Ultrasensitive fluorescence detection of gonyautoxins in seawater using a novel molecularly imprinted nanoprobe.

Gonyautoxins (GTXs), a group of potent neurotoxins belonging to paralytic shellfish toxins (PSTs), are often associated with harmful algal blooms of toxic dinoflagellates in the sea and represent serious health and ecological concerns worldwide. In the study, a highly selective and sensitive fluorescence nanoprobe was constructed based on photoinduced electron transfer recognition mechanism to rapidly detect GTXs in seawater, using specific entrapment of molecularly imprinted polymers (MIPs) combined with fluorescence analyses. The green emissive fluorescein isothiocyanate was grafted in a silicate matrix as a signal transducer and fluorescence intensity of the nanoprobe with a core-shell structure exhibited a strong enhancement due to efficient analyte blockage in a short response time. Under optimal conditions, the developed MIPs nanoprobe presented an excellent analytical performance for spiked seawater samples including a recovery from 94.44 % to 98.23 %, a linear range between 0.018 nmol L-1 and 0.36 nmol L-1, as well as good accuracy. Furthermore, the method had extremely high sensitivity, with limit of detection obtained as 0.005 nmol L-1 for GTXs and GTX2/3. Finally, the nanoprobe was applied for the determination of GTXs in seven natural seawater samples with GTXs mixture (0.035-0.058 nmol L-1) or single GTX2/3 (0.033-0.050 nmol L-1), and the results agreed well with those of a UPLC-MS/MS method. The findings of our study suggest that the constructed MIPs-based fluorescence enhancement nanoprobe was suitable for rapid, selective and ultrasensitive detection of GTXs, particular GTX2/3, in natural seawater samples.

Selective capture and determination of doxycycline in marine sediments by using magnetic imprinting dispersive solid-phase extraction coupled with high performance liquid chromatography.

Antibiotics are frequently used in aquaculture as feed additives and finally enter the marine environment that can pose potential threat to humans. In this study, magnetic molecularly imprinted nanocomposites were prepared by surface imprinting and applied as selective sorbents for specific capture of doxycycline. A multivariate approach based on response surface methodology with Box-Behnken design was adopted to optimize the dispersive solid-phase extraction of doxycycline from marine sediment. Three key parameters, including adsorbent amount and type of washing/eluting solvent, were screened. Under optimum conditions, the limit of detection was 0.03 µg g-1 with good linearity from 0.5 to 20 µg g-1 followed by HPLC detection. Finally, two sediment samples were analysed and satisfactory recoveries between 90.60 % and 93.76 % were obtained with acceptable relative standard deviations (≤4.12 %), suggesting a promising applicability of the developed method for efficient extraction and sensitive quantification of antibiotics in complex marine environmental matrix.

Rising CO2 will increase toxicity of marine dinoflagellate Alexandrium minutum.

Ocean acidification caused by increasing emission of carbon dioxide (CO2) is expected to have profound impacts on marine ecological processes, including the formation and evolution of harmful algal blooms (HABs). We designed a set of experiments in the laboratory to examine the effects of increasing CO2 on the growth and toxicity of a toxic dinoflagellate Alexandrium minutum producing paralytic shellfish toxins (PSTs). It was found that high levels of CO2 (800 and 1200 ppm) significantly promoted the growth of A. minutum compared to the group (400 ppm) representing the current CO2 level. The total yields of PSTs by A. minutum, including both intracellular and extracellular toxins, were significantly enhanced, probably due to the induction of core enzyme activity and key amino acids synthesis for PST production. More interestingly, high level of CO2 promoted the transformation from gonyautoxin2&3 to gonyautoxin1&4 and depressed the release of PSTs from inside to outside of the cells. All these processes collectively led to an apparent increase of A. minutum toxicity. Our study demonstrated that rising CO2 would increase the risk of toxic A. minutum based on the comprehensive analyses of different processes including algal growth and toxin synthesis, transformation and release.

Release of florfenicol in seawater using chitosan-based molecularly imprinted microspheres as drug carriers.

Novel molecularly imprinted polymer (MIP) microspheres using functionalized chitosan as eco-friendly substrates were prepared by surface imprinting method and applied as drug delivery carriers to provide extended-release of florfenicol (FF) in seawater. The chitosan-based composites were characterized by scanning electron microscopy and Fourier transforms infrared spectroscopy analyses. The swelling behavior, adsorption capability, and selectivity for FF were investigated. The results show that the MIPs possessed high drug loading saturation capacity and specific recognition affinity for FF. The release studies of MIPs as drug delivery carriers were evaluated in natural seawater. The microspheres exhibited slow sustained release profiles of FF and the release behavior conformed to the first-order kinetic equation. The imprinted microspheres as drug delivery devices would be a promising application for improving the efficacy of the antibiotic without exposing the ecological system to excess FF in aquaculture.

[Application of novel quantum dot-based molecularly imprinted fluorescence sensor in rapid detection].

A critical need in analytical chemistry is the efficient fabrication of selective and sensitive sensors to detect trace analytes in complicated samples. In recent years, fluorescence analysis has been widely used in environmental research and the life sciences due to its high sensitivity and simple operation. Quantum dots (QDs) are a new type of fluorescent nanomaterials. Owing to the quantum confinement effect, QDs possess excellent optical properties such as strong anti-bleaching ability, a narrow excitation and emission band, and tunable emission wavelength. As a hot labeling material, QDs are suitable for use in surface-modified analytical sensors employed in fields such as analytical chemistry, biology, and medicine. However, QD materials have a notable disadvantage, in that the actual sample matrix may contain some interferents with luminescent responses similar to those of the target; this decreases the selective ability of the fluorescence sensor. The surface modification of QDs via the molecular imprinting technique (MIT) is a promising solution to overcome this drawback. Molecularly imprinted polymers (MIPs) are a kind of "bionic" material that can carry out specific recognition and selective adsorption and hence, possess the unique properties of recognition specificity, structural predictability, good reproducibility, and excellent stability. Accordingly, MIPs have been widely employed in sensors as well as for drug delivery, catalysis, and solid phase extraction. Notably, QD-based molecularly imprinted fluorescence sensors combine the advantages of QDs and the MIT. Owing to their specific selectivity and high sensitivity, such sensors have been extensively developed for environmental monitoring, food detection, and biological analysis. However, there remain challenges associated with the preparation and application of the sensors: (i) single recognition: it is important to develop a composite sensor that can detect multiple target analytes from the actual samples at the same time during practical application; (ii) poor hydrophilicity: the actual sample is usually a liquid matrix; hence, it is imperative to determine an approach for improving the hydrophilicity of the sensor; (iii) the accuracy of fluorescence response and the resolution of visual detection need to be further improved; (iv) imprinting: it remains challenging to imprint biological macromolecules, viruses, and bacteria. Thus far, many researchers have made progress with regard to the preparation and application of the sensors. Accordingly, this work reviews approximately 20 papers published by the American Chemical Society, Elsevier, and other databases in the last five years to highlight progress in novel preparation methods and practical applications of QD-based molecularly imprinted fluorescence sensors for the sensitive analysis and rapid detection of trace substances. First, according to the different numbers of emission peaks in the fluorescence spectrum, three kinds of QD-based molecularly imprinted fluorescence sensors are introduced and the related recognition mechanisms are explained. Second, according to the different substances to be detected, this mini-review summarizes the latest research progress in sensors for the detection of ions, organic small molecules, biological macromolecules, as well as for the analysis of bacteria and viruses. Finally, existing challenges associated with the preparation and application of the sensors, as well as future development trends, are discussed.

Review on molecular imprinting technology and its application in pre-treatment and detection of marine organic pollutants.

Molecular imprinting technology (MIT) has been considered as an attractive method to produce artificial receptors with the memory of size, shape and functional groups of the templates and has become an emerging technique with the potential in various fields due to recognitive specificity, high efficient selectivity and mechanical stability, which can effectively remove background interference and is suitable for the pre-treatment and analysis of trace level substances in complex matrix samples. Nearly 100 papers about the application of MIT in the detection of marine pollutants were found through Science Citation Index Expanded (SCIE). On this basis, combined with the application of MIT in other fields, the pre-treatment process of marine environmental samples was summarized and the potential of four types of different molecularly imprinted materials in the pre-treatment and detection of marine organic pollutants (including antibiotics, triazines, organic dyes, hormones and shellfish toxins) samples was evaluated, which provides the innovative configurations and progressive applications for the analysis of marine samples, and also highlights future trends and perspectives in the emerging research field.

Fluorescence assay of oxytetracycline in seawater after selective capture using magnetic molecularly imprinted nanoparticles.

A comprehensive strategy for manufacturing a novel sorbent based on magnetic molecularly imprinted polymers (MMIPs) is addressed for selective capture of oxytetracycline from seawater. The novel MMIPs were synthesized by nano-Fe3O4 as sacrificial matrix and adsorption properties of the polymers demonstrate rapid adsorption kinetics, high adsorption capacity, and specificity towards oxytetracycline provided by the core-shell composite structure. After screening the critical parameters by multivariate optimization, a magnetic imprinting solid phase extraction method combined with fluorescence spectrophotometry (MMIP-SPE-FL) was constructed for sensitive determination of oxytetracycline in seawater samples. The results show a good linear response dependence on the spiking concentration of 3-100 µg L-1, and a satisfactory limit of detection of 0.7 µg L-1 after the MMIP-SPE preconcentration. Seven seawater samples from Jiaozhou bay were analyzed to give recoveries in the range of 89.75-107.65% with relative standard deviation values of less than 5.44% (n = 3).

First determination of extracellular paralytic shellfish poisoning toxins in the culture medium of toxigenic dinoflagellates by HILIC-HRMS.

Paralytic shellfish poisoning (PSP) toxins have received considerable attention in recent years because of their adverse effects on marine breeding industries and human health. In this study, a reliable method for the analysis of extracellular PSP toxins in the culture medium of marine toxic dinoflagellates was developed for the first time using graphitized carbon black-solid-phase extraction and hydrophilic interaction liquid chromatography-high-resolution mass spectrometry. The limit of quantification of typical PSP toxins in algal culture medium ranged from 0.072 µg/L to 0.151 µg/L under optimal conditions. Satisfactory absolute recoveries (87.5%-102.4%), precision (relative standard deviation ≤ 7.6%), and linearity (R2 ≥ 0.9998) were also achieved. In addition, the proposed method was applied to screen and determine the extracellular PSP toxins of two typical toxigenic dinoflagellates, Alexandrium minutum and Alexandrium tamarense. The total concentrations of the extracellular PSP toxins in A. minutum and A. tamarense over the whole growth period were within 2.0-735.5 and 2.0-19.2 µg/L, respectively. The concentrations of extracellular PSP toxins varied remarkably in the different growth stages of A. minutum and A. tamarense, and the contents of some extracellular PSP toxins were substantially higher than those of intracellular PSP toxins. Therefore, the extracellular PSP toxins released by toxigenic red tide algae cannot be ignored, and their environmental fate, bioavailability, and potential harm to aquatic environment need to be investigated in future studies.

Direct extraction and detection of malachite green from marine sediments by magnetic nano-sized imprinted polymer coupled with spectrophotometric analysis.

This research describes the application of magnetic molecularly imprinted nano-sized polymers (MMIPs) for the selective extraction and fast detection of malachite green (MG) from marine sediment samples followed by UV-Vis spectrophotometry. The novel material was prepared by surface imprinting using methacrylic acid as the functional monomer for fixing the template molecules. The polymers obtained at each step were thoroughly studied by transmission electron microscopy, FTIR spectroscopy and thermogravimetric analysis. Simultaneously, the adsorption performances of the resulting nanoparticles were analysed in detail and an excellent affinity with the MG was revealed. Further, the main parameters of magnetic molecular imprinted solid-phase extraction (MMIP-SPE) were screened via multivariate optimization methods. The magnetic nanoparticles were used as special adsorbents to directly extract MG from crude marine sediment extracts. The developed method exhibits satisfactory recoveries from the spiked samples, ranging from 80.40 to 92.96% with an RSD of less than 5.18% (n = 3).

Selective extraction and detection of norfloxacin from marine sediment and seawater samples using molecularly imprinted silica sorbents coupled with HPLC.

A simple, sensitive and effective method was developed and validated for selective adsorption and quantitation of norfloxacin (NFX) from marine sediments and seawater samples using the novel molecularly imprinted silica polymers as sorbents followed by high-performance liquid chromatographic analysis with diode array detection (HPLC-DAD). The imprinted particles were characterized by different techniques and the molecular recognition for NFX was also investigated. Several parameters affecting the extraction efficiency of molecularly imprinted solid-phase extraction (MISPE) process were optimized. Three sediment and seawater samples from Weihai bay in China were analyzed and the results show that satisfactory recoveries (77.2-98.7%) were achieved in seawater samples with low relative standard deviation (RSD) values less than 6.89% (n = 3), and the recoveries in sediments were in the range of 75.5-91.7%. The limits of detection are respectivley 2 µg L-1 in seawater samples and 5 µg kg-1 in sediments.

Fast extraction of chloramphenicol from marine sediments by using magnetic molecularly imprinted nanoparticles.

A straightforward method has been developed for selective separation of chloramphenicol (CAP) from marine sediment samples. Magnetic molecularly imprinted nanoparticles (NPs) of type Fe3O4@SiO2 were prepared via surface imprinting with CAP. The NPs were characterized by Fourier transform infrared spectroscopy, transmission electron microscopy and thermogravimetric analysis. They have perfect core-shell structure, excellent thermal stability, high affinity and selectivity to CAP. The imprinting factor and Scatchard analysis also reveal good specific recognition to the template. The imprinted NPs were applied as sorbents for fast and selective extraction of CAP from marine sediment samples. The experimental parameters affecting separation efficiency were optimized. Three marine sediment samples were analyzed. Following desorption with methanol/water (90/10,v/v), CAP was quantified by HPLC with DAD detection. The limit of detection is 0.1 µg L-1 with a good linear response between 0.1-20 mg L-1 of CAP concentration (R2 = 0.999, n = 3). The method exhibits satisfactory recoveries from spiked samples (77.9-102.5%) and has low relative standard deviations (<6.3%). The magnetic material can be used at least 5 times by the regeneration without any loss of selectivity and adsorption capability. Graphical abstract Schematic presentation of magnetic molecularly imprinted nanoparticles (MMIPs) as sorbent for fast extraction and chromatographic analysis of chloramphenicol (CAP) from marine sediments. CAP-MMIPs are synthesized by surface imprinting method using 3-methacryloxy propyl trimethoxy silane (MPS) as the silane coupling agent.

Selective separation and purification of ß-estradiol from marine sediment using an optimized core-shell molecularly imprinted polymer.

The core-shell molecularly imprinted polymers were optimized to provide reliable connections that allow molecularly imprinted polymers to be fixed on SiO2 surface for the efficient separation and purification of ß-estradiol from marine sediment for the first time. To achieve the goal, different preparation methods were used and finally the polymer using 3-methacryloxypropyltrimethoxysilane as coupling agent exhibited the best result, which further confirmed that 3-methacryloxypropyltrimethoxysilane played an indispensable role on improving the inter-particle connections. An offline molecularly imprinted solid-phase extraction with high-performance liquid chromatography method was successfully applied to the isolation and enrichment of ß-estradiol from marine sediment samples with high adsorption capacity, excellent clean-up efficiency, and great enrichment effect as well as high recovery (>90%) and accuracy (RSD < 8.5%, n = 3). It proved the successful grafting of molecularly imprinted polymers on SiO2 surface and the applicability of the offline molecularly imprinted solid-phase extraction method in the selective extraction and enrichment of ß-estradiol from marine sediment.

Selective detection of chloramphenicol based on molecularly imprinted solid-phase extraction in seawater from Jiaozhou Bay, China.

This study highlights an efficient sample pre-treatment method for preconcentration and detection of chloramphenicol in marine water using molecularly imprinted solid-phase extraction (MISPE). Chloramphenicol molecularly imprinted microspheres were prepared and evaluated on the base of morphology, capacity and selectivity. The imprinted microspheres exhibited specific recognition and high retention capability to chloramphenicol and were applied as special solid-phase extraction adsorbents. An off-line MISPE protocol has been optimized and a creative analytical method coupled to HPLC-DAD was successfully developed for the cleanup and determination of chloramphenicol in seawater samples. Method performance was satisfactory with recoveries ranging from 81 to 90% and relative standard deviation (RSD) was <4.93% (n = 3). Accuracy of the method was assessed at three spiking concentration levels and the limit of detection was 5 ng L-1. Finally, five seawater samples from Jiaozhou Bay of China were determined and the results showed that there was no chloramphenicol detected.

Selective isolation of gonyautoxins 1,4 from the dinoflagellate Alexandrium minutum based on molecularly imprinted solid-phase extraction.

Gonyautoxins 1,4 (GTX1,4) from Alexandrium minutum samples were isolated selectively and recognized specifically by an innovative and effective extraction procedure based on molecular imprinting technology. Novel molecularly imprinted polymer microspheres (MIPMs) were prepared by double-templated imprinting strategy using caffeine and pentoxifylline as dummy templates. The synthesized polymers displayed good affinity to GTX1,4 and were applied as sorbents. Further, an off-line molecularly imprinted solid-phase extraction (MISPE) protocol was optimized and an effective approach based on the MISPE coupled with HPLC-FLD was developed for selective isolation of GTX1,4 from the cultured A. minutum samples. The separation method showed good extraction efficiency (73.2-81.5%) for GTX1,4 and efficient removal of interferences matrices was also achieved after the MISPE process for the microalgal samples. The outcome demonstrated the superiority and great potential of the MISPE procedure for direct separation of GTX1,4 from marine microalgal extracts.

Preparation and characterization of magnetic molecularly imprinted polymers for selective trace extraction of dienestrol in seawater.

Highly selective and efficient magnetic molecularly imprinted polymers (MMIPs) were prepared using Fe3O4@SiO2 as a magnetic supporter, 3-methacryloxypropyltrimethoxy-silane (MPS) as a silane coupling agent, DIS as a template, methacrylic acid (MAA) as a functional monomer and ethyleneglycol dimethacrylate (EGDMA) as a cross-linker for the extraction of trace residuals of the synthetic estrogen dienestrol (DIS) in seawater, which is a concern worldwide for its endocrine disruption and carcinogenic danger to human health. The obtained MMIPs were demonstrated to have spherical morphologies, core-shell structures, large binding capacities, high efficiency and selectivity. These were characterized by scanning electron microscopy (SEM), transmission electron microscopy (TEM) and adsorption experiments. Owing to the specific binding sites, the MMIPs exhibited an almost three times higher adsorption capacity towards DIS (Qmax=4.68mgg-1) than magnetic molecularly non-imprinted polymers (MNIPs) (Qmax=1.72mgg-1). DIS in spiked seawater samples from the Weihai Bay of China was extracted and enriched by MMIPs, and satisfactory recoveries (87.3%-96.4%) with low relative standard deviation (RSD) values (2.03%-5.18%, n=5) were obtained. The limit of detection (LOD) of the method obtained was 0.16µgL-1, and the limit of quantitation was 0.52µgL-1 after MMIPs. No significant deterioration of the adsorption capacity of the MMIPs was observed after six rounds of regeneration. The results further demonstrated the applicability of the MMIPs method, a simple and straightforward method for the extraction and enrichment of DIS in seawater without any time-consuming procedures.

Microencapsulation of norfloxacin in chitosan/chitosan oligosaccharides and its application in shrimp culture.

Norfloxacin chitosan/chitosan oligosaccharide microcapsules (NCCM) were prepared by emulsion-chemical crosslinking method. The characteristics of obtained microcapsules were evaluated by scanning electron microscopy, Fourier transform infrared spectroscopy and release experiments. Cumulative release profile of norfloxacin from the chitosan microcapsules in natural seawater was measured and the controlled release of drugs was at a uniform rate in 48h. The chitosan microcapsules were applied onto the antibacterial study of the shrimp culture in natural seawater. It is observed that the seawater in the NCCM added groups was relatively clear and the biomass of Vibrio increased slowly in contrast to the control and norfloxacin groups. The inhibition rate of Vibrio in norfloxacin groups obvioursly decreased after the 5th day, whereas, it remained high and stable during experiment period in NCCM groups. The results showed that the chitosan microcapsules as release materials have excellent antibacterial effects on Vibrio in the farming of Penaeus vannamei Boone. The controlled release could obviously reduce dosage of antibiotics and delivery times, and effectively improve the utilization rate of norfloxacin drugs for shrimps.

Determination of ciprofloxacin in Jiaozhou Bay using molecularly imprinted solid-phase extraction followed by high-performance liquid chromatography with fluorescence detection.

A high selective pre-treatment method for the cleanup and preconcentration of ciprofloxacin in natural seawater samples was developed based on molecularly imprinted solid-phase extraction (MISPE). The ciprofloxacin imprinted polymers were synthesized and the characteristics of obtained polymers were evaluated by scanning electron microscopy, Fourier transform infrared spectroscopy and binding experiments. The imprinted materials showed high adsorption ability for ciprofloxacin and were applied as special solid-phase extraction sorbents for selective separation of ciprofloxacin. An off-line MISPE procedure was optimized and the developed MISPE method allowed direct purification and enrichment of the ciprofloxacin from the aqueous samples prior to high-performance liquid chromatography analysis. The recoveries of spiked seawater on the MISPE cartridges ranged from 75.2 to 112.4% and the relative standard deviations were less than 4.46%. Five seawater samples from Jiaozhou Bay were analyzed and ciprofloxacin was detected in two samples with the concentrations of 0.24 and 0.38µgL(-1), respectively.

Experimental and computational studies on molecularly imprinted solid-phase extraction for gonyautoxins 2,3 from dinoflagellate Alexandrium minutum.

An innovative and effective extraction procedure based on molecularly imprinted solid-phase extraction (MISPE) was developed for the isolation of gonyautoxins 2,3 (GTX2,3) from Alexandrium minutum sample. Molecularly imprinted polymer microspheres were prepared by suspension polymerization and and were employed as sorbents for the solid-phase extraction of GTX2,3. An off-line MISPE protocol was optimized. Subsequently, the extract samples from A. minutum were analyzed. The results showed that the interference matrices in the extract were obviously cleaned up by MISPE procedures. This outcome enabled the direct extraction of GTX2,3 in A. minutum samples with extraction efficiency as high as 83 %, rather significantly, without any need for a cleanup step prior to the extraction. Furthermore, computational approach also provided direct evidences of the high selective isolation of GTX2,3 from the microalgal extracts.

Determination of diethylstilbestrol in seawater by molecularly imprinted solid-phase extraction coupled with high-performance liquid chromatography.

An effective and highly selective molecularly imprinted material was prepared by suspension polymerization for the isolation and pre-concentration of synthetic estrogen diethylstilbestrol (DES) in seawater. The obtained MIPMs were proved to have more uniform size and porous structure, with maximum adsorption capacity of 8.43 mg g(-1) almost two times more than NIPMs (4.43 mg g(-1)). The MIPMs showed no significant deterioration of the adsorption capacity after five rounds of regeneration. An off-line molecularly imprinted solid phase extraction (MISPE) method followed by HPLC-DAD was proposed for the detection of DES in seawater, and recoveries were satisfactorily higher than 77%. Four seawater samples in aquaculture area were analyzed and 0.61 ng mL(-1) DES was detected in one sample. The result demonstrated that this method can be used for the rapid separation and clean up of trace residual of DES in seawater.

Determination of melamine in aquaculture feed samples based on molecularly imprinted solid-phase extraction.

This research highlights the application of highly efficient molecularly imprinted solid-phase extraction for the preconcentration and analysis of melamine in aquaculture feed samples. Melamine-imprinted polymers were synthesized employing methacrylic acid and ethylene glycol dimethacrylate as functional monomer and cross-linker, respectively. The characteristics of obtained polymers were evaluated by scanning electron microscopy, Fourier transform infrared spectroscopy and binding experiments. The imprinted polymers showed an excellent adsorption ability for melamine and were applied as special solid-phase extraction sorbents for the selective cleanup of melamine. An off-line molecularly imprinted solid-phase extraction procedure was developed for the separation and enrichment of melamine from aquaculture feed samples prior to high-performance liquid chromatography analysis. Optimum molecularly imprinted solid-phase extraction conditions led to recoveries of the target in spiked feed samples in the range 84.6-96.6% and the relative standard deviation less than 3.38% (n = 3). The aquaculture feed sample was determined, and there was no melamine found. The results showed that the molecularly imprinted solid-phase extraction protocols permitted the sensitive, uncomplicated and inexpensive separation and pre-treatment of melamine in aquaculture feed samples.