106.9%.

Identification of labile Zn sites in drug-target proteins.

Labile Zn fingers (Zfs) in proteins contain Zn-bound thiolates that can react with electrophilic agents, causing Zn(2+) ejection and protein unfolding. Such labile Zfs have been shown to be Cys4 or Cys3His cores whose Zn-bound Cys have no hydrogen bonds. Our aim here is to identify labile Zfs in proteins that are promising drug targets using these features. To prove the strategy used, we showed that five proteins with predicted labile Zfs reacted with Zn-ejecting agents, whereas five proteins with no or inert Zfs did not. The comprehensive set of labile Zfs provides new drug targets and guidelines to redesign Zn-ejecting compounds with improved specificity.

Pin1 inhibitors: Pitfalls, progress and cellular pharmacology.

Compelling data supports the hypothesis that Pin1 inhibitors will be useful for the therapy of cancer: Pin1 deficient mice resist the induction of breast cancers normally evoked by expression of MMTV-driven Ras or Erb2 alleles. While Pin1 poses challenges for drug discovery, several groups have identified potent antagonists by structure based drug design, significant progress has been made designing peptidic inhibitors and a number of natural products have been found that blockade Pin1, notably epigallocatchechin gallate (EGCG), a major flavonoid in green tea. Here we critically discuss the modes of action and likely specificity of these compounds, concluding that a suitable chemical biology tool for probing the function of Pin1 has yet to be found. We conclude by outlining some open questions regarding the target validation of Pin1 and the prospects for identification of improved inhibitors in the future.

Core-shell Au@ZIF-67-based pollutant monitoring of thiram and carbendazim pesticides.

A sensitive and stable substrate plays a vital role in the Raman spectroscopic techniques as an analytical method for detecting pesticides effectively from the environment. Enhancing signals from nanoparticles are weak and inconsistent in repeatability since analytes tend to degrade quickly under laser exposure. Herein, a novel substrate of Au@ZIF-67 is prepared on octahedral AuNPs by trapping pesticide molecules with small three-dimensional volumes by the flexibility of ZIF-67 for rapid detection with high sensitivity and stability. The two types of thiram and carbendazim pesticides, which are environmental pollutants that affect biodiversity, were successfully absorbed in Au@ZIF-67 nanostructures by adsorption-desorption equilibrium for analytical purposes in Raman spectroscopy. Spectra calculations of the thiram and carbendazim molecules on 8 atoms of Au using DFT were compared with the experimental data. The SERS enhancement factors for thiram and carbendazim were estimated to be 1.91 × 108 and 3.12 × 108, respectively, with the LOD values of trace amounts of ∼10-10 mol L-1. The novel substrate of Au@ZIF-67 is a propitious platform for detecting thiram and carbendazim in trace amounts, providing a helpful strategy for detecting residues with high performance in the environment at the laboratory and practical scales.

Two-Dimensional Printed AgNPs@Paper Swab for SERS Screening of Pesticide Residues on Apples and Pears.

In recent years, growing food safety and quality concerns have emerged and created an urgent need for the development of rapid and reliable food control technologies. This study proposes a novel surface-enhanced Raman spectroscopy (SERS) substrate printing technology that utilizes commercial filter paper functionalized by silver nanoparticles. We modified the Automatic TLC Sampler using a two-dimensional (2D) printer. The modification allows for various sampling modes which can be applied to 2D printing. The shape and size of nano silver on the substrate were determined, and the substrate sensitivity, uniformity, and stability were evaluated. As demonstrated by the experimental outcomes, the proposed technology is highly sensitive and reproducible, that is, the limit of quantitation was 10-5 mg/kg, and the spot-to-spot and block-to-block Raman intensity variations were below 4.2%. We also successfully applied the technology to pears and apples for thiram recognition, yielding outstanding detectability down to 2.5 × 10-6 and 3.9 × 10-7 mg/mL (equal to 2.5 × 10-3 and 3.9 × 10-4 mg/kg), respectively. These were well below the maximum residue limit (7 mg/kg). More importantly, the linear relationships between thiram levels and the SERS intensity allow for sensitive monitoring of minute variations in agricultural insecticide residues. This proposed detection method can realize in situ detection with a strong signal fingerprint.

Degradation of thiram in water, soil and plants: a study by high-performance liquid chromatography.

A comprehensive study was conducted to evaluate the persistence of thiram in water and soil under controlled conditions and on two plants, namely tomato and radish, in field conditions. In order to follow the decay of the pesticide, an HPLC procedure was developed employing an octadecyl endcapped RP-C18 column using a mixture of acetonitrile and water as the mobile phase and an ultraviolet detector. Studies conducted in water at different temperature, pH and organic content revealed that the persistence of the pesticide decreases with the increase in all the three variables. In the three different types of soils studied, the effect of pH was more or less apparent on a similar line. On average a slower decay was observed in the case of plants than in water and soil.

Chromosomal aberrations in cultured human lymphocytes treated with the fungicide, Thiram.

In vitro effects of different concentrations of Thiram were tested on human lymphocytes to determine, by means of the chromosome aberrations (CAs) assay, whether this fungicide could induce clastogenic damage. Evidences of the effect of Thiram on human lymphocytes were limited to sister chromatid exchange, micronuclei formation, and comet assays. We evaluated 0.01, 0.1, 1.2, and 12.0 µg/mL of Thiram, where 0.01 µg/mL represent the acceptable daily intake dose set by the World Health Organization and the Food and Agriculture Organization for fruit and vegetables, whereas 0.1, 1.2, and 12.0 µg/mL are its multiple values. Results indicated that human lymphocytes treated in vitro with Thiram at concentrations of 1.20 and 12.0 µg/mL significantly increased CAs frequency, compared with the negative control, whereas at lower concentrations (0.01 and 0.1 µg/mL), this effect was not observed. However, Thiram showed a clastogenic effect also at the concentration value of 1.2 µg/mL that represents a lower value with respect to the residue limits found in Italy for grapes, strawberries, potatoes, tobacco, and other fruits and vegetables. Finally, according to some evidence obtained from the study of other fungicides, Thiram produced a significant reduction in the mitotic index with increasing concentration.

Identification of degradation products of thiram in water, soil and plants using LC-MS technique.

In order to evaluate the deleterious effects of exposure to pesticides on a target population, a comprehensive study on their degradation in the various segments of ecosystem under varying environmental conditions is needed. In view of this, a study has been carried out on the metabolic pathways of thiram, a dithiocarbamate fungicide, in a variety of matrices namely water and soil under controlled conditions and plants in field conditions. The identification of degradation products was carried out in samples collected at various time points using LC-MS. The degradation products identified can be rationalized as originating by a variety of processes like hydrolysis, oxidation, N-dealkylation and cyclization. As a result of these processes the presence of some metabolites like dimethyl dithiocarbamate, bis(dimethyl carbamoyl) disulphide, bis(dimethyl dithiocarbamoyl) trisulphide and N-methyl-amino-dithiocarbamoyl sulphide was observed in all the cases. However, some different metabolites were observed with the change in the matrix or its characteristics such as cyclised products 2(N, N-dimethyl amino)thiazoline carboxylic acid and 2-thioxo-4-thiazolidine were observed only in plants. The investigations reflect that degradation initiates with hydrolysis, subsequently oxidation/dealkylation, followed by different types of reactions. The pathways seem to be complex and dependent on the matrices. Dimethyl dithiocarbamate and oxon metabolites, which are more toxic than parent compound, seem to persist for a longer time. Results indicate persistence vis-a-vis toxicity of pesticide and its metabolites and also provide a data bank of metabolites for forensic and epidemiological investigations.

Quantitative SERS sensing mediated by internal standard Raman signal from silica nanoparticles in flexible polymer matrix.

Attributed to poor signal uniformity and external interference, ultrasensitive surface-enhanced Raman spectroscopy (SERS) still faces difficulties in the reliable and quantitative detection of trace molecules. Here, a facile Ag/Si/sodium carboxy methyl cellulose (NaCMC) film with internal standard (IS) was promoted for quantitative determination of thiram. The effects of preparation conditions on SERS activity of the film were systematically investigated and then a flexible SERS substrate with high sensitivity and uniformity was fabricated. The enhancement factor was calculated to be 1.12 × 106 and SERS mapping was recorded with a relative standard deviation value of 19.8% by utilizing 4-mercaptobenzoic acid (4-MBA) as target molecule. Additionally, the dominant contribution of the IS from encapsulated Si nanoparticles (NPs) was confirmed in the quantitative assay of 4-MBA and thiram, facilitating attractive fitting coefficients (R2) as 0.991 and 0.998. Besides that, the proposed flexible film was conducted to scrub trace thiram from the surfaces of apple, orange, and cucumber, resulting in recoveries of 89%, 94%, and 91%. A smart and facile quantitative SERS substrate was developed here for monitoring trace biochemical molecules, verifying its potential utilizations in monitoring pesticide residues.

Sensitive and handy detection of pesticide residue on fruit surface based on single microsphere surface-enhanced Raman spectroscopy technique.

HYPOTHESIS: Surface-enhanced Raman spectroscopy (SERS) has become an emerging and reliable tool for detecting pesticide residues due to its high sensitivity, fast testing speed and easy sample handling. SERS active substrates are the key to achieve efficient and sensitive detection. However, for the most widely used noble metal nanoparticles, there are problems of high noble metal nanoparticle usage and random aggregation. The micron-scale Raman spot is focused on multiple randomly aggregated nanoparticles during the test, resulting in poor reproducibility. Therefore, the development of micron-scale cost-effective SERS substrates with good reproducibility and simple detecting method is of great significance in practical detection. EXPERIMENTS: Through deposition of silver nanoparticles (Ag-NPs) by chemical reduction on the surface of monodisperse sulfonated polystyrene (SPS) microspheres, micron-sized PS@Ag-NPs core-shell microspheres were prepared with excellent SERS activity. After that, two simple protocols (Method I and Method II) were explored for the determination of thiram on apple epidermis. FINDINGS: Based on our developed strategy of the single microsphere SERS technique, we successfully fabricated uniform PS@Ag-NPs substrate with high SERS activity and excellent detection sensitivity. The single microsphere SERS technique possesses the capability of anti-dilutability and the utilization of ultra-low PS@Ag-NPs microsphere dosage, realizing qualitative and quantitative detection of thiram on apple with detection limits far below the standard stipulated by China and the European Union.

Positive Patch Test Reactions to Carba Mix and Thiuram Mix: The North American Contact Dermatitis Group Experience (1994-2016).

BACKGROUND/OBJECTIVE: This study characterizes concomitant reactions to carba mix (CM) and thiuram mix (TM) in a large North American population. Because thiurams and dithiocarbamates have structural similarity, concomitant reactions are expected. METHODS: The 1994-2016 North American Contact Dermatitis Group data were analyzed. Patients with a final reaction interpreted as "allergic" to either CM or TM were included. RESULTS: A total of 49,758 patients were tested to both CM and TM. A total of 3437 (6.9%) had positive reactions to CM and/or TM including the following groups: CM+ only (n = 1403, 40.8%), TM+ only (n = 1068, 31.0%), or both (n = 966, 28.1%). A total of 47.5% of TM+ patients were positive to CM and 40.8% of CM+ patients were positive to TM. Male sex, occupationally related dermatitis, and hand involvement were significantly more common in individuals positive to CM and/or TM as compared with those who were negative (P < 0.0001). More than 80% of CM+/TM+ reactions were currently relevant. Gloves were the most common source of CM and TM; clothing and footwear were also frequent. CONCLUSIONS: Carba mix and TM remain important, clinically relevant allergens. Although significant concomitant reaction frequency was demonstrated, more than half of the patients reacting to either CM or TM would have been missed if both had not been tested, underscoring the importance of testing to both.

Dipentamethylene thiuram monosulfide is a novel inhibitor of Pin1.

Pin1 is involved in eukaryotic cell proliferation by changing the structure and function of phosphorylated proteins. PiB, the Pin1 specific inhibitor, blocks cancer cell proliferation. However, low solubility of PiB in DMSO has limited studies of its effectiveness. We screened for additional Pin1 inhibitors and identified the DMSO-soluble compound dipentamethylene thiuram monosulfide (DTM) that inhibits Pin1 activity with an EC50 value of 4.1 microM. Molecular modeling and enzyme kinetic analysis indicated that DTM competitively inhibits Pin1 activity, with a K(i) value of 0.05 microM. The K(D) value of DTM with Pin1 was determined to be 0.06 microM by SPR technology. Moreover, DTM specifically inhibited peptidyl-prolyl cis/trans isomerase activity in HeLa cells. FACS analysis showed that DTM induced G0 arrest of the HCT116 cells. Our results suggest that DTM has the potential to guide the development of novel antifungal and/or anticancer drugs.

Drug interactions with potential rubber closure extractables. Identification of thiol-disulfide exchange reaction products of captopril and thiurams.

Mixtures of thiuram disulfides are frequently used as accelerators in rubber stoppers for injectables and sterilized powders for injection. Rapid reactions of thiuram disulfides between themselves and with thiols yield mixed disulfides due to thiol-disulfide exchange. The possibility of exchange reactions of thiuram disulfides extracted from rubber stoppers and drug products containing pendant thiol groups have not been reported in the analysis of potential stopper extractables. In this paper we report the formation and identification of mixed thiuram disulfides of N,N,N',N'-dimethylthiuram disulfide (TMTD), N,N,N',N'-dibutylthiuram disulfide (TBTD), and captopril (a thiol-containing drug). A reversed-phase HPLC method was developed for the determination of TMTD, TBTD, captopril and their disulfides in aqueous vehicles, using a YMC ODS AQ column at 35 degrees C and mobile phases A and B consisting of acetonitrile:water:trifluoroacetic acid (TFA) (20:80:0.1) and acetonitrile:TFA (100:0.1), respectively. The captopril-TBTD and captopril-TMTD disulfides were identified by MS, with molecular ions at m/z 420.9 and m/z of 337.1, respectively. Possible structures for the fragment ions in the spectra are provided. Mixed captopril-thiuram formation was studied as a function of pH. Captopril-TMTD formation was enhanced at pH 6.0, reaching a maximum of 31.3% in 4.1h. At pH 4.0 and 2.2, the mixed captopril adduct product was still detected in solution after 20h. The impact of the formation of mixed disulfide products of thiol-containing drugs with thiurams in the HPLC profile of extractables and leachables studies is discussed.

A study towards release dynamics of thiram fungicide from starch-alginate beads to control environmental and health hazards.

In order to make the judicious use of thiram fungicide we have developed starch- and alginate-based controlled and sustained agrochemical delivery system in the form of beads using calcium chloride (CaCl2) as crosslinker. The beads were characterized by FTIR and swelling studies. To study the effect of composition of the beads on the release dynamics of fungicide (thiram), beads were prepared by varying the amount of starch, alginate and crosslinker in the beads. Formulation characteristics like entrapment efficiency, bead size, percentage equilibrium swelling of the beads and diffusion mechanism for thiram release have been evaluated. Maximum (93.33+/-2.89)% swelling and maximum (80.67+/-0.83)% thiram release has occurred in the beads prepared with 15% starch, 1% alginate and 0.1M crosslinker solution. In most of the formulations the entrapment efficacy of thiram has been observed more than 90% and the values for the diffusion exponent 'n' have been obtained >1 which shows that the release of fungicides occurred through Case II diffusion mechanism.