Ochratoxin A (OTA) is one of the major mycotoxins causing severe effects on the health of humans and animals. Ochratoxin alpha (OTα) is a metabolite of OTA, which is produced through microbial or enzymatic hydrolysis, and one of the preferred routes of OTA detoxification. The methods described here are applicable for the extraction and quantification of OTA and OTα in several pig and poultry matrices such as feed, feces/excreta, urine, plasma, dried blood spots, and tissue samples such as liver, kidney, muscle, skin, and fat. The samples are homogenized and extracted. Extraction is either based on a stepwise extraction using ethyl acetate/sodium hydrogencarbonate/ethyl acetate or an acetonitrile/water mixture. Quantitative analysis is based on reversed-phase liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). Method validation was successfully performed and the linearity, limit of quantification, accuracy, precision as well as the stability of the samples, were evaluated. The analyte recovery of the spiked samples was between 80 and 120% (80–150% for spiked concentrations ≤ 1 ng/g or ng/mL) and the relative standard deviation was ≤ 15%. Therefore, we provide a toolbox for the extraction and quantification of OTA and OTα in all relevant pig and poultry matrices. Aspergillus ochraceus [Aspergillus and Penicillum species [Mycotoxins are toxic secondary metabolites produced by filamentous fungi contaminating food and animal feed . One of chraceus , but lat species . OTA is  species . There a species ,7,8,9,10 species ,12.There is a broad range of exposure routes to OTA. Different types of food and feed can be contaminated, including cereals like wheat, rice, rye, maize, and barley . TherefoIngested OTA can easily be absorbed and further distributed to different organs. Nevertheless, natural detoxification into less-/nontoxic metabolites, such as ochratoxin alpha (OTα) and phenylalanine, is possible by enzymes and microbes in the gut ,18,19. TIn human and animal matrices many methods for the detection of OTA are either based on liquid extraction from tissue or feed ,24,25,26v/v) mixture worked best. Regardless of the matrix extracted, the analyte concentration was determined by reversed phase LC-MS/MS using external calibration in neat solvent. Possible extraction losses and matrix effects during ionization were compensated using 13C-labelled OTA and OTα as internal standards, which were added to the samples prior to extraction. The aim of this work was to develop simple methods for the extraction and quantification of OTA and OTα in different pig and chicken matrices. The matrices of interest were feces or excreta, urine (only available from pig), plasma, DBS, skin and fat (only from chicken), liver, kidney, muscle, and feed for pigs and poultry. For the method development and validation, blank matrices were homogenized and spiked with OTA and OTα at different levels, covering the full linear range of quantification for each different matrix. Afterwards, the samples were extracted using ethyl acetate followed by re-extraction in a reduced amount of sodium hydrogencarbonate solution, and another re-extraction in a reduced amount of ethyl acetate. A similar stepwise liquid/liquid extraction was already described by Monaci et al.  or GiacoTo determine if the method is fit for its intended purpose, a single-laboratory method validation was carried out, and the linearity of the calibration function, intraday and interday precision, accuracy, LOQ, LOD, specificity, and analyte stability in processed samples were evaluated. 13C-labelled OTA and OTAα) was applied for the linear regression with a weighting factor of 1/analyte concentration, as recommended by Gu et al. [2) was > 0.99.The linearity of the external calibration for OTA and OTα was assessed by preparing four individual calibration series from seven individual concentration levels between 0.25 ng/mL and 250 ng/mL in neat solvent . For theu et al. . The cal13C-labelled internal standard correction for OTA and OTα was used for all matrices except for DBS of pigs. DBS of pigs showed already without internal standard correction analyte recoveries between 88–116%, and therefore internal standard correction was not necessary. Accuracy and precision were determined in all matrices using blank samples spiked with OTA and OTα. Samples with different spiking levels were extracted on two independent days to calculate the interday precision. The intraday precision was calculated from an independent triplicate prepared on the same day. The intraday precision  and interday precision  showed a relative standard deviation (RSD) ≤ 15% for all matrices. For all different matrices evaluated, the accuracy of the method was between 81% and 147%. Analyte recovery of over 120% was determined only for low-level concentrations at the LOQ for poultry plasma samples, and liver, kidney, and muscle samples from pigs. Therefore, the set acceptance criteria for the analyte recovery of 80–120% (80–150% for spiked concentrations ≤ 1 ng/g or ng/mL) of the spiked concentration were met. To fulfill all validation criteria according to accuracy and precision, The LOQ was estimated by spiking experiments. It is the smallest measured content of the analyte above which the determination can be made with a specified accuracy and precision, as recommended by the Guidance for Industry 208 from the Food and Drug Administration . The LODThe specificity of the method is given by the retention time and the multiple reaction monitoring (MRM) transitions. Blank samples were extracted and measured to evaluate the endogenous response. No interfering peaks were detected at the specific retention time of OTA at 1.6 min and OTα at 1.3 min, except for pig plasma and pig DBS samples. In these samples, a small peak at the specific retention time of OTA was found, with an approximate S/N of LOD. As this peak was visible in all measured MRM transitions for OTA, we assume that this is not an interfering peak, but rather the analyte itself. We considered that the used plasma and blood samples were not completely OTA-free.As analyte stability in processed samples is crucial for reliable results, the analyte stability of OTA and OTα in ready-to-inject samples was evaluated at different temperatures and storage periods. The analytes were stable in all evaluated matrices. At room temperature after 7 days of storage, slightly higher recoveries  were observed in poultry matrices. All other tested storage conditions fulfilled the acceptance criteria of an analyte recovery of 80–120% compared to the initial measured value. Nevertheless, for long-term storage of processed samples, storage at −20 °C would be recommended.The intention of this work was to develop and validate methods for extraction and quantification of OTA and OTα in different pig and poultry matrices. The European Commission EC/2016/1319) has published recommendations for OTA in feed, with maximum thresholds of 50 µg/kg for pig feed and 100 µg/kg for poultry feed /1319 has,34. We pThere are already many methods described for the detection and quantification of OTA, and sometimes also OTα, in different matrices. The methods presented here are similar to those described by Monaci et al.  or GiacoAs sometimes the available quantity of samples is very limited, the robustness of the method regarding the sample amount was evaluated. Starting from 1 g or 0.1 g of excreta, the extraction parameters were scaled down for the smaller sample size. However, no influence on the results was observed, confirming the ruggedness of our method. Moreover, different blood volumes dried on protein saver cards were evaluated regarding analyte recovery. The same results were obtained, regardless of the spotted blood volume. This is in accordance with results of Osteresch et al., where different blood volumes had almost no impact on the results for OTA . The samThe detection of OTA and its metabolite OTα in important pig and poultry matrices was established and validated. The developed methods allow the extraction and quantification of OTA and OTα in feed, plasma, DBS, liver, kidney, and muscle samples from pigs and chickens. Furthermore, extraction protocols for feces and urine samples from pigs as well as excreta, skin, and fat samples from chickens are available. Extraction is similar for all matrices, and either based on the green solvent ethyl acetate or on rather small volumes of ACN/water. Quantitative analysis of the analytes is done using LC-MS/MS. Method development was focused on the detection of biologically relevant concentrations. Therefore, the LOQ is between 0.5 ng/g in tissue matrices and 10 ng/g in feed or feces/excreta. The accuracy and precision of the methods were evaluated, and the analyte recovery was between 80 and 120% (80–150% for spiked concentrations ≤ 1 ng/g or ng/mL) of the initially spiked concentration and the RSD was ≤ 15%. Analyte stability in processed samples was tested at room temperature, at 4 °C and −20 °C for different storage periods. For long term storage −20 °C is recommended. The presented methods are easily established and feasible to monitor OTA exposure or for the use in feeding trials of pig and poultry to evaluate OTA-mitigating feed additives.13C-labelled OTA as internal standard were obtained from Romer Labs . The 13C-labelled OTα was produced in-house by enzymatic degradation of 13C-labelled OTA into 13C-labelled OTα and phenylalanine. An analytical standard for creatinine was purchased from Merck . All standards were stored as recommended by the supplier. Ortho-phosphoric acid 85%, formic acid, ACN HPLC grade, and acetic acid were obtained from VWR . Ethyl acetate HPLC grade and ACN HPLC-MS/MS grade were purchased from Chem-Lab . Methanol (MeOH) LC-MS grade was bought from Honeywell . Sodium hydrogencarbonate and Whatman 903 protein saver cards were obtained from Merck . Ultrapure water was produced in-house using a Milli-Q IQ 7000 water purification system from Merck .The analytical standards for OTA, OTα, and 13C-labelled OTA were purchased with a concentration of 10 µg/mL already dissolved in ACN. The 13C-labelled OTα was prepared by proprietary enzymatic degradation of 13C-labelled OTA. Progress of conversion of 13C-OTA to 13C-OTα was checked by LC-MS/MS. After complete conversion 13C-OTα was extracted five times with ethyl acetate. The combined ethyl acetate phases were dried completely under reduced pressure and reconstituted in ACN to a final concentration of 6.34 µg/mL. The same enzymatic reaction can be performed with any other OTA hydrolyzing enzymes, such as Carboxypeptidase A, under suitable reaction conditions.Standard stock solutions for OTA, OTα, and v/v/v) to the respective target concentrations between 1 µg/mL and 10 ng/mL. The prepared solutions were used for spiking the matrices at different concentration levels. Internal standard was prepared by dilution and mixing of 13C-OTA and 13C-OTα in ACN/water/formic acid (50/49/1 v/v/v) to 1 µg/mL, 0.25 µg/mL, or 0.1 µg/mL. Samples were spiked with internal standard during sample preparation, as described below. Further dilutions of internal standard were prepared and used for the external calibration in neat solvent.Stock solutions of OTA and OTα were mixed and used directly or optionally further diluted in ACN/water/formic acid , liver, kidney, muscle, skin, and fat samples were obtained in-house from the center of applied animal nutrition  or from the local slaughterhouse. Blood was analyzed using dried blood spots. All samples except feed, lyophilized feces or excreta, and DBS samples were stored at −20 °C until sample preparation. Feed, lyophilized feces or excreta samples were stored at room temperature in the dark until sample preparation. DBS were stored at 4 °C in the dark after drying overnight at room temperature. For all matrices quantitative determination was based on external calibration in neat solvent.v/v/v) and MeOH/acetic acid (99.8/0.2 v/v) was used as solvent B. Flow rate was set to 0.8 mL/min and a gradient was used during the chromatographic run, containing 0 min–1.6 min 0% B constant, 1.6 min–1.65 min linear gradient to 100% B, 1.65 min–2 min 100% B constant, and 2 min–2.05 min linear gradient to 0% B. The total run time was 3 min. The measured MRM transitions were 114 > 86  as quantifier and 114 > 44  as qualifier. The retention time of creatinine was 1.36 min. Quantitative determination of creatinine was based on a serial dilution of standards in MeOH/water (10/90 v/v). After determination of the creatinine values, urine samples were diluted individually with ultrapure water to 10 µM creatinine. If the creatinine value was below 10 µM, the samples were used directly. The concentration of analytes in urine depends, among others, on the amount of water the animal was drinking. To reduce variation in the results, urine samples were diluted to a specified concentration of creatinine. Determination of the creatinine concentration of each urine sample, and afterwards the individual dilution factor, urine samples were brought to room temperature, mixed shortly on a vortex shaker, and diluted 1:10,000 in ultrapure water. Creatinine concentration was determined by LC-MS/MS in positive mode. An Agilent 1290 Infinity II system was coupled to a Sciex Triple Quad 5500 mass spectrometer. Chromatographic separation was achieved on a Gemini 5 µm C18 column 150 × 4.6 mm with a suitable precolumn from Phenomenex . The column compartment was heated to 30 °C and the injection volume was set to 2 µL. Solvent A consisted of MeOH/water/acetic acid (40/59.8/0.2 13C-OTA and 13C-OTα. Then 800 µL extraction solution consisting of ethyl acetate/phosphoric acid 85% (99/1 v/v) were added and mixed by manual shaking to make sure that the samples did not stick to the bottom of the vial, before mixing vigorously for 10 min on a vortex shaker with adapter for 2 mL reaction tubes. The samples were centrifuged for 5 min at 19,000 rcf. Finally, 50 µL of the supernatant were transferred to an HPLC vial with 0.2 mL silanized insert and mixed with 50 µL ACN. The samples (diluted urine samples to 10 µM creatinine or plasma samples) were thawed and mixed shortly on a vortex shaker. An aliquot of 200 µL was transferred to a fresh Eppendorf tube and spiked with 8 µL of internal standard solution containing 1 µg/mL 13C-OTA and 13C-OTα. Then 6 mL 1 M phosphoric acid were added and mixed shortly, followed by the addition of 30 mL ethyl acetate. The samples were extracted on an end-over-end shaker for 60 min at 80 rpm. The samples were centrifuged for 10 min at 3200 rcf. Afterwards, 8 mL of the supernatant were transferred to a fresh 50 mL falcon tube and mixed with 4 mL 0.1 M sodium hydrogencarbonate solution (pH 8.2). The samples were mixed for 1 min on the end-over-end shaker at 80 rpm and centrifuged for 10 min at 3200 rcf. Then 3 mL of the water phase were transferred to a fresh 15 mL falcon tube and mixed with 70 µL ortho-phosphoric acid 85%. Finally, 1.5 mL ethyl acetate were added to the sample and the samples were mixed for 1 min on the end-over-end shaker at 80 rpm. The samples were centrifuged for 10 min at 3200 rcf and 50 µL of the supernatant were transferred to an HPLC vial with a 0.2 mL silanized insert and mixed with 50 µL ACN.Fresh feces and excreta were freeze dried. For extraction, 1 g of the sample was weighed in a 50 mL falcon tube and spiked with 30 µL of the mixed internal standard solution containing 1 µg/mL To test the robustness of the method, the used sample amount for extraction was reduced to 100 mg and all following steps were adapted to this smaller amount. 13C-OTA and 13C-OTα. The same procedure as already described in 5.3.2 for feed, feces, and excreta samples was used for further sample preparation. As the expected analyte concentrations are lower in tissue compared to feed or feces/excreta, an additional step for analyte concentration was added. Therefore, 1 mL of the supernatant after the second ethyl acetate extraction step was transferred to a fresh Eppendorf tube and dried completely under reduced pressure. The samples were reconstituted in 100 µL ACN/water/formic acid (50/49/1 v/v/v) in the ultrasonic bath for 10 min. Afterwards the samples were mixed vigorously for 10 min on a vortex shaker. Subsequently, the samples were centrifuged for 5 min at 19,000 rcf before 50 µL of the supernatant were transferred to an HPLC vial with a 0.2 mL silanized insert. Fresh tissue samples were frozen and stored at −20 °C in sealed bags. For homogenization, the samples were cooled in liquid nitrogen for approximately 1 min and pulverized using a ball mill  for 30 s with 30 Hz in two cycles. For extraction, 1 g of the sample was weighed in a 50 mL falcon tube and spiked with 30 µL of the mixed internal standard solution containing 0.1 µg/mL For the preparation of DBS, 50 µL whole blood were spotted on the protein saver cards and dried overnight at room temperature. To test the influence of the spotted volume, in addition 75 µL and 100 µL blood were spotted.v/v) were added and mixed. The samples were placed on an end-over-end shaker for 60 min at 80 rpm. Afterwards, 100 µL of the liquid were transferred to an HPLC vial with a 0.2 mL silanized insert. For DBS of pigs, the whole blood spot  was cut out from the protein saver card and placed in a 1.5 mL Eppendorf tube. Then 800 µL extraction solution containing ACN/water (70/30 13C-OTA and 13C-OTα were spiked in the tubes. Then 800 µL extraction solution of ACN/water (70/30 v/v) were added and mixed. The samples were placed for 60 min on an end-over-end shaker at 80 rpm. An aliquot of 500 µL of the supernatant was dried completely under reduced pressure at 60 °C and reconstituted afterwards in 62.5 µL ACN/water/formic acid (50/49/1 v/v/v). Samples were mixed vigorously for 10 min on a vortex shaker and afterwards centrifuged for 10 min at 19,000 rcf. Finally, 50 µL of the supernatant were transferred to an HPLC vial with a 0.2 mL silanized insert. A similar approach was used for DBS from chicken. Here, 50 µL of whole blood were spotted on protein saver cards and dried overnight. The entire spot was cut out and placed in a 1.5 mL Eppendorf tube. Additionally, 10 µL mixed internal standard solution containing 250 ng/mL of v/v) containing 0.1% formic acid and eluent B ACN/water (95/5 v/v) containing 0.1% formic acid. A gradient was used for separation: 0 min–0.25 min 20% B constant, 0.25 min–2 min linear gradient to 100% B, 2 min–2.50 min 100% B constant, and 2.50 min–2.51 min linear gradient to 20% B. The total run time was 3 min. Over the whole gradient the flow rate was set to 1 mL/min. The measured MRM transitions were optimized using direct infusion of the analytes. The optimized parameters for each analyte are listed in Quantification was based on reversed-phase LC-MS/MS in negative MRM mode. Therefore, an Agilent 1290 Infinity II system was coupled to a Sciex QTRAP 6500+ mass spectrometer. Chromatographic separation was achieved on a Kinetex 2.6 µm EVO C18 column 150 × 2.1 mm with a suitable precolumn from Phenomenex . The temperature of the column oven was set to 40 °C and 4 µL injection volume were used. The optimum chromatographic conditions were achieved with eluent A water/ACN  in the range from 0.25 ng/mL to 250 ng/mL. Linear regression was performed with a weighting factor of 1/analyte concentration, as recommended by Gu et al. [2 was calculated and the acceptance criteria was set at R2 >0.99.Linearity was assessed by preparing four individual calibration series through diluting defined concentrations of OTA, OTα, u et al. . The R2 Accuracy and precision were evaluated by analyzing blank samples, which were spiked with defined concentrations of OTA and OTα at two days at two different levels. The accuracy was calculated using the following equation:The acceptance criteria for accuracy were for spiking levels above 1 ng/g or ng/mL an analyte recovery of 80–120% of the initial spiked concentration. For spiking levels less or equal to 1 ng/g or ng/mL 80–150% analyte recovery was accepted. For intraday precision, the samples were spiked and analyzed in triplicate on the same day. For the interday precision two independent extractions were analyzed, as planned for further routine analysis. For the intraday and interday precision the acceptance criteria were set to RSD ≤ 15%. To calculate the RSD, the following equation was used:For the determination of the LOQ, spiking experiments were used. As defined in the Guidance for Industry 208, LOQ is the smallest measured content of analyte which can be determined with specified accuracy and precision . LOD wasSelectivity is given by the retention time and the specific MRM transitions of the analytes. Furthermore, the endogenous response of blank samples was evaluated. A signal of interferences at the retention time of the analyte below a S/N ≤ 3:1 was acceptable.Samples were extracted and extracts ready to inject were stored at different storage conditions in triplicate. The stability in process samples was evaluated at room temperature up to 7 days, in the fridge at 4 °C up to 7 days, and in the freezer at −20 °C up to 7 days for liver, kidney, and muscle samples, as low analyte concentrations are expected in combination with a high matrix content after drying and reconstitution, or at −20 °C up to 30 days for all other matrices. The acceptance criteria were 80–120% analyte recovery compared to the initial measured concentration.


Then, we divide the electrolyte into four parts: organic electrolytes; ionic liquid electrolytes; aqueous electrolytes; and solid-state electrolytes. Here, we aim to find electrode materials with a high specific capacity of charge and discharge at lower temperatures. Meanwhile, high-electrical-potential cathode materials and low-potential anode materials are also found. Furthermore, their stability in air and performance degradation in full cells and half-cells are analyzed. As for the electrolyte, despite the aspects mentioned above, its electrical conductivity in low temperatures is also reported.With the rapid development of electric power, lithium materials, as a rare metal material, will be used up in 50 years. Sodium, in the same main group as lithium in the periodic table, is abundant in earth’s surface. However, in the study of sodium-ion batteries, there are still problems with their low-temperature performance. Its influencing factors mainly include three parts: cathode material, anode material, and electrolyte. In the cathode, there are Prussian blue and Prussian blue analogues, layered oxides, and polyanionic-type cathodes in four parts, as this paper discusses. However, in the anode, there is hard carbon, amorphous selenium, metal selenides, and the NaTi Nowadays, with the rapid development of mobile electronic devices and increasing energy consumption, people’s demand for lithium-ion batteries (LIBs) is expanding . HoweverHowever, reported SIBs are mostly tested at room temperature, and SIBs that are able to work at low temperatures have not received much attention. With the development of SIBs, low-temperature performance is a factor that researchers have to consider. As reported, one of the factors limiting the large-scale application of SIBs is its poor performance at low temperatures . The mai6]3·nH2O) and PBAs (AxM[M′(CN)6]y·Z1−y·nH2O, A = Na, K; M, M′ = Fe, Mn, Ni, etc.; 1 < x < 2, 0 < y < 1; Z: vacancies) have the following advantages. First, their high energy density with a small volume changes per cycle and obtains a favorable rate capability in ambient temperatures. Second, their synthetic process is simple, which makes it suitable for large-scale industrial manufacturing [2Fe[Fe(CN)6] cubic nanoparticles on carbon nanotubes (CNT) to form a robust composite material (PB/CNT) as shown in + insertion and extraction process. In addition, a larger lattice parameter of the perovskite framework of PB reduces the activation energy of sodium-ion diffusion, and the concatenate of CNT in PB nanocrystals maintains favorable electric contact at low temperatures. The excellent performance of PB and the unique structure allowed PB/CNT cathodes to achieve a discharge capacity of 142 mAh/g at a 0.1 C rate (1 C = 100 mA/g in this review), as shown in 2Ni[Fe(CN)6] cathode, which was named as PBNi-ES. The ES method reported in the article significantly reduces the amount of combined water in the PBNi. In the characterization of SEM, it can be clearly seen that the PBNi prepared through the ES method has more pores and smaller particle size. This new structure has better stability under multiple cycles and a high current density, which also directly enhances the transmission efficiency of Na+ and e−. In 1.71Mn[Fe(CN)6]0.94□0.06·1.66H2O was composed through a facile in situ polymerization method. Combined with 3, 4-ethylenedioxythiophene, (NH4)2S2O8 and deionized water, the as-synthesized powder was named as MnHCF@PEDOT-20. The special prepared method and Mn/Fe in PBA promotes the infiltration of these two elements. It further decreases phase change in the cathode, enhancing the cell’s low-temperature performance. It is also worth mentioning that at a low temperature of −10 °C, NaPF6 was more stable in the electrolyte, while the degradation of transition metals was reduced. Hence, after a long run of 500 cycles, it still had a remarkable specific capacity of about 70 mAh/g, i.e., nearly 83% compared with its initial capacity, and it had an obvious voltage stage at 3.4 V. Recently, Jerry Barker et al.  was shaped as cylindrical lanterns. The cathode manifested as a NASICON type underwent two main processes when charge–discharge begun. In the first Na+ transfer reaction, there is a phase changing process which is a transportation from original Na3Fe2(PO4)3 to Na4Fe2(PO4)3. Then, as Na+ keeps transferring, the Na4Fe2(PO4)3 in cathode turned into Na5Fe2(PO4)3. Assembled with a hard carbon anode, the full cell delivered an ideal specific capacity of 74.6 mAh/g in 0 °C and 40 mAh/g at −20 °C with rate of 1 C, as shown in 4Fe3(PO4)2P2O7 structure combined with Mn2+ and modified with graphene as an SIB cathode. Both Mn2+ doping and graphene modifying effectively enhanced the ion transition; thus, the system showed impressive low-temperature performance. At a rate of 0.2 C at −20 °C, a specific capacity of 85 mAh/g can be obtained. Furthermore, at 0.5 C, its capacity retention is still 96.8% after 180 cycles at −20 °C, indicating that it has more favorable stability in extreme conditions compared with Na4Fe3(PO4)2P2O7 without Mn2+ doping.In another paper, Yan Wang et al.  synthesia et al.  prepared3V2(PO4)2F3 has a special polyanionic structure that consists of a 3D framework with V2O8F3 bi-octahedra connected by PO4 tetrahedra and large tunnels where Na+ is mobile upon extraction–insertion reactions. Laurence Croguennec et al. [3V2(PO4)2F3. Additionally, changes in the phase diagram upon cycling were observed by operando X-ray diffraction and other methods, as shown in 3V2(PO4)2O2F nano-tetraprisms, as shown in In the structure of fluorophosphates, highly electronegative F can improve the output voltage. Nac et al.  produced3V2O2(PO4)2F with reduced graphene oxide to enhance its electronic conductivity. It is proved that the open framework is more conductive to sodium-ion migration during charging–discharging processes at low temperatures. Hence, when combined with the polymer electrolyte, the Na3V2O2(PO4)2F/Na half-cell demonstrated specific capacities of 107 mAh/g, 84 mAh/g, 74 mAh/g, and 58 mAh/g, even under temperatures of −5 °C, −15 °C, −20 °C, and −25 °C at 1 C, respectively. After cycling at 1 C for 190 cycles under −25 °C, it still has a capacity retention of 99.6%, demonstrating superior low-temperature stability, as shown in 4Fe3(PO4)2(P2O7)/C nanocomposite via a facile one-step sol–gel method with high phase purity and uniform carbon coating, owing to the 3D sodium diffusion pathways and high sodium diffusion coefficient in Na4Fe3(PO4)2(P2O7), as shown in Maowen Xu et al.  hybridiz3V2(PO4)3 is used as a cathode material, as shown in Herein, some representative cathode materials were selected, and +. At the same time, CN− increases the opening of the perovskite surface. Meanwhile, the smaller charge and the three bonds in CN− further reduce the contact between Na+ and the anionic p-π electrons through the passing site face. In following experiments, the introduction of carbon nanotubes further enhanced the conduction of Na+; hence, the cathode has superior electrochemical performance at low temperatures. As mentioned above, the cathode materials obtained by introducing structural adjustment methods or through special preparation methods in the work of other research groups also exhibited outstanding low-temperature performance. Furthermore, PB and PBAs, which originated with the group of Professor John B. Goodenough of the University of Texas at Austin in 2010, were used in SIBs as part of large-scale commercialization attempts produced by Novasis Energies, Inc. (Novasis). This cathode still has stable cycling performance at low temperatures, while other SIB cathode materials have rarely been reported for large-scale commercial applications. We believe that PB and PBAs are the most ideal cathode materials in the future. In the end of the cathode materials’ section, it is worth mentioning that PB and PBAs are also our significant options. The unique perovskite skeleton in this cathode material has a large lattice parameter, which facilitates the diffusion of NaAs for SIB anode materials, hard carbon (HC) is the most practicable. HC is usually pyrolyzed by a high-molecular-weight polymer, so these carbons are difficult to graphitize at high temperatures above 2800 °C. Common HC includes resins such as phenolic resins, polyalcohols, epoxy resins, organic polymers, pyrolysis carbons, and carbon blacks (such as acetylene black). Due to the large interlayer spacing of hard carbon, it is conducive to the diffusion of ions between carbon layers and is also conducive to rapid charge and discharge. HC has a high specific capacity and is a promising carbon anode material with many current applications and research studies. However, at low temperatures, its kinetics slow down, resulting in serious polarization inside the battery. The aggravated polarization further reduces the electrochemical plateau to around 0 V. Due to the decrease in voltage, the deblocking ability of sodium ions to graphite anodes is greatly weakened, leading to an unfavorable specific capacity. This makes the low potential, which is originally an advantage of anode materials, one of the factors for the deterioration of electrochemical performance. Meanwhile, at lower temperatures, the formation of sodium dendrites is more likely to damage the SEI membrane, thereby reducing its stability and specific charge–discharge capacity. 3SO3/diglyme electrolyte, reactions in the battery were accelerated at low temperatures. At a rate of 0.5 C, the reversible capacity of the HC electrode is about 320 mAh/g at −5 °C, 310 mAh/g at −15 °C, and 302 mAh/g at −25 °C, as shown in Scientists aim to solve these problems through covering carbon and graphite on it. To conquer this bottleneck, M.R. Palacín et al.  develope3.5V2(PO4)2F3 cathode. A discharge capacity of 400 mAh/g was achieved for the half-cell and 244 mAh/g was achieved for the full cell at a rate of 0.1 C at −20 °C. However, the electrochemical performance of the full cell decayed dramatically at −20 °C because of more serious solid electrolyte interphase layers produced, compared with that in 25 °C after cycling revealed by the spectrum, as shown in 2 at a low temperature of −20 °C. Moreover, its capacity retention at −20 °C is around 87% compared to that obtained at ambient temperatures. In summary, because of the favorable electrochemical performance of HC below 0 °C, it can be constantly used to couple with the target cathode to study the low-temperature performance of full cells, which also proved the practicality of the HC anode.Moreover, Biao Zhang et al.  used lon2, CoSe2, and CuSe have received widespread attention for their low reaction energy consumption and superior theoretical specific capacity. However, when the temperature is reduced, during the sodium/disodium process, the conductivity of electrons and ions is low and the volume expansion is severe, resulting in electrode chalking and a loss of electrical contact, leading to extremely low cycle stability. Researchers improved their low-temperature performance by designing the structure of the unique Se anode material or modifying it with carbon material to accelerate the transfer of electrons and ions during charge and discharge, while increasing the active site. Se materials have also attracted extensive attention as a non-toxic, cheap, and abundant material. Amorphous selenium (a-Se), as a semiconductor, has marvelous electrical conductivity and a comparable high volumetric capacity, which make it attractive for anode material in SIBs. A-Se is also an abundant, environmentally friendly, and chemically stable material. Compared with carbon-based materials which is difficult to form intercalation compounds in Na-ion batteries, the a-Se alloy compounds have excellent electrochemical performance. Transition metal selenides such as MoSe2, FeSe+. When cycled in half-cells with the Na counter electrode, it exhibited favorable electrochemical performance when the temperature went down. For example, it showed a reversable specific capacity of 250 mAh/g at −5 °C and 180 mAh/g at −15 °C at a rate of 2 C. Moreover, it still had satisfactory capacity retentions of 96.2% and 98% after 1000 cycles, as shown in Xing-Long Wu et al.  coated a+ diffusion efficient. Moreover, as a widely acknowledged semiconductor, SnSe is abundant and environmentally friendly. In addition, it has a high theoretical capacity of 780 mAh/g for sodium-ion storage. In the research of Ming Zhang et al. [+ diffusion. In addition to the merits of carbon nanotubes mentioned above, the nanofiber ZnSe structure can also play a supporting role because of its superior bearing ability. It can stretch when the pressure is reduced and prevent the structure from collapsing when the pressure increases, resulting in improved electrochemical performance at low temperatures. For instance, the half-cell delivered a discharge capacity of 267.0 mAh/g at a rate of 1 C. Moreover, when it came to 600 cycles, an 83.3% capacity was still retained, demonstrating better cycling performance than pure ZnSe at −10 °C, as shown in Compared with amorphous selenium, metal selenides own higher electrical conductivity; thus, they are more favorable in low-temperature SIBs. From the SEM image in g et al. , a SnSe@g et al.  also cho7Se8@C structure with Fe and Se material. One of the advantages of this system is that one-dimensional carbon nanotubes in the three-dimensional structure facilitates the transfer of zero-dimensional Fe7Se8 nanospheres, as shown in 7Se8@C anode was assembled with a Na3V2(PO4)2O2F cathode which already showed an ideal capacity when the temperature decreased. The full cell exhibited a specific capacity of 166 mAh/g after 440 long-term cycles at 0.5 C. This means that its capacity fading per cycle was reduced to about 0.15% at an extremely low temperature of −25 °C, as shown in 2/rGO hybrids with a 3D hierarchical structure, prepared in a traditional hydrothermal way, were developed by Ye, Zhizhen et al. [In another work, Jing-Ping Zhang et al.  construcn et al. . Not onl2(PO4)3 electrode, hindering their application in electric vehicles. NaTi2(PO4)3 (NTP), with a sodium superionic conductor (NASICON) structure, has large ion channels for the insertion and extraction of sodium ions at a faster rate, thereby obtaining high electrical conductivity. At the same time, NTP can also be made into a one-dimensional structure, which is rich in sodium insertion sites. NTP also has the advantages of high capacity and stable structure, which gives it excellent electrochemical performance at low temperatures. To enhance the multiplicity performance of NTP, carbon coating is more adopted by scientists since it not only effectively promotes the conductivity of it, but also reduces particle size and prevents the oxidation of metal ions. Chunhua Chen et al. [3V2(PO4)3 (NVP). Then, he coated the mixture with graphene-like layers to synthesize the new nano-porous electrode (NTP@C-2), as shown in Low electrical conductivity and the ion diffusion coefficient affect the multiplicity performance of the NaTin et al.  mixed tr3(VOPO4)2F, and an ionic liquid electrolyte, which manifested as an ideal low-temperature electrochemical performance. It will be discussed in the ionic electrolyte section. Zhanliang Tao et al. [+ and brings about fast reaction kinetics of the cell. Moreover, when assembled with the Ni(OH)2 cathode and the 2 M NaClO4 electrolyte, the full cell demonstrates remarkable low-temperature rate performance through a dual-ion reaction. When at the high of 10 C, after 500 cycles, the capacity retention showed an unusual perfect capacity retention of 100% at −20 °C which was much higher than that of 25 °C. The full cell also delivered a specific capacity of 82 mAh/g at 0 °C, 78 mAh/g at −10 °C, and 70 mAh/g at −20 °C with a voltage stage of 1.25 V, as shown in In addition, Ruben-Simon Kühnel et al.  designedo et al.  coated N1.25Ti0.75O4 was also prepared and investigated in a high-crystalline structure as an all-climate anode material [1.25Ti0.75O4 has typical 1D channels of post-spinel, wherein the channel skeletons are constituted by VO6 and TiO6 octahedrons in the form of vertices or edge-shared linking, as shown in 1.25Ti0.75O4, this material can achieve a high DNa+ of 3.48 10−11 cm2 s−1, even when the temperature dropped to −25 °C. Hence, assembled with the Na0.8Ni0.4Ti0.6O2 cathode, the full cell’s discharge capacity can reach 93 mAh/g at a 0.2 C rate, as shown in Besides the above-mentioned three types of anodes, high-crystallinity Ti-based oxide NaVmaterial . NaV1.25In summary, representative anode materials were selected and plotted according to voltage and electrical properties, as shown in +/Na and high cycling stability during de/sodiumization. The smooth electrochemical plateau of HC is attributed to the intercalation and deintercalation of Na+ into graphite interlayer gap which is larger than 0.37 nm. HC, which has been widely used as a node material, also has higher stability and better electrochemical performance than other anode materials at low temperatures. The most important thing is that the precursor of HC is cheap and environmentally friendly, which has incomparable practical advantages compared to other anode materials. General HC precursor materials are composed of various biopolymers, including hemicellulose, cellulose, lignin, pectin, protein, free sugars, etc. In our article, both longan peel and tissue paper used as precursor materials showed satisfactory low-temperature performance. The main progress now lies in annealing the precursor and further improving the porosity inside the HC to increase the Na+ diffusion rate and improve its electrochemical performance at low temperatures.Among all the above-mentioned anode materials, HC anode materials are most valued due to their low potential versus Na+, but also participates in the formation of the SEI [As an important component of SIBs, the electrolyte is a medium for ion conduction that provides pathway to support electrochemical reactions on electrodes . The ele the SEI . Therefo the SEI . Low-tem the SEI , but als the SEI . In addi the SEI . All theTherefore, electrolytes that can be applied under low-temperature conditions should have the following characteristics: (a) favorable stability; (b) good ionic conductivity; (c) a wide electrochemical window; (d) no reaction with other battery components; (e) high safety and low toxicity, and (f) can meet the cost requirements for practical applications. The above characteristics of electrolytes essentially depend on the property and selection of the electrolyte salt . While a4, NaPF6) with different carbonic acid derivative . Meanwhile, the performance of electrochemical SIBs can be significantly improved by changing the composition and ratio of carbonic acid derivatives or slightly adding some other additives. At the same time, matched with suitable anode and cathode materials, the SIB can be manifested given its favorable electrochemical capacity and stability. Among sodium salts, NaClO4 is more abundantly studied at low temperatures. In addition, the PC solvent has a freezing point of −48 °C and an outstanding dielectric constant which effectively avoids any solidification of the electrolyte system and improves the low-temperature performance of SIBs by increasing the ionic conductivity. However, the single-component PC is less compatible with the electrodes, so efforts are devoted to solving the problem with additives [4 dissolving in PC with the addition of 2 vol% FEC exhibited a specific capacity of 94 mAh/g at a 0.2 C rate with an 8.7% capacity decline compared to that the 25 °C in the half-cell test with a NaV1.25Ti0.75O4 cathode at −20 °C. Combined with the Na0.8Ni0.4Ti0.6O2 anode, the Na0.8Ni0.4Ti0.6O2//NaV1.25Ti0.75O4 full cell manifested a specific capacity of 93 mAh/g at a 0.2 C rate and maintained a 84% retention capacity over 200 cycles at a 1 C rate. Chungang Wang et al. [Organic electrolytes are usually a mixture of sodium salts  as an electrolyte combined with PB/CNT and NVP@C, before then being tested in a half-cell. The PB/CNT half-cell disclosed 155 mAh/g and 105 mAh/g specific capacities in 0.3 C at 0 °C and −25 °C, respectively. Even at a 2.4 C rate and super long cycles of 1000, the cell could still maintain high capacity retentions of 81% and 86%, respectively. As for the NVP@C, an oblivious decay of the specific capacity can be noticed with the temperature decreasing. As the temperature changed by ranges of 0 °C, −10 °C, and −20 °C, the specific capacity changed from 120 mAh/g to 116 mAh/g at a 0.2 C rate. Meanwhile, at −20 °C, with a rate of 10 C, the retention capacity could still remain 75% after 500 cycles. In the works of XingLong Wu et al. and JingPing Zhang et al. [4 with a EC/PC cosolvent system which magnificently enhanced the specific capacity. The specific capacity of the cl-Fe7Se8@C half-cell reached 425 mAh/g with a current density of 0.2 A/g, which decreased to 350 mAh/g from 0 °C to −25 °C. Compared with the performance at 2 C at room temperature, the capacity retentions of the cell reached about 96% at 0 °C, 82% at −15 °C, and 68% at −25 °C. Meanwhile, the experiment of 3DSG composite half-cells demonstrated a high specific capacity of 375 mAh/g at −5 °C, 300 mAh/g at −15 °C, and 250 mAh/g at −25 °C, respectively, in a large current density of 2 A/g. Moreover, the retention capacities were 96.2%, 98%, and 90.4%, respectively, over 1000 cycles at 20 C. Conventional solvent EC has a high dielectric constant and outstanding membrane formation, but its high melting point and viscosity limit the application of SIBs at low temperatures, so EC needs the addition of a suitable co-solvent to improve the low-temperature performance of SIBs . It is cg et al. ,50, the 4 was investigated in the low temperatures of 0 °C and −25 °C by Jian-Ming Dai et al. [+ and decreases in conductivity, as characterized in 4 dissolved in the EC/DEC (1:1 vol) cosolvent, it delivers inconspicuous capacity fluctuation as the temperature decreased [Moreover, cosolvents formed by EC and other carbonic acids also performed stable low-temperature electrochemical functions. EC dissolved with DMC (1:1 vol) after adding 1 M NaClOi et al. . Assemblecreased . In the ecreased , at a ra4 dissolved salts in the electrolyte, NaPF6 is manifested as no big difference in electrochemical performance. Meanwhile, it is proven to have an ideal low-temperature performance. In the research of the carbon-covering HC anode, 1 M NaPF6 solution in EC and PC mixture (1:1 wt) with an additive of 10 wt% DMC was chosen by M.R. Palacín et al. as the electrolyte. At low temperatures of 0 °C and −15 °C, it delivered favorable specific capacities of 260 mAh/g and 265 mAh/g at a rate of 0.1 C, respectively, which showed a slight degradation compared to that at 25 °C (290 mAh/g). Additionally, Yang-Kook Sun et al. [6 solution. Attributing to the low-temperature function of the unique cathode and the favorable electrolyte, the full cell sandwiched by HC anodes demonstrated appreciable specific capacities of 130 mAh/g and 100 mAh/g at a 0.75 C rate with slightly different retention capacities of 89% and 92% after 100 cycles at a 0.75 C rate. In 4 and NaPF6 solvents with organic solvents have a wide electrochemical window and ideal thermodynamic stability. Meanwhile, in the group of Huang, Yunhui et al. [3V2(PO4)2O2F half-cell. By assembling into Na/ Na3V2(PO4)2O2F batteries, a high capacity retention of 93.1% was realized after 1000 cycles at 1 C. Under an ultra-high rate of 30 C and a low temperature of −30 °C, the discharge capacities still reached 89.2 and 92.1 mAh/g, respectively. This also proves that SIBs with different electrode materials mentioned above, combined with conventional organic electrolytes, exhibit practical electrochemical capabilities at low temperatures. Therefore, with the easy synthesis processes, the conventional organic electrolyte is the most suitable choice for SIB at low temperatures.Furthermore, compared with NaClOn et al.  tested a−3 S/cm and 3.8*10−4 S/cm, respectively. Ionic liquids are ionic, noncombustible, and nonvolatile, and salt-like electrolytes can provide broad electrochemical window, as well as high stability and safety. Moreover, it has a broad temperature range that gives a promising low-temperature application horizon . Imidazo3C1pyrrFSI) was proven to have a favorable electrical conductivity (9.8*10−4 S/cm) at a low temperature of 0 °C. When NaFSI-C3C1pyrrFSI was assembled with NaCrO2, about 85% and 51% of the initial discharge capacities were maintained (100 mAh/g and 60 mAh/g) at 0 °C and −10 °C, respectively. It is also worth mentioning that after a series of temperature changes from 90 °C to −20 °C and back to 90 °C, the discharge capacity returned almost completely to its initial value, indicating that no significant degradation occurred at −20 °C, as shown in 3C1pyrrFSI was assembled with Na2FeP2O7, the half-cell demonstrated a specific capacity of 80 mAh/g at 0 °C, 67 mAh/g at −10 °C, and 42 mAh/g at −20 °C [14TFSI-NaTFSI  mixtures and proved that with a molar fraction of 9:1, the electrolyte showed remarkable conductivity (2.2*10−3 S/cm) at −30 °C, as shown in Meanwhile, pyrrole ionic liquids are cyclic quaternary amine ionic liquids with similar physicochemical and structural properties as chain quaternary amine ionic liquids. In the study of Rika Hagiwara et al. , NaFSI-[t −20 °C . Moreovet −20 °C , they in4 dramatically decreases the freezing point of the electrolyte and improving the low-temperature performance. At an extremely low temperature of −50 °C, the electrolyte conductivity reached 1.1*10−4 S/cm. Furthermore, the electrochemical function was tested by sandwiching the NaTi2(PO4)3@C (NTP@C) anode and carbon cathode. The full cell with the 2M-0.3 electrolyte exhibited an excellent specific capacity (ca. 68 mAh/g) and cycle performance (95%) over 100 cycles at a rate of 0.665 C. As demonstrated by Zhanliang Tao et al. [4 can be used as an electrolyte without any additives in extreme environments. Sandwiched by the NaTi2(PO4)3 anode and the nano Ni (OH)2 cathode, the full cell delivered favorable characteristics through a dual-ion reaction. It showed specific capacities of 82 mAh/g at 0 °C, 78 mAh/g at −10 °C, and 70 mAh/g at −20 °C, with an average voltage of 1.25 V. Moreover, the capacity retention remained 85% under a rate of 10 C after ultralong life of 10,000 cycles. Then, combined with SEM results, it can be concluded that there is no pulverization or agglomeration on the surface of the electrode at low temperatures. SIBs based on aqueous electrolytes are safe, environmentally friendly, inexpensive, and less corrosive; thus, they are considered for different perspectives and applications . Simultao et al. , NaClO4 0.44MnO2 cathode and a Zn anode to reduce the irreversible proton co-insertion caused by aqueous media. The electrolyte consists of a 1 M sodium acetate in the 5:1 v/v ethanol–water solution compared with different ethanol–water volume ratios disclosed by contact angle tests . −5 S/cm, which is very similar to that of Xu’s work. Assembled with the polyanionic cathode Na3V2O2(PO4)2F and the PFSA electrolyte, the cell shows a highly reversible sodium-ion extraction–insertion process, resulting in remarkable low-temperature electrochemical performance. Meanwhile, due to the perfect match among the NASICON cathode and the PFSA-Na membrane, the system shows limited polarization. The half-cell delivered suitable discharge capacities, high voltage, and stable low-temperature performance, as mentioned in the cathode part. Liquid electrolytes have safety hazards and flammability risks when vehicles are hit or the temperature rises sharply which has been leaked in lithium-ion battery energy vehicles. The lithium-ion battery car combustion incidents that have occurred in recent years have brought people’s concerns back to battery safety. Furthermore, the higher viscosity of the electrolyte at low temperatures causes more severe polarization, which is an important reason for the electrochemical performance degradation of liquid electrolytes. Thus, solid-state sodium batteries (SSIBs) have emerged as an attractive choice to solve these problems. Maowen Xu et al.  prepared4/TEP-VC electrolyte which acted as a plasticizer. Finally, the target electrolyte was obtained. It was proven that Na dendrites, which seriously impact SEI formation, are hindered by quasi-solid P electrolytes. Moreover, the investigation elucidated that it could promote the synthesis of an interface between the NVP cathode and the electrolyte, which suppressed the vanadium’s dissolution and enhanced the batteries’ safety. Therefore, when assembled with the NVP anode, the half-cell exhibited stable electrochemical performance in low temperatures. A 70 mAh/g specific capacity was achieved at −10 °C with a retention capacity of 84.8% after 50 cycles at 0.1 C, as shown in 4 in the PC and 5% FEC into the photocuring agent ethoxylated trimethylolpropane triacrylate (ETPTA) and 1% 2-hydroxy-2-methylpropiophenone, before being dropped on a cellulose film and cured with ultraviolet light. Due to the resultant, the QSSE exhibited remarkable ionic conductivity of 7*10−4 S/cm at a low temperature of 0 °C and 8*10−4 S/cm at a low temperature of −10 °C.In another group, Guanglei Cui et al.  reported3Zr2Si2PO12-based composite to form polymer electrolyte. As for the constitution, the weight percentage ratio is 70: 15: 15. Here, 70% is the Na3Zr2Si2PO12-based composite, 15% is poly (vinylidene fluoride-co-hexafluoropropylene) (abbreviate as PVdF-HFP), and 15% is 1 M NaCF3SO3/TEGDME. Meanwhile, when the temperature goes down, PVdF-HFP effectively promotes the stability of the interface of two solid materials. The application of ceramic powder not only plays a similar role as the PVdF-HFP binder, but it also facilitates the transformation of Na+, resulting in high conductivity in low temperatures. Unlike at low current densities, the addition of strong ionic forces to the conductive ceramic at high current densities can facilitate the transfer of sodium ions in the electrolyte shell, resulting in the promising electrochemical performance of conductive ceramics at high rates, as shown in −4 S/cm at 0 °C, 1*10−4 at −10 °C, and 2*10−5 S/cm at 0 °C, as shown in 2O3 nanoparticles 3 coating, NaCrO2 nanowire fabrication, or hierarchical columnar structure design, can reduce the attenuation of battery performance at low temperatures. As for polyanionic-type cathodes, C coating, CNTs, and C nano wares are also effective ways of advancing low-temperature cathode performance. By comprehensively considering cathode performance at low temperatures, many materials have a specific capacity of about 100 mAh/g and a relatively stable voltage window of about 3 V, showing bright application potential.PB and PBAs are widely used as traditional cathode materials in SIBs. Combined with CNTs, the ionic conductivity of cathode material was enhanced, and the charge–discharge specific capacity and cycle stability were also promoted. Simultaneously, some special cathode structures, such as cubic nanoparticles and crystalline particles, also make the batteries, showing ideal stability at low temperatures. Then, layered oxides composed by transition metals exhibit favorable electrochemical performance. Further structural modifications, such as NaTi2(PO4)3 can also obtain favorable electrochemical properties by combining sp2-type carbon or high-crystallinity NaV1.25Ti0.75O4 in the anode. Considering representative materials, we found that HC is still the material with the lowest voltage and largest specific capacity.HC is mostly used as an anode material in SIBs. At low temperatures, some effective modifications to HC can also play a role in promoting its performance. Similar to cathodes, in some special ways, such as C coating or structure-like micropores, HC can increase the conductivity of the anode to obtain an ideal battery-specific capacity. Amorphous selenium and metal selenides, on the other hand, use Se in combination with the metal elements Zn, Fe, or Sn as the anode, and can obtain a lower charge and discharge potential after modification with CNTs or graphite. NaTi+ transfer rate in the electrolyte is slowed down, and the ion conductivity of the electrolyte and the diffusion rate of Na+ in the active material will be reduced. The charge transfer process is difficult at the solid–liquid phase interface on the surface of material. In the four electrolytes mentioned in this paper, the traditional organic electrolytes need to be mixed with organic mixtures of different proportions to improve its efficiency. The ionic liquid electrolytes have the highest ion transfer rate and dielectric constant. Aqueous electrolytes also have ideal conductivity at low temperatures and are environmentally friendly. Solid-state electrolytes have relatively safe qualities, can effectively avoid the fire hazard on organic electrolytes, and have been proven to have practical low-temperature performance. This article provides feasible suggestions for the future development of SIBs at low temperatures.In the end, under low-temperature conditions, the Na


Mn, but significantly increased DO concentrations throughout the experimental period. Moreover, ecological aquaculture with submerged macrophytes showed strong effects on the phytoplankton and bacterial community compositions. In particular, the M group ponds had higher phytoplankton density and mainly included Cyanobacteria, whereas the EM group had lower phytoplankton density and mainly included Chlorophyta. Moreover, higher alpha diversity, as determined by Ace and Simpson index values, was detected for bacterial communities in the EM group ponds. Furthermore, PCoA clearly grouped the bacterial communities according to the two culture modes throughout the culture period. These results indicate that ecological aquaculture with submerged macrophytes can improve water quality, control Cyanobacterial blooms, and affect the diversity and composition of bacterial communities. These valuable effects seem to be beneficial and consistent to maintaining aquaculture ecosystem stability.Traditional aquaculture ponds are one of the most vulnerable ecosystems; thus, ecological aquaculture is increasingly valued for its beneficial ecological properties and ecosystem services. However, little is known about ecological aquaculture of largemouth bass with submerged vegetation. Here, we designed three ecological ponds of cultured largemouth bass with submerged macrophytes (the EM group) and three ponds with traditional aquaculture (the M group) to reveal the response of water quality, and phytoplankton and bacterial communities, to submerged macrophyte bioremediation during a 90-day culture period. We observed that Cyanobacterial outbreak occurred in the M group ponds from day 7 to the end of the experiment; however, there were no Cyanobacterial blooms in the EM group ponds throughout the culture period. Compared with the M group ponds, the EM group ponds, which had submerged hydrophytes, had significantly decreased concentrations of TP, TN, and COD The largemouth bass were obtained from Zhanglin Fishery Co., Ltd.  and allowed to acclimate to a pond for 2 weeks, and they were fed commercial fish food . Subsequently, fish with an initial body weight of 14.50 ± 0.23 g were arbitrarily allocated to the six ponds used in this study (0.17 ha) with the same stocking density of 43.48 g·m−3. During the experiment, fish were fed two times daily (08: 00 and 18: 00) to apparent satiation. Three months before the start of the experiment 10 l pond water each pond and stored at 4°C to maintain the bacterioplankton community.The experiment was conducted at the Yang Zhong experimental base . In this study, the three ponds without submerged macrophytes were considered the control group (M), and the three ponds with submerged macrophytes were considered the experimental group (EM). One month before the start of the experiment 2.2.−1 MS-222, body weight, body length, body thickness and liver and visceral weights of fish were measured. Total of 30 l of water samples were collected by the mixing method of uniform five-point sampling in the same pond on days 0, 10, 20, 30, 60, and 90; and mixed samples of water from the upper, middle, and lower portions of the pond at each point. M group samples were labeled as MW_1, MW_2, MW_3, MW_4, MW_5, and MW_6, respectively, whereas EM group samples were labeled as EMW_1, EMW_2, EMW_3, EMW_4, EMW_5, and EMW_6, respectively . Thirty fish in each group were collected separately at the beginning and the end of the experiment, and anaesthetized with 100 mg·Lectively . Water d methods . AdditioWt – W0) × 100 /W0, SGR (% day−1) = (lnWt – lnW0) × 100/days, HSI (%) = (hepatosomatic weight/body weight) × 100.where W0 is the initial body weight and Wt is the final body weight.In addition, we calculated the weight gain rate (WGR), specific growth rate (SGR), and hepatosomatic somatic index (HSI) as parameters reflecting growth performance, as follows: WGR (%) = (2.3.Y), Y = (ni/N) i× f , throughout this study, Y > 0.02 was considered a dominant species.Phytoplankton counts and species identification were carried out in the laboratory with an optical microscope  and a stereoscope , respectively. Further calculations were evaluated with the Mcnaughton dominance index  following the manufacturer’s protocols. The DNA from all samples was examined for purity by electrophoresis on 1.0% agarose gels, and DNA concentrations were detected by a Nano Drop 2000 spectrophotometer .The V3–V4 regions of 16S rRNA of microorganisms were amplified by PCR with primers 338F (5′-ACTCCTACGGGAGGCAGCAG-3′) and 806R (5′-GGACTACHVGGGTWTCTAAT-3′). PCR was conducted in a 20-μL volume, which contained 4 μL 5× FastPfu buffer, 2 μL 2.5 mmol/l dNTPs, 0.8 μL Primer (5 μM), 0.4 μL FastPfu polymerase, and 10 ng DNA template. PCR were performed in triplicate for each sample. The thermocycling conditions were as follows: initial denaturing at 95°C for 3 min; followed by 27 cycles of 30 s at 95°C, 30 s at 55°C and 30 s at 72°C; and final extension at 72°C for 10 min.2.5.PCR products were purified with the AxyPrep DNA Gel Extraction Kit  following the manufacturer’s instructions. PCR products were then eluted with tris HCl and detected by 2% agarose gel electrophoresis, and detected and quantified using QuantiFluor™-ST . Subsequently, we purified and constructed the sequencing library. The purified amplified fragments were constructed into PE2*300 library according to the Illumina MiSeq platform , and sequenced by the MiSeq PE300 platform .p < 0.05).Bioinformatic analysis of the water microbiota was carried out using the Majorbio Cloud platform2.6.p < 0.05 between groups were identified as significant, in this experiment, p-value generally refers to the difference of two treatments at the same time point. All data were analyzed using IBM SPSS Statistics v26.Data were collated using Microsoft Excel and all results are presented as the means ± SD values. Student’s t-test was used to analyze differences in water quality, phytoplankton composition, and microbial communities between the two treatments. Differences with 3.3.1.Micropterus salmoides in EM group than M group had significantly higher body length, body thickness, body weight, weight gain rate (WGR), and specific growth rate . Whereas fish in the EM group showed a significantly lower HSI than those in the M group (p < 0.05).The growth performance was significantly divergent between EM and M groups . SpecifiMn (p < 0.05), but significantly increased DO concentrations on day 0, 10, 20, 60 and 90 (p < 0.05). NH4+-N in the EM group ponds was significantly lower than that in the M group ponds on day 10 (p < 0.05); however, in other periods, NH4+-N was more similar and the concentrations were all less than 0.2 mg/l. NO2−-N and NO3−-N in the EM group ponds were significantly higher than those in the M group ponds on days 20, and 30 (p < 0.05), but there were no significant differences on days 60 and 90 (p > 0.05).The physicochemical properties of water samples are shown in 3.2.3.2.1.During the culture experiment, a Cyanobacterial outbreak occurred in the M group ponds from day 7 to the end of the experiment ,B. Howev3.2.2.6 to 3.06 × 107 cells/L, which was extremely significantly higher than that of the EM group for every month in the experimental period  showed that genera Chroococcus and Microcystis, which are cyanobateria, had total values over 50% in the M group, whereas genera Chlorella and Scenedesmus, which belong to Chlorophyta, had values close to 50% in the EM group . Subsequently, the Ace index of the EM group was similar on days 60 and 90, but it was also significantly higher than that of the M group in the same sampling period , significantly increased from day 10 (p < 0.05). However, there were no dissimilarities between the two groups on day 90 , whereas CL500-29_marine_group, Mycobacterium, norank_f__67–14, Acidibacter, and norank_f__Saprospiraceae were negatively correlated with DO and pH (p < 0.05). Furthermore, Microcystis_PCC-7,914, norank_f__norank_o__PeM15, norank_f__67–14, and Acidibacter exerted very significant positive correlations with TP (p < 0.01), which can be used as an indicator of trophic status. In addition, Dinghuibacter and Sphingorhabdus were found to be significant negatively correlated with TN (p < 0.05).A Spearman’s correlation analysis was performed to evaluate the potential associations between water physicochemical parameters and bacterial communities . We foun4.Intensive aquaculture is on the rise worldwide and includes introducing formulated feed into production systems. However, in most countries, unstable and uncontrolled traditional intensive production has led to environmental and social problems . Given tMn is an indicator of organic pollution  can be found in the article.The animal study was reviewed and approved by Freshwater Fisheries Research Center (FFRC), Chinese Academy of Fishery Sciences (CAFS).GX: accepted and designed research, reviewed the manuscript and approved the final manuscript. PX: reviewed the manuscript and approved the final manuscript. ZN: designed research, guided experimental methodology, wrote the manuscript, reviewed the manuscript and approved the final manuscript. ZZ: feeding, sampling, conducting the experiment, data analysis, wrote the manuscript and approved the final manuscript. HZ: sampling, collected and analyzed the data, read and approved the final manuscript. YS: sampling, feeding and approved the final manuscript. JG and JcG: sampling, data analysis, approved the final manuscript. All authors contributed to the article and approved the submitted version.This work was supported by National Key Research and Development Program of China (No. 2019YFD0900301), and the earmarked fund for CARS-46.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


The detection limit for TBZ was 6.2 × 10−7 M. The response time of the sensor for TBZ was 8 s, and its signal drift was only 1.7 mV/h. The new sensor is applicable for direct potentiometric determination of TBZ in complex real samples, such as fruit peel. The accuracy of TBZ determination is confirmed using the standard addition method.Direct potentiometric measurements using solid-state sensors have a great potential for thiabendazole (TBZ) determination, considering simplicity, accuracy, and low cost. Modifying the sensing material of the sensor with multi-walled carbon nanotubes (MWCNTs) leads to improved analytical properties of the sensor. In this study, a new potentiometric solid-state sensor for TBZ determination, based on MWCNTs modified with a sulfate group, and TBZ ion as sensing material was developed. The sensor exhibited a Nernstian response for TBZ (60.4 mV/decade of activity) in a working range between 8.6 × 10 Thiabendazole (TBZ) is a benzimidazole derivative with a wide area of application as a fungicide. It is one of the most detected pesticides in the United States of America and Europe . TBZ is Chromatographic methods are the most commonly used for TBZ determination ,13,14,15This paper describes the preparation of a new hybrid material based on MWCNTs modified with a sulfate group and TBZ ion and its application as a sensing material in the liquid membrane for a new solid-state sensor for TBZ determination. This research aimed to develop a simple, fast, accurate, and improved potentiometric method for TBZ determination, which was possible due to introducing modified MWCNTs as a sensor material. A newly developed potentiometric sensor was characterized, and its applicability was demonstrated in pure TBZ solutions and fruit samples.3H , and n-hexane ) and nitrate salt of TBZ (prepared according to [3)3 9H2O , and absolute ethanol ) were used for the new sensing material (MWCNT-OSO3−TBZ+) preparation. Then, 100 mg of MWCNT-OSO3H was mixed with 2 mL of water and placed in an ultrasonic bath to form a stable homogenous suspension. A TBZ solution (c = 1 × 10−2 M) was added dropwise in suspension until a flaky precipitate was formed, and it was stirred using a magnetic stirrer for 24 h. The precipitate was washed with water and centrifuged (5 min at 5000 rpm) three times, followed by drying for 24 h at 60 °C. The newly prepared MWCNT-OSO3−TBZ+ (0.0018 g), tetrahydrofuran , dimethylformamide , poly(vinyl chloride) , and dibutyl sebacate  were mixed in order to prepare the liquid membrane of the sensor. The assembling of the new sensor and drop-casting of graphene layer  and membrane mixture on the graphite core of the sensor were performed according to [MWCNT-OSOrding to , using Mrding to  using TBrding to .HCl  was used to prepare the analyte (TBZ). All salt solutions for interference measurements, L(+)-ascorbic acid , citric acid , glucose , fructose , imazalil , fuberidazole , carbaryl , carbendazim , 2-phenylphenol  and NaOH  were prepared using analytical grade chemicals. Deionized water (conductivity of 0.055 µS/cm) was used to prepare all solutions.Bananas, oranges, lemons, clementines, and limes were purchased from the local store in Osijek (Croatia) and were not declared bio fruit. Bananas originated from Ecuador, oranges and clementines from Greece, and lemons and limes from Spain.An ultrasonic bath  was used to prepare the sensor membrane and TBZ solution. Measurements were performed using 794 Basic Titrino coupled with 806 Exchange unit, 826 mobile pH meter, 728 stirrer, Tiamo software , and in-house software. SEM images of the electrode surfaces were taken by SEM  using back-scattered electron detector. Samples were analyzed without gold coating.3−TBZ+ sensor as the working electrode and silver/silver chloride electrode  as the reference electrode. Conditioning the sensor in TBZ solution (c = 2.0 × 10−3 M) for 15 min and short calibration (concentration range between 1.0 × 10−6 and 1.0 × 10−3 M) were performed daily before measurements in order to test the sensor performance.All measurements were performed at room temperature with magnetic stirring and without ionic strength adjustment, using a newly developed MWCNT-OSOc = 2.0 × 10−3 M) was prepared using HCl (c = 2.0 M) addition. pH in the TBZ solution was 2.6.The TBZ is slightly soluble in water but can be soluble in acidic media; thus, the solution (c = 2.0 × 10−3 and 5.0 × 10−5 M) was incrementally added to distilled water (V = 20 mL) using in-house software. Dynamic response measurements were performed by adding TBZ (c = 2.0 × 10−3 and 5.0 × 10−4 M) to distilled water (V = 50 mL) with 45 s pause between additions. Signal drift was measured in TBZ solution . The influence of potential interferences ) was investigated using the fixed interference method [c = 1.0, 1.0 × 10−1 and 1.0 × 10−2 M) were used for measuring the influence of pH on the sensor response. For real samples preparation, chopped peel of bananas, oranges, lemons, clementines, and limes (m = 100 g) were placed in a glass and covered with distilled water , followed by pH adjusting to 2.6. After 24 h, the samples were filtered through the gauze and measurements were performed (V = 15 mL).For the response measurements, TBZ  was used as the analyte. The investigated concentration range was 2.5 × 10−8 to 1.0 × 10−3 M. The measured response can be seen in In response, characterization of the new MWCNT-OSO3−TBZ+ sensor revealed a Nernstian response (60.4 mV/decade of activity) to TBZ in a concentration range between 8.6 × 10−7 and 1.0 × 10−3 M. Due to its wide concentration range, as well as its low limit of detection, it could be applicable for TBZ determination in various real samples.The new MWCNT-OSO3−TBZ+ sensor with other potentiometric sensors for TBZ determination is presented in 3−TBZ+ sensor has significantly better analytical properties, such as lower limit of detection, wider measuring concentration range, faster response and prolonged lifetime.The comparison of the new MWCNT-OSO−7 and 1.0 × 10−4 M. The resulting response is presented in 3−TBZ+ sensor has a fast response, with an average response time of 8 s.Dynamic response time is another sensor characteristic that is also important for its applicability for TBZ determination. It represents the time needed to attain 90% of the final potential value after a sudden change in TBZ concentration . The res3−TBZ+ sensor was measured in TBZ solution (c = 2.0 × 10−3 M) using linear regression analysis, and it is described by the equation: E (mV) = −0.0007 × t(s) + 360.28. The calculated signal drift was −1.7 mV/h.The stability of the sensor response can be observed using signal drift. It represents a change in the sensor potential as a function of time. Signal drift of the new MWCNT-OSOc = 1.0 × 10−2 or 2.5 × 10−4 M) and variable concentration of TBZ . The potentiometric selectivity coefficients were then calculated by mathematically adjusting the Nikolskii–Eisenman equation (Equation (3)) to experimental data using Solver (Microsoft Excel).The sensor’s selectivity is one of its most important properties that determine its applicability for TBZ determination in real samples. It can be estimated by calculating potentiometric selectivity coefficients  was used. The results are presented in TBZ is slightly soluble in water. Its solubility increases in acidic media; thus, it was important to investigate the influence of pH on the sensor response. For that purpose, TBZ solution .It can be observed that the sensor potential was stable in the pH range between 2 and 4, followed by a decrease in potential. Although the working pH range of the new MWCNT-OSO3−TBZ+ sensor is a TBZ determination using direct potentiometric measurements; thus, its applicability was tested in pure TBZ solutions and fruit peel samples.The main purpose of the new MWCNT-OSOc = 5.0 × 10−4, 1.0 × 10−4, 5.0 × 10−5, 1.0 × 10−5, and 3.0 × 10−6 M) were used. This investigation aimed to check the accuracy of the new MWCNT-OSO3−TBZ+ sensor for TBZ determination. After measuring the potential in the TBZ solution, its concentration was calculated using a calibration graph (response of the sensor) and the corresponding equation of a line. This procedure was followed by determining TBZ using the Gran method [c = 2.0 × 10−3 M) in analyte solution (TBZ solution of known concentration) and by measuring the corresponding potential, which responds to TBZ according to Equation (4):k is constant, and c is the concentration of TBZ. After rearrangement, it follows:As aqueous samples, five TBZ solutions  and (6), and after rearrangement, it follows:After every standard addition of TBZ, its concentration can be calculated as follows:c = 1.0 × 10−5 M, three repeated measurements).After constructing a plot of Based on the results, it can be concluded that both direct potentiometry and the Gran method can be used for TBZ determination in real samples. Less accurate results, but still acceptable, were obtained in samples with lower TBZ concentration, which was expected considering that many factors can influence direct potentiometric measurements. The Gran method yields slightly more accurate results because it is based on multiple standard additions; thus, it was used to determine TBZ in oranges, lemons, bananas, clementines, and limes.c = 2.0 × 10−3 M) was added to each fruit sample. The resulting data are presented in Peels of oranges, lemons, bananas, clementines, and limes were used as real samples. The fruit was purchased from the local store and peeled. Then, 100 g of fruit peel was chopped and covered with distilled water, followed by a pH adjustment to 2.6. After 24 h and filtration, determination of TBZ using the Gran method was performed. The accuracy of TBZ determination and potential influence of the matrix were investigated using the standard addition method, where 5 mL of TBZ .A new potentiometric solid-state MWCNT-OSOeviously . In addi


Cu was biomagnified whereas Pb and Ni were biodiluted in offshore fish. Hg presented significant biomagnification in both of nearshore and offshore fish. Finally, the target hazard quotient of Hg (1.41) in MSCS fish exceeded the standard limit, which was posed by high Hg concentration and consumption rate of offshore fish.Seawater and fish were collected from nearshore  and offshore  regions of the South China Sea (SCS) to determine the heavy metals (HMs) pollution status and biomagnification characteristics. Results show that Cu in PRE seawater was moderately contaminated. Overall pollution risk of seawater were PRE (3.32) > MSCS (0.56), whereas that of fish was MSCS (0.88) > PRE (0.42). δ Heavy metals (HMs), which are distributed ubiquitously in both natural and anthropogenic regions ,2,3, areMarine ecosystems have long and complex food web structures, which may enhance the bioaccumulation ability of HMs in marine organisms ,12,13. T13C/12C = δ13C) is applied for identifying the dietary sources of marine organisms, and nitrogen (15N/14N = δ15N) is well known to determine the TL and investigate the biomagnification of contaminants for marine food webs  concentrations for sampled species of nearshore and offshore sampling sites, where HMi is the concentration of metal i in biological sample. The biomagnification factor (BF) of each HM was estimated with Equation (5), where a is the slope from the linear regression in Equation (4).A linear regression model, as shown in Equation (4), was used to fit the TLs and logi is the averaged concentration of HM i for each fish species (dw); M is the average moisture content in fish in different areas  [The estimated daily intake (EDI) of each HM via fish consumption was calculated using the following Equation (6) :(6)EDI=Cectively ); IR is  region) ; EF is eIn addition, the target hazard quotient (THQ) refers to the risk of noncarcinogenic effects. If THQ < 1, the exposure level is less than the reference dose . This sunce dose . Inorgannce dose . The RfDp < 0.05) of the correlation was assessed by the statistic software SPSS 19.0  for Windows system. HM concentrations in all samples were compared by the T-test method at a 5% significance level to describe statistics.Statistical significance (p < 0.05) concentrations of Ni (1.35–7.98 μg/L), Cu (6.44–10.1 μg/L), As (0.503–7.60 μg/L), Cd (<LoD—1.76 μg/L), Pb (<LoD—1.24 μg/L), and Hg (1.45—10.2 ng/L) compared to MSCS seawater. This work observed Ni, Cu, and Hg in MSCS seawater ranging from 1.21–1.66 μg/L, 0.935–1.49 μg/L, and 0.770–1.78 ng/L, respectively; whereas the concentrations of As, Cd, and Pb in all of the MSCS seawater were not detected (<0.001 μg/L). In this study, no HM concentration in seawater exceeded the Level II of China National Environmental Quality Standards for Surface Water , and theMean concentrations of Ni, Cu, As, Cd, Pb, and Hg in fish samples are listed in p < 0.05) (As shown in  < 0.05) . The ove13C, δ15N values and TL for fish collected in the PRE and MSCS are given in 13C values in PRE fish were from −24.77‰ to −16.44‰; whereas in MSCS fish, they ranged from −22.24‰ to −12.54‰. Significant differences (p < 0.05) of δ13C values were found between PRE and MSCS fish, which were MSCS (−16.67‰ ± 2.00‰) > PRE (−20.59‰ ± 2.43‰) (δ± 2.43‰) A.13C during trophic transfer, δ13C values are an efficient tool to track diet sources of aquatic organisms [13C in marine fish were widely reported in the Yellow Sea (−27.62 ‰ to −14.54 ‰) [13C. The variances observed in the stable isotope ratio of δ13C may predominately reflect significant differences of primary organic matter in the daily diets of fish from different geographical regions [13C values [13C values in PRE and MSCS fish may be attributed to the distinct carbon sources in nearshore and offshore areas. PRE fish possessed obviously negative δ13C values compared with MSCS fish, which suggested that more terrestrial organic matter was assimilated by fish living in nearshore regions, while MSCS fish displayed the relatively positive δ13C values, indicating the diet sources of marine organic carbon [Due to less enrichment or depletion of δrganisms . A wide 14.54 ‰) , and eve14.54 ‰) . By cont14.54 ‰)  and Taih14.54 ‰) , general regions . It has C values ,53. Hencc carbon .15N values of PRE fish  were significantly higher (p < 0.05) than those in MSCS fish , as shown in  [Scoliodon laticaudus sample in the PRE that exhibited δ15N value of 16.6‰. Both δ15N enrichment during trophic transfer and varied δ15N baseline values in the primary producer (mediated by varied primary nitrogen sources) are able to alter the δ15N values in aquatic food webs [15N baseline in the PRE (δ15N = 5.72) was about two times higher than that in the MSCS (δ15N = 2.69)  , except ood webs . This wo = 2.69) . Human N = 2.69) , the TL p < 0.05) in PRE fish, as well as Cu  and Hg  in MSCS fish, displayed significant biomagnification; while Ni  and Pb  of MSCS fish displayed an obvious decrease with TL ( with TL . Neitherp < 0.05). Positive correlations in environmental media generally represent similar sources of HMs [This study also presents the results of correlation analysis (CA) and hierarchical clustering (HC) of HMs and TL of fish in s of HMs . Howevers of HMs . Thus, ts of HMs ; and poss of HMs . HC analThe EDI and THQ for fish collected from the PRE and MSCS regions could evaluate the HM risks of nearshore and offshore fish intake, which are listed in This study demonstrates that the pollution profiles of HMs in seawater of the PRE and the MSCS are at a safe level, except for the moderate pollution of Cu in PRE seawater. The CPI values indicate that fish in the MSCS, rather than the PRE, bioaccumulate more HMs. Distinct C and N stable isotope ratios reveal the differences of habitat in marine fish from nearshore and offshore regions of the SCS. Compared to other HMs, Hg is the unique HM which exhibits biomagnification in both nearshore and offshore areas and is correlated with TL. Environmental behaviors of Hg in marine ecosystems need to be investigated in future. In addition, the THQ of Hg in offshore (THQ of Hg = 1.41) fish of the SCS have exceeded the safe limit. Given the significant biomagnification ability and potential health risks of Hg, we propose that further works need to pay more attention to tracing bioaccumulated Hg sources and developing consumption advisories of marine fish, to minimize the potential environmental impacts of Hg.