To describe the very early vault changes in the first month after Implantable Collamer Lens (ICL) implantation and to evaluate the effect of preoperative biometric factors on vault.Eighty-three eyes from eighty-three subjects with complete data who met follow-up requirements were recruited in this retrospective study between May 2019 and March 2020. We quantitatively assessed the postoperative vault at 2 h, 1 day, 1 week, and 1 month following implantation. Associations between the postoperative vault and age, ICL size, spherical equivalent (SE), axial length (AL), central corneal thickness (CCT), flat keratometry (K), steep K, mean K, anterior chamber depth (ACD), crystalline lens thickness (LT), white-to-white (WTW) diameter obtained by three devices, horizontal and vertical sulcus-to-sulcus (STS) diameter, bright and dark pupil sizes (BPS and DPS) and DPS-BPS were investigated using Spearman’s correlation analysis and stepwise multiple regression analysis.p < 0.001), followed by horizontal STS , crystalline LT  and vertical STS , significantly influenced the vault at 1 month after the operation. The multiple regression equation was expressed as follows: central vault (µm) = -1369.05 + 657.121 × ICL size- 287.408 × horizontal STS − 432.497 × crystalline LT − 137.33 × vertical STS (adjusted R2 = 0.643).The mean vault values at 2 h, 1 day, 1 week, and 1 month after ICL implantation were 672.05 ± 30.72, 389.15 ± 28.33, 517.23 ± 30.76 and 530.12 ± 30.22 μm, respectively. Significant differences were found in the vault values at 2 h, 1 day and 1 week after the operation. The ICL size (β = 0.942; After ICL implantation, the vault decreased and then increased, but it did not return to the vault value 2 h after surgery. The ICL size, horizontal and vertical STS and crystalline LT are key factors for predicting postoperative vaulting.The online version contains supplementary material available at 10.1186/s12886-021-01963-x. The Implantable Collamer Lens  is a safe and effective option to correct myopia –4. With Many factors can influence the vault after ICL implantation. Lee et al.  found thVery early vaulting changes, which are often used in contralateral eye surgery strategies, are often ignored by researchers. Therefore, the present study revealed the early change process in vaulting through observations from 2 h to 1 month after ICL implantation and established preoperative biometric factors that might contribute to vault formation and prediction.This retrospective study was approved by the Lixiang Eye Hospital of Soochow University Institutional Review Board and adhered to the tenets of the Declaration of Helsinki. All the patients were examined, treated and followed at the refractive surgery centre of Lixiang Eye Hospital between May 2019 and March 2020. Eighty-three eyes from eighty-three subjects with complete data were recruited for this study. Informed consent was obtained from each subject before surgery.2. None of the patients had ciliary body cysts, obvious cataracts, glaucoma or retinal disease history, or systemic diseases. One eye was randomly selected for the subjects who had surgery on both eyes.The inclusion criteria for this study included patients aged 18–45 years, patients with myopia between − 0.50 and − 21.00 DS, patients with astigmatism between 0 and − 6.00 DC, patients with an anterior chamber depth  equal to or greater than 2.80 mm, and patients with an endothelial cell density greater than 2000 cells/mmAll the patients had undergone complete ophthalmic examinations, including uncorrected and best corrected distance visual acuity evaluation, slit-lamp microscopy, tonometry , and fundus examination using a three-mirror lens. The refractive dioptre was measured using a standard phoropter and converted into the spherical equivalent (SE), which was calculated as the original spherical dioptre plus a half of astigmatism. Flat K, steep K, mean K, central corneal thickness (CCT) and ACD were obtained using a Scheimpflug camera . The bright and dark pupil sizes (BPS and DPS) were measured using an OPD-Scan III device . Axial length (AL) and crystalline lens thickness (LT) were obtained using an IOLMaster 700 . Ultrasound biomicroscopy  equipped with a 50-MHz transducer was performed to measure the horizontal and vertical sulcus-to-sulcus (STS) diameter after instillation of proparacaine . Horizontal corneal diameter and white-to-white distance (WTW) measurements were performed using a Pentacam, OPD-Scan III device and IOLMaster 700.All examinations were performed in a room with constant temperature and humidity controlled by an air conditioning system. Phoropter, Pentacam and OPD-Scan III examinations were conducted in the darkroom, and all other examinations were conducted under standard room lighting conditions. Each test was performed by the same experienced physician or technician.ICL implantation followed a standard procedure and was performed by the same experienced surgeon . After topical anaesthesia  was applied and hyaluronic acid  was injected into the anterior chamber via a 3.0-mm temporal corneal incision using an injector cartridge, an ICL V4c model (VICMO or VTICMO) was implanted and then placed in the posterior chamber. Next, the hyaluronic acid was completely removed from the eye using a manual Irrigation/Aspiration (I/A) instrument. All surgeries were uneventful, and no intraoperative complications were observed. Following surgery, tobramycin 0.3 % dexamethasone 0.1 % eye drops  were administered four times daily for the first 5 days, three times daily for the next 5 days and two for the last 5 days. The power calculations for the ICL were performed according to the manufacturer’s guidelines using a modified vertex formula . All ICLAll subjects had undergone vault measurement using a Pentacam at 2 h, 1 day, 1 week and 1 month after ICL implantation. The patient placed their chin on the chin rest and their forehead against the forehead strap and was asked to open both eyes and fixate on the blue fixation target in the centre of the black background. Fifteen Scheimpflug image enhancement models were used to obtain anterior segment images. The image quality was checked using the quality factor value for each eye. Two experienced technicians blinded to the treatment groups independently measured the centre vault value in the Pentacam Scheimpflug image using the device’s built-in image analyser. The vault measurement was centred on the optical axis and appeared as a white dashed line on the screen. Each technician obtained three measurements, which were averaged. If the difference between the two technicians was less than 30 μm, the average value of the six measurements was included in the analysis, and if the difference value was equal to or more than 30 μm, the measurements were repeated until the difference was less than 30 μm.SPSS 18.0  was used to perform the data analysis and the Kolmogorov-Smirnov test was performed for all measurement data. The data with a normal distribution were expressed as means ± standard deviation (SD); otherwise, the data were expressed as medians and quartiles. Repeated measures analysis of variance was used to calculate the vault change, and post hoc comparisons among time points were performed using Bonferroni correction. A paired sample t test was used to assess the correlation between the postoperative vault at 2 h and 1 day and postoperative vault at 1 month. Spearman’s correlation analysis and stepwise multiple regression analysis were used to examine associations between 1-month ICL vaulting and the other variables. The independent variables included age, ICL size, SE, AL, CCT, flat K, steep K, mean K, ACD, LT, WTW obtained using three devices, horizontal and vertical STS, BPS, DPS, and DPS-BPS. A p-value of less than 0.05 was considered significant.The average age of the subjects was 27.21 ± 5.07 (range: 18 to 38) years, and 32.53 % (27/83) of the subjects were male. Table Table p < 0.001) and that between vault values at 1 day and 1 month after surgery was 0.823 (p < 0.001). The paired difference between vault values at 2 h and 1 month after surgery was 141.93 ± 136.66 μm  and that between 1 day and 1 month after surgery was − 140.96 ± 159.36 μm .The paired sample correlation between vault values at 2 h and 1 month after surgery was 0.879 (p < 0.05). Table p < 0.001), the ICL size , horizontal STS  and vertical STS . The multiple regression equation was expressed as follows: central vault (µm) = -1369.05 + 657.121 × ICL size − 287.408 × horizontal STS − 432.497 × crystalline LT − 137.33 × vertical STS. The formula requires further validation. The R, R2 and adjusted R2 of the model were 0.814, 0.660 and 0.643, respectively. Figure According to Spearman’s correlation analysis, the vault at 1 month after surgery was positively correlated with ACD, WTW and the ICL size and negatively correlated with crystalline LT , indicating that the crystalline LT is an excellent predictive variable. Our results also showed that pupil size can influence the postoperative vault, but the vault cannot be predicted by the preoperative pupil size, including BPS, DPS or the difference between the two. According to our results, the vault value at 1 month after surgery was positively correlated with ACD, WTW and ICL size and negatively correlated with crystalline LT. However, we believe that such results are of little clinical significance because the ICL size was an important factor affecting postoperative vaulting. In our study, ICL size was not a continuous variable, so it had a significant impact on the results of Spearman’s correlation analysis. We are confident in the results of multivariate analysis. We found that the ICL size, followed by horizontal STS, crystalline LT and vertical STS, significantly influenced 1-month postoperative vaulting. Previous studies have shown that the ciliary sulcus is vertically oval –39. Becahttp://www.accessdata.fda.gov/cdrh_docs/pdf3/p030016c.pdf)). According to our results, neither WTW measured by any instrument nor ACD was a reliable predictor of postoperative vaulting, which was also the consensus of many similar studies [2 = 0.144), only one size of ICL was included in this study, and the axial direction of ICL placement was not considered, which would affect the results. Chen et al. [2 = 0.320 and 0.297) and small sample size (38 eyes for the V4 group and 39 eyes for the V4c group) make the results unsatisfactory. Recently, Igarashi et al. [2 (0.41) was still not completely satisfactory. The NK formula developed by Nakamura et al. [2 of the multiple regression was 0.68, and the adjusted R2 was 0.666. In subsequent validation, a moderate vault was achieved in 92.1 % of cases by applying the formula [2 of 0.666 vs. 0.643). The CLR and crystalline LT both describe the morphology of the lens, and the distance between scleral spurs and the horizontal and vertical STS describe the anatomical morphology of the posterior chamber. Additionally, the sample size of our study was larger (83 eyes vs. 46 eyes) than theirs, and the crystalline LT was easier to measure. To identify the presence of ciliary body cysts, UBM is an important preoperative examination before ICL implantation [Conventionally, the manufacturer’s recommendation for ICL size refers to only two parameters: WTW and ACD (Visian ICL Product Information: Visian ICL For Myopia. Available at  studies , 35, 42. studies  obtainedn et al.  developea et al. , 43 is lantation , 44. TheThere are certain limitations in this study. First, although the sample size was larger than those in some similar studies, it still must be supplemented in subsequent studies. Second, we only developed the prediction formula, and verification of the formula must be performed. We intend to publish later. Our study was conducted only among Han Chinese, and further study is needed to determine whether ethnic differences will impact the results. Finally, vault measurement may itself be variable because it is performed manually, and we did not measure the agreement of the interobservers. We used the method of averaging two independent measurers after the measured difference was less than 30 μm.In summary, we described very early changes in the ICL vault in the first month, starting at 2 h after ICL implantation, and we found that the ICL size, followed by the horizontal STS, crystalline LT and vertical STS, significantly influenced the vault 1 month after surgery. We hope our findings and new formulas will be helpful for surgeons when choosing the appropriate ICL size.Additional file 1.


The disability rate of spinal cord injury (SCI) is extremely high, and stem cell inhibition is one of the most effective schemes in treating the spinal cord, but the survival rate is extremely low after stem cell transplantation, so it cannot be widely used in clinic. Studies have revealed that loading stem cells with biological scaffolds can effectively improve the survival rate and effect after stem cell transplantation. Therefore, this research was devised to analyze the repair effect of thiolated chitosan nanocarriers scaffold carrying de-epithelized human amniotic epithelial cells (HAECs) on SCI. And we used thiolated chitosan as nanocarriers, aiming to provide a reliable theoretical basis for future clinical practice. Through experiments, we concluded that the Tarlov and BBB scores of rats with SCI were raised under the intervention of thiolated chitosan carrying HAECs, while the inflammatory factors in serum, oxidative stress reaction in spinal cord tissue, apoptosis rate of nerve cells, and autophagy protein expression were all suppressed. Thus, the thiolated chitosan carrying HAECs may be applied to treat SCI by suppressing autophagy protein expression, oxidative stress response, and release of inflammatory factors in spinal cord tissue, which may be a new clinical therapy for SCI in the future. Even though we cannot understand exactly the therapeutic mechanism of thiolated chitosan carrying HAECs for SCI, the real clinical application of thiolated chitosan carrying HAECs needs to be confirmed by human experiments. SCI is one of the primary causes of limb dysfunction . For theAs a kind of stem cell, HAECs have been proven to be effective in transplantation therapy for central nervous system diseases , 12. MorForty SPF SD rats, half male and half female, weighing 280–320 g, were from Zhuhai BesTest Bio-Tech Co., Ltd., with license number SYXK (Guangdong) 2020–0229. They were fed adaptively for one week. Rats were randomly assigned to 4 groups: transplanted HAECs , transplanted thiolated chitosan carrying HAECs , no processing , and control group (CG).2 for 90 s, and then the incision was closed. In CG, a microvascular clamp was not applied to treat the spinal cord, and the others were the same as abovementioned. A week later, rats in GA were transplanted with HAECs . The epithelial cell layer of HAECs was eliminated by trypsinase digestion, and 5 μL (2 cells/μL) cells were obtained and implanted into the transection site of model rats. In CG and MG, rats were implanted with the same amount of physiological saline. The chitosan (500 mg) was obtained and mingled with 8 mL of hydrochloric acid (1 mol/L). Then, the deionized water was supplemented to 50 mL, and then 1.38 g of 1-(3-dimethylaminopropyl)-3-ethyl carbodiimide hydrochloride was supplemented to continue. The pure thioglycollic acid (0.5 mL) was added and changed it to pH 5.0 with 2 mol/L sodium hydroxide. They were placed at ambient temperature for 6 hours in the dark, and then dialyzed with hydrochloric acid (5 mmol/L), hydrochloric acid (5 mmol/L), 1% sodium chloride, and hydrochloric acid (1 mmol/L), respectively, for 12 hours each time. Freeze-drying was applied to prepare the thiolated chitosan solid. The thiolated chitosan (1.25 mg) was added to the 5-5'-dithio-bis-nitrobenzoic acid solution (0.5 mg/mL) for 3 h. After that, it was centrifuged for 15 min  to obtain the supernatant, and the OD value (412 nm) was tested via an enzyme-labeling instrument. The morphology of thiolated chitosan was observed under a scanning electron microscope (SEM). HAECs and the prepared thiolated chitosan were cultivated for 24 hours, and then transplanted to rats in GB according to the scheme in 2.3. The concentration of HAECs was changed to 5 × 103/hole and inoculated it into 96-hole plate. After 24 h, the culture medium was replaced, the thiolated chitosan was supplemented, and cultivated for 24 h, 48 h, and 72 h, respectively. Next, 20 μL MTT solution (5 mg/mL) was added. After 4h, the supernatant was discarded and DMSO (150 μL) was supplemented. The OD value (570 nm) was tested via an enzyme-labeling instrument. The growth of HAECs cells with thiolated chitosan was compared with that without thiolated chitosan. In the three groups, rats were assessed by the Tarlov score [rd, 5th, and 7th days after transplantation. The higher the score, the better the situation.In GA, GB, and MG, rats were anesthetized with pelltobarbitalum natricum intraperitoneally and then fastened on the fixed mount. Next, the skin and muscle at T9-T10 level were carefully cut layer by layer, the neural plate at T9-T10 was excised, and the spinal cord was clamped with a microvascular clamp at a pressure of 20 g/cmov score  and the 3 for a subsequent test. ELISA was used to test inflammatory factors such as interleukin-6 (IL-8) and interleukin-10 (IL-10) and tumour necrosis factor-α (TNF-α) in rat serum, and the kits were all from Invitrogen, the States. Spinal tissue samples were collected from samples in 2.9. The contents of malondialdehyde (MDA) and superoxide dismutase (SOD) in tissues were tested via xanthine oxidation and thiobarbituric acid. The kit was purchased from Beijing Biolab Technology Co., Ltd. and Sigma-Aldrich, the States. Spinal tissue samples were collected from samples in 2.9. They were fastened in 4% paraformaldehyde for 72 hours and prepared into paraffin sections (5 μm). Then, it was dyed in accordance with the specifications of the TUNEL apoptosis kit  and counted under a microscope to calculate the apoptosis rate of nerve cells. Spinal cord tissue was put in RIPA lysate , and protein concentration was verified via BCA . SDS-PAGE electrophoresis was applied to transfer the film and seal it for 1 h. Beclinl , LC3 , P62 , and GAPDH primary antibody  were supplemented overnight. Then, the film was washed with TBST and HRP-labeled goat anti-rabbit IgG secondary antibody was added . ECL  was applied for detection, and the gray value was calculated.After transplantation for 7 days, all the rats were killed by cutting their necks under anesthesia. The abdominal aorta blood (5 mL) was drawn and centrifuged for 10 min  to obtain serum, which was put in a refrigerator at −80°C. In addition, the spinal cord tissue from the T9–T10 injured segment of rats was obtained and cut into small pieces of 1 mmt-test was applied for comparison between groups. Univariate analysis of variance and LSD back testing were applied for comparison among groups. P < 0.05 was considered significant.All results were represented as (mean number ± standard deviation) and analyzed by SPSS19.0. Independent sample P < 0.001, According to the test, the sulfhydryl group in 1 g of thiolated chitosan was approximately 214.52 ± 14.26, which was higher than in nonthiolated chitosan (26.42 ± 4.86) , (31.42 ± 7.42 pg/mL), and (52.62 ± 9.57 pg/mL), respectively. In GB, they were (11.87 ± 3.42 pg/mL), (22.42 ± 6.72 pg/mL), and (37.84 ± 6.65 pg/mL), respectively. In MG, they were (29.42 ± 5.63 pg/mL), (42.85 ± 10.52 pg/mL), and (84.63 ± 11.74 pg/mL), respectively. In CG, they were (9.62 ± 2.05 pg/mL), (12.84 ± 4.26 pg/mL), and (18.93 ± 4.71 pg/mL), respectively. The levels of IL-6, IL-10, and TNF-α in MG were the highest, followed by GA. The levels of IL-10 and TNF-α in GB were lower than those in GA but higher than those in CG  in GA, (14.83 ± 2.93%) in GB, (42.82 ± 5.63%) in MG, and (7.63 ± 1.18) in CG. Compared with the four groups, the apoptosis rate of nerve cells in MG was the highest, followed by GA, and GB was lower than GA but higher than CG  are becoming more and more complex and serious . As one Firstly, we tested the sulfhydryl groups in the prepared thiolated chitosan and found that the content of sulfhydryl groups in the thiolated chitosan increased obviously. Previous studies have revealed that the sulfhydryl groups can greatly destroy the intramolecular hydrogen bonds of chitosan, improve the water solubility of chitosan, and form disulfide bonds with the sulfhydryl groups of mucin molecules, thus increasing the mucosal adsorption effect of nanoparticles and the absorption rate of encapsulated drugs in vivo . TherefoThe nanodrug-carrying scaffolds of thiolated chitosan can ameliorate the survival rate of HAECs. The thiolated chitosan carrying HAECs may be applied to treat SCI by suppressing autophagy protein expression, oxidative stress response, and release of inflammatory factors in spinal cord tissue, which may be a new clinical therapy for SCI in the future.However, we cannot understand exactly the therapeutic mechanism of thiolated chitosan carrying HAECs for SCI due to the limited conditions, and the improvement of thiolated chitosan for HAECs' shortcomings may not only ameliorate the cell survival rate and the binding rate with the body, but also have a deeper and potential relationship between them, which is worth exploring and discovering. Nevertheless, the real clinical application of thiolated chitosan carrying HAECs needs to be confirmed by human experiments.


Administration of a single oral therapeutic dose (30 mL) of Mist Nibima corresponds to ingestion of 559.2 ± 7.662, 505.5 ± 6.930, 312.6 ± 4.285, 76.8 ± 1.028, and 51.0 ± 0.699 µg of methyl cryptolepinoate, cryptolepine, quindoline, 11-isopropylcryptolepine, and hydroxycryptolepine, respectively. This translates into a corresponding daily dose of 1677.6 ± 22.986, 1516.5 ± 20.790, 937.8 ± 12.855, 230.4 ± 3.084, and 153.0 ± 2.097 µg of methyl cryptolepinoate, cryptolepine, quindoline, 11-isopropylcryptolepine, and hydroxycryptolepine. These results could now serve as tools for authentication, standardization, and quality control of Mist Nibima to ensure its chemical and pharmacological consistency and safety.Mist Nibima is an essential herbal medicine used to treat malaria, bacterial, yeast, and COVID-19 infections. However, the drug has not been standardized and its active chemical ingredients are also not known. This study employed physicochemical, organoleptic, qualitative, and quantitate phytochemical analysis to established standards for Mist Nibima. Additionally, UHPLC was used to quantify the alkaloid cryptolepine in the drug using calibration curve. The chemical ingredients in Mist Nibima were thereafter characterized using UHPLC-MS. Organoleptic evaluation shows that Mist Nibima is a very bitter, cloudy, broom yellow decoction with the following physicochemical parameters: pH = 6.10 ± 0.08 (at 28.3°C), total solid residue = 5.34 ± 0.27%w/v, and specific gravity = 1.0099 ± 0.0000. The total alkaloid (23.71 ± 1.311%) content of the drug is 3 times its total saponins (7.993 ± 0.067%) content. Methyl cryptolepinoate (37.10%), cryptolepine (33.56%), quindoline (20.78%), 11-isopropylcryptolepine (5.16%), and hydroxycryptolepine (3.14%) were the active chemical ingredients in the drug with the concentrations of 18.64 ± 0.255, 16.85 ± 0.231, 10.42 ± 0.143, 2.56 ± 0.034, and 1.70 ± 0.023  Cryptolepis sanguinolenta produced by the Center for Plant Medicine Research (CPMR), Mampong-Akuapim, Ghana. C. sanguinolenta (Lindl.) Schlechter is a climbing, twining, perennial shrub which is a member of the Apocynaceae family. The plant is locally known in Ghana as Nibima or Kadze in the Twi and Ewe dialects, respectively + of C16H12N2 which is indicative of cryptolepine with a molecular mass (Mm of 232.28 g/mol) +. This represents the molecular formula of C18H14N2O2 with the molecular mass of 290.0 g/mol. This data agreed with the molecular weight of 290 and [M + H]+ of 291 reported for methyl cryptolepinoate [Compound 2 produced a fragment ion at m/epinoate . CompounThe mass spectrum of the second compound in Mist Nibima is shown in z at 275 represents [M + 2H + Na]+ which is indicative of a sodiated adduct of C16H12N20. This is indicative of hydroxycryptolepine which has a molecular mass (Mm) of 248 g/mol. The increase in Mm to 250 g/mol is due to the presence of the 18O isotope in hydroxyl group instead of 16O. Furthermore, the m/z observed at 259.2 in the spectrum is indicative of [M + 2H + Na]+ - CH3. These MS features observed for compound 3 confirmed that it is hydroxycryptolepine. Hydroxycryptolepine was isolated from C. sanguinolenta with [M + H]+ of 249 which led to Mm of 248 g/mol [Compound 3 produced at least 10 distinct peaks in its mass spectrum. The peak with m/48 g/mol .The mass spectrum of compound 4 is shown in z at 277. This represents the [M + H]+ of 11-isopropylcryptolepine which has a molar mass of 276 g/mol. Furthermore, the m/z at 259 and 245 represents [M -H-NH3]+ and [M-2H–2CH3]+, respectively, for 11-isopropylcryptolepine. Therefore, compound 5 was confirmed to be 11-isopropylcryptolepine [The mass spectrum of the 5th compound in Nibima is shown in tolepine .The names, RT, and concentrations of the five active chemical ingredients/compounds identified in the herbal drug using the UHPLC-MS characterization are presented in C. sanguinolenta root. Water is a very polar solvent due to its strong hydrogen bonds. Hence, the most polar compounds were to be extracted in larger quantities into the drug compared to the less polar ones. However, this principle is true for methyl cryptolepinoate, an ester which dissociate into a cation and anion when in water, but not for hydroxycryptolepine. This indicates that the amount of compound extracted by boiling water depended not only on the polarity of the compound, but also on the quantity of the compound present in the plant material. Hence, hydroxycryptolepine might be the smallest constituent produced by the batch of the plant used to produce the drug. The dose of Mist Nibima as indicated on the label is 30 mL to be taken orally, 3 times daily after meals. Based on the results in µg of methyl cryptolepinoate, cryptolepine, quindoline, 11-isopropylcryptolepine, and hydroxycryptolepine, respectively. This translates into a corresponding daily dose (30 mL three (3) times) of 1677.6 ± 22.986, 1516.5 ± 20.790, 937.8 ± 12.855, 230.4 ± 3.084, and 153.0 ± 2.097 µg of methyl cryptolepinoate, cryptolepine, quindoline, 11-isopropylcryptolepine, and hydroxycryptolepine.The result indicates that methyl cryptolepinoate with 37.10% composition was the major component of Mist Nibima. This was followed by cryptolepine with 33.56%. The remaining three components followed in the decreasing order of quindoline (20.78%), 11-isopropylcryptolepine (5.16%), and hydroxycryptolepine (3.14%). Mist Nibima is a decoction (boiling water extract) of C. sanguinolenta has been widely studied for its biological activities and chemical constituents. Phytochemical investigations of organic extracts and fractions of the plant led to isolation of about eighteen indolo quinoline alkaloids in addition to three other derivatives of cryptolepine. They are cryptolepine, isocryptolepine, cryptosanguinolentine, neocryptolepine, cryptotackieine, quindoline, cryptospirolepine, cryptolepicarboline, cryptomisrine, 11-isopropylcryptolepine, cryptolepinone, biscryptolepine, hydroxycryptolepine, cryptoheptine cryptoquindoline, cryptolepinoic acid, methyl cryptolepinoate, and ethyl cryptolepinoate [C. sanguinolenta including Mist Nibima were not known. This study has chromatographically and spectroscopically characterized the individual chemical compounds in Mist Nibima and also determined their concentration for the first time.epinoate , 15–20. The UHPLC chemical finger printing analysis has shown that Mist Nibima contains five compounds out of about eighteen alkaloids previously isolated from the plant, since the chemical fingerprint has shown that the drug contains five compounds (five peaks) with specific RTs. It is now possible to discriminate contaminants in any future batch of the drug or be able to tell if an additional peak detected in any future batch of Mist Nibima is being produced by the plant or is an adulterant by using this chemical fingerprint as a reference standard. The chemical finger print chromatogram obtained for Mist Nibima could, therefore, serve as reference standard in determination of adulteration and substandard batches of the drug in future productions.Furthermore, knowing the standard concentration of a drug makes it possible to distinguish between a substandard batch with significantly lower concentration of the active ingredients or a batch with significantly high concentrations of the active ingredients which may be toxic. These situations may arise as a result of lapses in the raw material sourcing and the manufacturing process. Since the respective compounds and their standard concentrations in Mist Nibima are now known, the necessary adjustments can be made to any batch which does not meet these standard concentrations of the active ingredients in order to bring it to the level of the reference concentrations of its active ingredients. For instance, the concentration of active ingredients in a lower concentrated batch of a herbal medicine can be increased to the standard levels by evaporations under controlled heat to a required volume or addition of a calculated amount of a high concentrated sample, whereas a batch with higher concentration of active ingredients of the drug can be diluted with calculated amount of water or low concentrated sample to reduce it to the standard concentration.The present study has established physicochemical, organoleptic, qualitative, and quantitative phytochemical standardization and quality control parameters for Mist Nibima, a herbal decoction used for the treatment of malaria, COVID-19, and many other diseases. Five indole alkaloids, namely, quindoline, methyl cryptolepinoate, hydroxycryptolepine, cryptolepine, and 11-isopropylcryptolepine, and their corresponding concentrations have also been identified as the active chemical constituents in Mist Nibima for the first time. These results will be used as tools for authentication, standardization, and quality control of this essential herbal medicine to ensure its consistent chemical and pharmacological characteristics and safety.


There was a significant reduction in with an increase in the cutting days (12.70–6.53% protein). Fiber increased (48.79–72.99% NDF) when fertilization treatments were included and cutting days increased. The elements that were included in fertilization  showed a higher foliar content and also presented a reduction in foliar content with growth of the plant. Treatments with fertilization showed a nutrient uptake increase for all the elements up to 60 days, where a reduction in uptake capacity was observed. King grass is a plant with a high nutrient uptake capacity and, therefore, with high biomass and nutrient production. This is an advantage since it can be used in multiple applications, such as animal feed, biofuel production, and as a substrate for biodigestion, among others.The research was conducted to determine the effects of cutting interval and fertilization on the nutritional quality, nutrient uptake, and biomass production of King grass. The experimental design was a randomized complete block, using 4 blocks and 8 treatments per block; treatments consisted of 4 ages of cutting , with fertilization and without fertilization. The results showed increases of up to 72,000 kg ha In this context, varieties of Pennisetum purpureum are highly used as an animal feed alternative mainly in the dry seasons, due to high biomass production and as a strategy to complement grazing so as to increase the load capacity and fill the shortage of quality pastures3.In tropical regions, cattle farming bases its feed on the production of pastures and forage because of its low cost of production and availabilityPennisetum purpureum) high biomass production has positioned it as an important forage material for renewable energy production4. Gallego determined that King grass is one of the plants with the greatest potential for butanol production thanks to its productive characteristics, biomass production, and solvent yield5. Cardona classified it as a high-performance perennial grass with the capacity to adapt to various soil conditions and a high potential for ethanol production, even showing higher yields than sugarcane bagasse6.King grass’s , finding that as the cutting days increased, fiber levels also increased and crude protein levels and digestibility of DM significantly decreased. Additionally, they found that the maximum values of crude protein in leaf were 10.1% at 90 cutting days and decreased to 5.6% at 180 days11. Consequently, studies should be carried out in each production area related to the use of fertilizers for the purpose of maximizing plant productivity based on agroclimatic and soil characteristics to obtain models that maximize yield12.Biomass production and nutrient concentration in forage is determined by its availability in soils, therefore, the interpretation of soil analyses and the application of fertilizers based on this interpretation is a determining factor in crop developmentThis research determined biomass production, nutrient concentration, and nutrient uptake from King grass based on cutting days and fertilization.(+) kg−1), low available phosphorus (16.2–18.4 mg kg−1), medium exchangeable potassium (0.21–0.24 cmol(+) kg−1), medium exchangeable calcium (3.52–3.71 cmol(+) kg−1), low exchangeable magnesium (0.42–0.47 cmol(+) kg−1), low available sulfur (5.47–5.81 mg kg−1), normal exchangeable aluminum content (0.21–0.28 cmol(+) kg−1), high available copper (4.16–4.82 mg kg−1), high available zinc (3.51–3.97 mg kg−1), medium available manganese (5.68–8.92 mg kg−1), high available iron (141–162 mg kg−1), and medium boron available (0.29–0.38 mg kg−1).The research was developed at La Esmeralda farm located in the municipality of Circasia Quindío, in the Colombian coffee region, located at 4° and 38′ 24″ north latitude and 75° and 38′ 26″ west longitude at an elevation of 1660 m, with an annual rainfall of 2400 mm and an average temperature of 19 °C. The study was carried out in an Andisol soil with an acidic pH (5.4–5.6), high organic matter content (8.6–8.9%), high cation exchange capacity (21.3–24.8 cmol2. Treatments (T) consisted of 4 ages of cutting , with fertilization and without fertilization. In the fertilization treatments, 3400 kg fertilizer per hectare-year distributed according to the area and cutting day was applied, the day after each cut. Fertilizer nutrient concentration was 23.6% N, 12.5% P2O5, and 19.3% K2O. T1, T2, T3, and T4 were designated as the cuts after 30, 45, 60, and 90 days, respectively, with fertilization, and T5, T6, T7, and T8 were the cuts at 30, 45, 60, and 90 days, respectively, without fertilization. Moreover, in T1, T2, T3, and T4, 27.9, 41.9, 55.9 and 83.8 g m2/cut of fertilizer were applied, respectively.A randomized complete block design with 4 blocks and 8 treatments per block was used, having a total of 32 experimental units per 40 m13. In addition, a regression analysis was performed to determine the relationship between biomass production, leaf-stem ratio, DM, crude protein, NDF, ADF, with cutting days. Analyses were performed with SAS and MATLAB R2021a packages.Experimental data were taken after the second cut to have a stabilization period of the treatments and 4 experimental cuts were made for each treatment. The analyzed variables were biomass production, leaf-stem ratio, dry matter (DM), crude protein, neutral detergent fiber (NDF), acid detergent fiber (ADF), foliar content, and nutrient uptake of N, P, K, Ca, Mg, Na, Cu, Zn, Mn, Fe, B and nutrient recovery efficiency. Data were subjected to a variance analysis (ANOVA). When there were differences (P < 0.05), Duncan’s multi-range test was used2 were taken per experimental unit, each square meter was individually weighed, and the biomass production and leaf/stem ratio were determined. Approximately 4 kg were collected per experimental unit for bromatological analysis, nutrient concentration, and nutrient absorption. Once the sample was taken, it was weighed and dehydrated at 60 °C for subsequent analysis. Soil analyses were carried out in each experimental unit at the beginning of the experimental process. Bromatological and soil analyses were performed using the Weende and Van Soest methodology, and for N the Kjeldahl methodology was used. Moreover, K, Ca, Mg, Mn, Na, Fe, Zn, and Cu were analyzed using atomic absorption spectroscopy, P through UV–Vis spectroscopy (Bray II), B using UV–Vis spectroscopy (modified Olsen), and organic carbon by Walkey and Black methodology. The pH was studied by the potentiometric method, cation exchange capacity by sodium chloride (titration), and the interchangeable aluminum extracted with 1 N potassium chloride (titration). Nutrient recovery efficiency was calculated using the Eq.  and has tended to stabilize , where the growth rate decreased until 60 days of cutting (T3), at which point biomass production was the highest , with increases up to sixfold in accumulated production. This indicated that King grass crops must be managed with fertilization systems.Bromatological contents of King grass were statistically different (P < 0.01) in each analyzed variable. Crude protein showed a progressive linear reduction when cutting days increased Fig. , being hThere was a linear increase in fiber with the age of cutting as a result of stem growth in treatments 2, 3, and 4; in treatments with fertilization, rapid plant development was observed  for all analyzed variables, except sodium (Na). Cutting age had a significant effect on mineral content; as the number of days increased, the plant´s mineral content was significantly reduced. Treatments with fertilization had a reduction in the concentration of minerals between 13.1 and 48.6% and treatments without fertilization between 9.7 and 31.3%. Elements incorporated into fertilization  had higher values on the same cutting days and showed increases of 17.9, 27.3, and 12.5% for N, P, and K respectively, demonstrating the effect of fertilization on the foliar contents of plants. Elements that were not incorporated into fertilization  had higher values in non-fertilization treatments where the plant was less developed  for all analyzed variables, with significant variations in fertilization and cutting days. The plant increased nutrient uptake up to 60 days in treatments with and without fertilization; in contrast, at 90 days, a decrease in nutrient uptake for all elements was observed and a reduction in nutrient production capacity was also observed. When fertilization was implemented, the grass showed a significant increase in its nutrient uptake capacity with absorption up to 3.4-fold higher. Although nutrient uptake in treatments without fertilization was lower, it remained high  , reported biomass productions of 5.88, 8.08, 10.68, and 13.42 ton of DM cut ha−1, respectively3. Uvidia15 obtained the same trend with cuts at 30, 45, 60, 75, and 90, reporting equally ascending values between 5 and 19 t ha−1 year−1. Data similar to those were found in non-fertilization treatments in this study, but significantly higher in treatments with fertilization. The high biomass production in the systems with fertilization gives King grass an important potential in the development of animal feeding systems, production of biofuels, composts and substrate for biodigestion, among others.Martínez and González evaluated six varieties of 3, in a study about King grass production with different cutting ages reported a leaf:stem ratio of 0.93, 0.85, 0.76, and 0.71 at 45, 60, 75 and 90 cutting days, respectively. Although they observed smaller variation than the ratio found in this study, the same trend was observed. Several authors concluded that this behavior was due to the maturity of the plant and its productive characteristics17.Luna3. These values were similar to those found in this study, indicating that nutritional quality in King grass is reduced with cutting age, while structural carbohydrates are significantly increased, which has been related to the increase in cellulose and hemicellulose production. Uvidia found the same trend in the relationship between protein and fiber content15.Luna also reported crude protein contents between 7.93 and 9.41, where the lowest value was obtained for a higher cutting agePennisetum purpureum with different cutting days, found a protein production per year between 3 and 3.7 t without statistical differences14. In this work, the production of protein per hectare year ranged between 2.22 and 10.75 t ha−1 year−1 and the higher value was found in T3 where cuts were made at 60 days with fertilization. Martínez and González also found Ca concentrations between 0.21 and 0.59% and P concentrations between 0.24 and 0.46%, values similar to those found in this study for the various treatments14.Martínez and González, in a study of 12 determined the effect of cutting age on mineral content in Pennisetum pedicellatum and found a reduction in mineral content, however, only Mg and Ca were significant (P < 0.05) with contents between 0.44 and 0.31% for Ca, similar to those found in this study, and contents between 398 and 278 mg kg−1 for Fe, which were higher than those found in this study. Zn contents ranged between 19.3 and 17.2 mg kg−1, which were lower than those found in this study. Similarly, Mn was between 54.0 and 80.2 mg kg−1, lower than those found here, except in treatment with fertilization and cutting at 90 days. Mg was between 0.52 and 0.36% higher than those found here, demonstrating that there is an important effect of cutting age and soils on nutrient concentration in plants.AsmarePennisetum clandestinum , and Lolium perenne . However, its high biomass production and carrying capacity  makes it a plant with a great potential to increase productivity and land use. The use of King grass in systems combined with forage plants of high nutritional quality  or its establishment in farms of small- and medium-sized beef cattle producers would allow achieving production models with high carrying capacity and adequate nutritional intake.King grass has low nutritional contents (low protein and high fiber contents) compared to high nutritional quality grasses such as Permanent cutting systems without fertilization lead to continuous nutrient uptake, resulting in a progressive loss in the soil's ability to provide nutrients to the plant and, consequently, a progressive reduction in biomass and nutrient production. Additionally, the application of fertilizers (NPK) stimulates the absorption of other nutrients, achieving a more than twofold increase in absorption after 60 days compared to treatments without fertilization. This guarantees high yields, but could also lead to soil fertility losses. For this reason, fertilizers should be formulated based on soil analysis and plant nutritional requirements Table  to promo22. Singh, in an experiment about the effect of nutrient omissions in wheat, found that maximum productivity and nutritional quality was achieved when a balanced mix of fertilizers  was applied, the omission of a nutrient had a direct effect by reducing concentrations and absorption of these elements in the crop23.Knowledge of plant nutritional requirements and development of fertilization systems based on these requirements ensures maximum crop productivity and sustainability over time−1; the highest values of N, P, and K uptake were 144.2, 53.5, and 142.6 kg ha−1, respectively, and the lowest values were 60.5, 14.4, and 63.4 kg ha−1, respectively24. Rosado, in a study on nutrient absorption, evaluated the effect of the application of three nitrogen fertilizers and six nitrogen doses  on Panicum maximum cv. ‘Mombasa’ macronutrient uptake and found that nutrient absorption had a significant increase with fertilization; N absorption increased 320 kg ha−1 year−1, and the same trend was found for P and K25. Those results were similar to those found in this study with an effect of fertilization on the absorption capacity and nutrient production of plants.Adeniyan studied the effect of the combination of N, P, and K on nutrient absorption and corn yield using a granular and foliar fertilizer in doses of 100, 80, 60, and 40 kg ha32; these values were lower than those found in this study, demonstrating again the very high capacity of the plant to take advantage of nutrients from fertilizers and soil and the importance of studying and understanding the mechanisms used by the plant to achieve these efficiencies.Studies carried out in different crops  in which nutrient recovery efficiency was determined, the highest efficiency reported for nitrogen, phosphorus and potassium was 91%, 38% and 88% respectively


Now solid renal tumors ≤4 cm is the most common, especially the subtype of clear cell renal cell carcinoma (ccRCC) of malignant kidney tumors in clinical. However, there is not specific characteristics of contrast‐enhanced ultrasound (CEUS) be recommended by the EFSUMB Guidelines in distinguish the essence of the kidney tumor with different sizes. Therefore, this meta‐analysis aimed to assess the ability of CEUS to diagnose solid ccRCC (sccRCC) ≤4 cm. We comprehensively searched the Cochrane Library, Embase, PubMed, and Web of Science databases from their inception to 28 July 2020, for studies reporting the CEUS features of sccRCC lesions ≤4 cm. Additional articles were identified through the Chinese National Knowledge Infrastructure database. Studies were selected independently by two investigators and the relevant data were extracted. Discrepancies were resolved via discussion with the senior author. Study quality was assessed using the Quality Assessment of Diagnostic Accuracy Studies‐2 tool, and the sensitivity and specificity of each study were determined and plotted as a receiver operating characteristic curve. Ten studies were included in this meta‐analysis. Hyperenhancement showed medium sensitivity (67%–89%) and specificity (42%–75%) for diagnosing sccRCC ≤4 cm, fast‐in contrast agent and heterogeneous enhancement showed high diagnostic abilities (area under curve (AUC) 0.74–0.84), but the presence of a pseudocapsule and fast‐out contrast agent had poor diagnostic ability (AUC <0.70). The combination of hyperenhancement and iso‐enhancement showed high sensitivity (98%) for diagnosing sccRCC ≤4 cm. Hyperenhancement, fast‐in contrast agent, and heterogeneous enhancement may be specific features that could help to identify sccRCC ≤4 cm, while the presence of a pseudocapsule and fast‐out of contrast agent may have low diagnostic values. The combination of multiple indexes may improve the diagnostic value of CEUS for sccRCC ≤4 cm. Now solid renal tumors ≤4 cm is the most common, especially the subtype of clear cell renal cell carcinoma (ccRCC) of malignant kidney tumors in clinical. However, there is not specific characteristics of contrast‐enhanced ultrasound (CEUS) with the EFSUMB Guidelines be recommended in distinguish the essence of the kidney tumor with different sizes. Therefore, this meta‐analysis aimed to assess the ability of CEUS to diagnose solid ccRCC (sccRCC) ≤4 cm. CT and MRI have high diagnostic capability in detecting renal tumor, but they do not apply to all the crowd for ionizing radiation (CT) or exorbitant fees (MRI). CUS provides a fast, safe, repeatable, and cost‐effective method for revealing the basic characteristics of lesions, such as location, size, shape, border, echo, and blood supply. However, it does not recognize tumor microflow and detection rate of small renal tumors is lower than CT, especially those less than 2 cm.2The study did not require approval from the institutional Research Ethics Board. The study protocol has been published in PROSPERO . The study was conducted in accordance with the QUORUM reporting guidelines.The study addressed the following issues, according to the “PICOS” framework: P (patients): adults (age >18 years) with a solid renal mass ≤4 cm, possibly representing a ccRCC2.1A literature search was performed using the Embase, PubMed, Web of Science, and Cochrane databases from their inception to 28 July 2020, with no language restrictions. The keywords included: “clear cell renal cell carcinoma,” “ccRCC,” “renal clear cell carcinoma,” “contrast‐enhanced ultrasound,” “contrast‐enhanced ultrasonography,” “contrast ultrasonography,” “ultrasound contrast imaging,” and “CEUS.” Additional studies were identified through the Chinese National Knowledge Infrastructure database. Titles and abstracts were reviewed and eligibility was determined independently by two investigators with disagreements resolved by consensus, and full‐text versions of relevant studies were retrieved for further evaluation.2.2The inclusion criteria were as follows: (1) adults with sccRCC ≤4 cm; (2) CEUS implemented before clinical intervention; (3) data retrievable to calculate a 2 × 2 contingency table; (4) acceptable reference standard (pathology) used for all patients; (5) CEUS characteristics of the mass described qualitatively or quantitatively; and (6) study patients not a subset of patients from another included paper. In the event of overlap between study samples, the latest results were used.The exclusion criteria were as follows: (1) studies with fewer than 30 cases; and (2) conference papers or secondary literature, such as experience exchanges, abstracts, lectures, and reviews.2.3All retrieved articles were exported into Endnote 9.3.3, and all duplicate studies were removed. Each article was assessed independently by two investigators, and eligible studies were recruited according to the selection criteria. The investigators extracted the relevant data into a data extraction table (Microsoft Excel), and any discrepancies were discussed with a third reviewer to reach an agreement. The following data were gathered: first author's surname, year of publication, geographical location, language, ethnicity, sex, age, number of sccRCCs and lesions other than sccRCCs ≤4 cm, lesion length, study design, CEUS instrument, contrast agent type and dose, mechanical index, “gold standard,” true positive (TP), true negative (TN), false positive (FP), and false negative (FN) CEUS characteristics. The definitions were as follows: (1) hyper‐, iso‐, and hypo‐enhancement at peak lesion enhancement were defined according to the lesion‐enhancement degree compared with the renal cortex; (2) “fast‐in” and “slow‐in” indicated an arrival time of the contrast agent in the lesion before or after its arrival in the adjacent renal cortex, and “fast‐out” and “slow‐out” indicated the relative outflow of the agent from the tumor, respectively; (3) homogenous enhancement was defined as a lesion with full enhancement without any defects, and heterogeneous enhancement was defined as a lesion with some unenhanced areas; (4) a rim of perilesional enhancement was considered to represent the pseudocapsule; (5) TP and FN were CEUS characteristics of sccRCC ≤4 cm, TP was present, FN was absent; and (6) FP and TN were CEUS characteristics of lesions other than sccRCC ≤4 cm, FP was present, TN was absent.2.4The Quality Assessment of Diagnostic Accuracy Studies‐2 (QUADAS‐2) tool2.5I2 tests were used to evaluate the potential heterogeneity between studies. I ≤25% indicated low heterogeneity, >25% to ≤50% indicated mild heterogeneity, >50% to ≤75% indicated moderate heterogeneity, and I2 > 75% indicated high heterogeneity. If I2 > 50%, a random‐effect model was used.p < 0.05 indicated statistical significance.All data were calculated using STATA 15.1 and Review Manager 5.3 software. Quality assessment was performed using Review Manager 5.3. The following indexes were calculated to assess the diagnostic ability of the CEUS characteristics: sensitivity, specificity, diagnostic odds ratio (DOR), negative likelihood ratio (NLR), positive likelihood ratio (PLR), area under the curve (AUC) with 95% confidence intervals (CIs), summary receiver operating characteristic curve (SROC), and percentage of patients with the CEUS characteristics. The 2 × 2 data were summarized in forest plots of sensitivity and specificity for each CEUS characteristic. Sensitivity and specificity on a per‐feature basis were plotted in SROC space. Cochran's Q‐statistic and 33.1A flow diagram of the study selection procedure is presented in Figure The assessment of the methodological quality of the included articles using the QUADAS‐2 tool is depicted in Figure The inappropriate exclusion of patients and lack of reviewer blinding to the final pathologic diagnosis were the main contributors to overestimation of diagnostic accuracy. The remaining QUADAS‐2 domains were all considered to have low risk of bias and low concern for all studies.3.23.2.1p < 0.05, I2 = 82.57%) and specificity  of heterogeneous enhancement showed significant heterogeneity, with a combined sensitivity of 0.67 (95% CI: 0.51–0.79) and a combined specificity of 0.75 (95% CI: 0.45–0.92). The SROC with pooled AUC was 0.75 (95% CI: 0.71–0.78). The summary PLR and NLR were 2.7 (95% CI: 1.1–6.3) and 0.44 (95% CI: 0.32–0.62), respectively, the DOR was 6 (95% CI: 2–17), and the pooled OR was 5.882 (95% CI: 2.168–15.968).The sensitivity  and specificity  also showed significant heterogeneity, with a combined sensitivity of 0.70 (95% CI: 0.49–0.84) and a combined specificity of 0.67 (95% CI: 0.45–0.84). The SROC with pooled AUC was 0.74 (95% CI: 0.70–0.77). The summary PLR and NLR were 2.1 (95% CI: 1.4–3.3) and 0.45 (95% CI: 0.30–0.69), respectively, the DOR was 5 (95% CI: 3–9), and the pooled OR was 5.118 (95% CI: 2.758–9.496).For hyperenhancement, the sensitivity  and the specificity showed significant heterogeneity . The combined sensitivity was 0.89 (95% CI: 0.81–0.94) and the combined specificity was 0.42 (95% CI: 0.18–0.69). The SROC with pooled AUC was 0.84 (95% CI: 0.80–0.87). The summary PLR and NLR were 1.5 (95% CI: 0.9–2.5) and 0.27 (95% CI: 0.11–0.63), respectively, the DOR was 6 (95% CI: 2–21), and the pooled OR was 5.991 (95% CI: 1.711–20.978).The sensitivity of fast‐in contrast agent showed slight heterogeneity  and specificity  of the presence of a pseudocapsule both showed significant heterogeneity, with a combined sensitivity of 0.54 (95% CI: 0.26–0.79) and a combined specificity of 0.63 (95% CI: 0.46–0.77). The SROC with pooled AUC was 0.63 (95% CI: 0.59–0.67). For the absence of a pseudocapsule, the sensitivity  and specificity  also showed significant heterogeneity, with a combined sensitivity of 0.46 (95% CI: 0.21–0.74) and a combined specificity of 0.37 (95% CI: 0.23–0.54). The SROC with pooled AUC was 0.37 (95% CI: 0.33–0.41). For fast‐out contrast agent, the sensitivity showed significant heterogeneity  and the specificity showed mild heterogeneity , and the combined sensitivity was 0.62 (95% CI: 0.42–0.78) and the combined specificity was 0.28 (95% CI: 0.18–0.41). The SROC with pooled AUC was 0.34 (95% CI: 0.30–0.39). The sensitivity for homogeneous enhancement showed slight heterogeneity , the specificity showed significant heterogeneity , the combined sensitivity was 0.41 (95% CI: 0.33–0.50), and the combined specificity was 0.19 (95% CI: 0.05–0.54). The SROC with pooled AUC was 0.35 (95% CI: 0.731–0.39). The summary PLR, NLR, and DOR were all low for all the above features.The sensitivity  showed slight heterogeneity and the specificity  showed low heterogeneity. The DOR was 5 (95% CI: 3–9), the pooled OR was 10.522 (95% CI: 5.106–21.682), and the summary PLR and NLR were low.Hyper‐iso‐enhancement showed high sensitivity  and poor specificity . The SROC with pooled AUC was 0.56 (95% CI: 0.52–0.60) for hyper‐iso‐enhancement (≤0.70). The sensitivity  was observed in two studies published in 2016, and slight heterogeneity  was also found in two studies that focused on lesions within 4 cm. Regarding fast‐in contrast agent, slight or significant heterogeneity was seen in all aspects. For fast‐out contrast agent, slight heterogeneity  was found in six studies in Asian patients, slight heterogeneity  was observed in six studies using SonoVue, and significant heterogeneity  was detected in four studies focusing on lesions within 3 cm. Regarding the presence/absence of a pseudocapsule, there was significant heterogeneity in two studies published in 2016, three studies published in English, three studies in Asian populations, two studies of lesions within 3 cm, and six studies using SonoVue. For heterogeneous enhancement, slight heterogeneity was observed in five studies in English, four studies in Asians, two studies focused on lesions within 3 cm, and five studies using SonoVue. For homogeneous enhancement, there was slight heterogeneity in four studies in English , four studies in Asian populations , three studies focusing on lesions within 4 cm , and five studies using SonoVue . The above results identified these factors as potential sources of heterogeneity.Subgroup analyses were conducted based on the year of publication, language, ethnicity, lesion length, and contrast agent type to investigate potential sources of heterogeneity. There was no significant heterogeneity for hyperenhancement in any of the aspects. For hyper‐iso‐enhancement, slight heterogeneity  encodes an oxygen‐sensing mediator protein (pVHL) in the hypoxia signaling pathway.VHL,The mechanism of angiogenesis in ccRCC is shown in Figure EphB4/ephrin‐B2 is widely involved in physiological and pathological angiogenesis during embryonic and postnatal development, and is considered to be molecular markers of arteriovenous development.The sccRCC characteristics of hyperenhancement, heterogeneous enhancement, and fast‐in contrast agent depend on the structural and functional abnormalities of the tumor blood vessels. The current analysis indicated that hyperenhancement, fast‐in contrast agent, and heterogeneous enhancement had high diagnostic ability (AUC ≥0.70) for sccRCC lesions ≤4 cm, with sensitivities of 70%, 89%, and 67% and specificities of 67%, 42%, and 75%, respectively. These results were in accordance with previous studies that reported “hyperenhancement,” “fast‐in,” and a “peritumoral hyperenhanced rim (pseudocapsule)” as CEUS characteristics of ccRCC.4.2The presence of a pseudocapsule is related to tumor size, as confirmed by Atri et al.Most studies indicated that ccRCC demonstrated fast‐out contrast agent properties. However, the current analysis suggested that fast‐out contrast agent had a poor detection ability, with a sensitivity of only 0.62, low specificity, and poor overall diagnostic ability (AUC 0.34). The apparent discrepancy between these results and those of others studies may be related to the intra‐tumoral heterogeneity. The fast‐out loss of contrast agent occurs via arteriovenous fistulas or by normal physiological reflux of renal veins. However, intra‐tumoral heterogeneity derived from clonal and sub‐clonal tumor cells may affect the differentiation degree and malignancy level of the lesion,5This study has several limitations. First, we did not include any gray literature or unpublished studies, because these were usually unavailable. Second, most of the included studies were from China and most of the study populations were Asian, with very few international studies including non‐Asian populations. This may have affected the results, and, further studies are needed to confirm the validity of the results in different populations. Third, the study data were from different regions and affiliations, and the CEUS images were evaluated by clinicians with different experiences, potentially affecting the diagnostic results. Finally, the number of included studies was small, making it impossible to perform some subgroup meta‐regression and sensitivity and specificity analyses, thus highlighting the need for more studies in the future. More research is also needed to examine the quantitative parameters in sccRCC ≤4 cm, and to explore the qualitative and quantitative parameters in lesions >4 cm. Due to the inability to extract multiple CEUS features from the same lesion, we were unable to analyze the combined diagnostic values of multiple features for sccRCC lesions ≤4 cm, and we aim to address this issue in further studies.Hyperenhancement, fast‐in contrast agent, and heterogeneous enhancement may have high diagnostic ability for sccRCC lesions ≤4 cm. Hyperenhancement and iso‐enhancement showed high sensitivity for diagnosing sccRCC ≤4 cm, suggesting that the combined index may be particularly useful for detecting these lesions. In contrast, presence of a pseudocapsule and fast‐out of contrast agent might not aid the diagnosis of sccRCC lesions ≤4 cm.The authors declare that they have no conflict of interest.Institutional Review Board approval and informed consent from the patient are not required because this is a systematic review of published literature.


The activity and selectivity performances toward pollutant removal of the electrode materials were investigated by electrochemical measurements and constant potential electrolysis, suggesting that Ti electrode exhibited the best NO3−/NO2− reduction and N2 production efficiencies. In-situ Fourier transform infrared spectroscopy was used to study the in-situ electrochemical information of pollutants conversion on electrode surfaces and propose their reaction pathways. The effects of main operating parameters  on the removal efficiencies of COD and TN were studied. Under optimal conditions, COD and TN removal efficiencies from simulated wastewater reached 92.7% and 82.0%, respectively. Additionally, reaction kinetics were investigated to describe the COD and TN removal. Results indicated that COD removal followed pseudo-first-order model; meanwhile, TN removal followed zero-order kinetics with a presence of NH4+ and then followed pseudo-first-order kinetics when NH4+ was completely removed. For actual pharmaceutical wastewater treatment, 79.1% COD and 87.0% TN were removed after 120 min electrolysis; and no NH4+ or NO2− was detected.In this work, an electrochemical method for chemical oxygen demand (COD) and total nitrogen  removal from wastewater using a divided electrolysis cell was developed, and its process optimization was investigated. This process could effectively relieve the common issue of NO A large quantity of wastewater produced from industrial manufacturing process is contaminated by two often-regulated primary contaminants: organic matter and nutrients, such as nitrogen in the forms of ammonia, nitrate, and nitrite ,2. Biolo4+ and proved the superiority of electrochemical wastewater treatment [3− or NO2− have been reported [3O4/Ti, Fe2O3/Ti, and Cu cathodes on NO3− removal, and Co3O4/Ti exhibited better NO3− removal efficiency.Electrochemical method is proposed as a promising alternative for wastewater treatment because of its high efficiency, versatility, easy operation, and environmental compatibility ,15,16. Greatment ,19. For reatment  reportedreported ,22. Su ereported  presente4+, NO3−, and NO2−. In our previous work [4+ over-oxidation on anode and NO3−/NO2− over-reduction on cathode. Actually, many literatures indicate that the over-oxidation or over-reduction is a common phenomenon in the process of nitrogen-containing wastewater treatment by electrochemical method [3− and NO2−. Li at al. [3− reduction with Fe cathode, and the concentration of NO3− decreased from 100.0 to 7.2 mg-N·L−1 in 3 h, but 51.1 mg-N·L−1 NH4+ was generated as the by-product. In order to relieve the issue of over-reduction/oxidation, it is essential to find a suitable reaction cell and key operating parameters to perform the balance between cathodic reduction and anodic oxidation. Li et al. [Current studies for wastewater treatment by electrochemical method mainly concentrate on the removal of a single pollutant or two mixed pollutants, and there are few investigations on the electrochemical treatment of wastewater containing COD, NHous work , the feal method ,25,26, wl method  studied i at al.  investigi et al.  and Mandi et al.  reportedi et al. ,32 also 3−/NO2− removal. The in-situ electrochemical information of COD/NH4+ oxidation and NO3−/NO2− reduction on electrode surfaces were explored by in-situ Fourier transform infrared spectroscopy (in-situ FTIR). The key operating parameters, i.e., initial pH value, chloride concentration, and current density, were studied to perform cathodic reduction of NO3−/NO2− and anodic oxidation of COD/NH4+. Additionally, kinetics and reaction mechanism were also proposed to describe the redox process. Finally, an actual pharmaceutical wastewater was investigated to verify the electrochemical performance for COD and TN removal.In this work, an electrochemical process for COD and TN removal was optimized, and its feasibility to relieve the issue of over-oxidation or over-reduction was evaluated. Ti, Cu, and stainless steel were selected as cathode material to study their reduction performance for NO−1 Na2SO4 solution. Sodium chloride was added into the solution to maintain the concentration of chloride ion. Actual pharmaceutical wastewater, which was characterized by low biodegradability and complex composition, was collected from a pharmaceutical factory . The wastewater characteristics are summarized in Simulated wastewater containing COD and TN was prepared by dissolving ethyl acetate, ammonium sulfate, sodium nitrate, and sodium nitrite into 0.1 mol L2 electrode (10 cm × 10 cm) was used as anode, while Ti, Cu, and stainless steel were selected as cathode, respectively. A volume of 600 mL wastewater was circulated between anode and cathode chamber by a peristaltic pump.Electrochemical experiments were conducted in a divided electrolysis cell . Ti/PbO2Electrochemical measurements  and constant potential electrolysis were executed for the choice of a proper cathode with an electrochemical workstation. The different cathodes were used as the working electrode; platinum sheet and saturated calomel electrode (SCE) were employed as the counter and reference electrode, respectively. In-situ FTIR spectra were obtained by a FTIR spectrometer equipped with a self-made spectroelectrochemical cell .COD, TN, and ammonia were determined by the dichromate, alkaline potassium persulfate, and Nessler reagent spectrophotometry, respectively ,13. Anio2, Ti/IrO2, Ti/RuO2, and BDD were compared for treatment of typical industrial wastewater, suggesting that Ti/PbO2 anode was suitable for COD and NH4+ removal [3−/NO2− removal efficiencies due to their high stability and low cost.In the electrochemical system for COD and TN removal, both anode and cathode materials are conclusive for optimizing the redox process . The inv removal ,36. Simi−1. Results indicated that Ti electrode had a hydrogen evolution potential of −1.542 V, which was more negative than those of Cu (−1.532 V) and stainless steel (−1.466 V), meaning that a side reaction of hydrogen evolution was least likely to occur on Ti electrode. 3−/NO2− reduction. Obvious cathodic peaks and extended broad waves were observed with the addition of NO3−/NO2− into the blank solution, which meant the direct NO3−/NO2− reduction occurred. Cathodic peaks of Ti, Cu, and stainless steel were at about −1.10, −0.99, and −0.82 V for NO3− reduction, respectively. For NO2− reduction, cathodic peaks of −0.68 and −0.81 V were observed for Ti and stainless steel (not observed on Cu), respectively.3−/NO2− reduction with three electrodes at a constant potential of −1.26 V are shown in 3− reduction was obtained with the Ti cathode, while that with stainless steel was the lowest. The inset of 2− and NH4+ were the intermediates or by-products during the NO3− reduction process [2 selectivity of these three electrodes was Ti (63.3%) > Cu (48.9%) > stainless steel (31.5%). This result might be related to the relative hydrogen evolution potential shown in 4+ as the NO3− reduction by-product was favored in a potential region closing to the hydrogen evolution reaction region; in other words, more negative hydrogen evolution potential led to higher N2 selectivity. The information regarding NO2− reduction from wastewater with different cathodes as working electrode is shown in 2− reduction for Ti, Cu, and stainless steel were 10.46, 8.67, and 5.96 mg-N in 5 h; the N2 selectivities with these three electrodes were 76.6%, 68.4%, and 51.8%, respectively. According to the results shown in 3−/NO2− reduction decreased in the following order: Ti > Cu > stainless steel; the Ti electrode was therefore selected for subsequent experiments to study the optimal reaction conditions.The removal and selectivity of NO process ,38. The To explore solid/liquid interfacial phenomena, i.e., identify the intermediates, adsorbed species, and products during pollutants removal, in-situ FTIR spectroscopy was applied in the studies of oxidation and reduction processes . In the −1 during COD electro-oxidation. Two negative infrared (IR) bands are observed at about 1271 and 1709 cm−1, representing the C−OH and C=O scissors vibration in COOH, respectively. The downward band at 2314 cm−1 is assigned to CO2 absorption. The results indicated that the organic matter was decomposed to low molecule acid and then converted to CO2.As shown in 4+ are detected at 1597 and 3005 cm−1. The weak bands located at 980 and 1666 cm−1 represent the absorption of NH3,ads. Besides, dispersed bands are observed around 2000–3000 cm−1. According to the study by Griffiths and Haseth [3+, NH2+, and NH+. A band at 1238 cm−1 can be also observed, indicating the generation of NO2− [2− band was observed, the vibration of NO3− was not shown. This phenomenon might be related to the operating conditions. For example, Kapałka et al. [4+ electro-oxidation because of the concentrations of generated active radicals, which was reported by Anglada et al. [4+ electro-oxidation in this system was proposed as shown in Equation (1):d Haseth , the com of NO2− . Though a et al.  indicatea et al. . On the 3− are given in −1 is ascribed to the existence of NO3−. The peak of 1265 cm−1 suggests the presence of NO2− in electrolysis. Dispersed bands are presented at ca. 1800–3000 cm−1, indicating the vibrations of NH2+ and NH3+ [−1 are observed and assigned to absorbed NH3,ads, and the weak band at 1485 cm−1 is confirmed as NH4+. As shown in 2− appears at 1230 cm−1. Similar to the spectra of NO3−, NH3,ads generated at the bands of 1014, 1639, and 3116 cm−1 in the electro-reduction of NO2−, and then, the NH4+ is detected at 1408 cm−1. Additionally, weak bands at about 1843 and 2854–2924 cm−1 are discovered with electrolysis time, which are ascribed to volatile by-products of NO and NO2, respectively [4+ oxidation, nitrate/nitrite reduction and products distribution also highly rely on operating parameter, such as optimum voltage, electrode distance, and initial pH [3−/NO2− electro-reduction mechanism was speculated in Equation (2), according to the FTIR spectra.The in-situ FTIR spectra measured during electro-reduction of NOand NH3+ . Three cectively . Similaritial pH . In thisAs stated above, operating parameters can make great influences on the performance of electrochemical process; moreover, actual wastewater is known for its characteristics of complexity and variety. Therefore, main factors, including initial pH value, chloride concentration, and current density, were investigated on COD and TN removal.3−, NO2−, and NH4+ removals. As shown in 3− and NO2− reduction than neutral or alkaline condition due to the H-atom adsorbed on the surface of electrode, which was necessary for the NO3−/NO2− indirect reduction [4+ was negative, suggesting that more NH4+ was generated in electrolysis rather than being removed. This could be explained by the over-reduction of NO3−/NO2−; i.e., NH4+ was generated as by-product in the process, indicating that the side reaction of over-reduction was the fundamental reason for limiting the TN removal in this electrochemical system. Taking into account the above results, initial pH value of 6 was selected for the further experiments.eduction . Howevereduction . Moreove− concentration varied from 0 to 1500 mg·L−1. The results indicated that, as Cl− was added, the removal of COD improved from 77.4% (0 mg·L−1·Cl−) to 88.6% (250 mg·L−1·Cl−), then increased slightly when further increasing Cl− concentration. The TN removal efficiency increased from 18.1% to 26.0%, 46.7%, 82.0%, and 87.0% with the increase of Cl− from 0 mg·L−1 to 250, 500, 1000, and 1500 mg·L−1, respectively. Besides, COD removal at different Cl− concentrations followed pseudo-first-order kinetics. Meanwhile, the process of TN removal matched a zero-order model as NH4+ existed and then fitted on pseudo-first-order kinetics as NH4+ removed completely. According to the removal efficiency and rate constant, it seemed that Cl− concentration of 1000 mg·L−1 was suitable for the next investigations.−2; meanwhile, the TN removal efficiency increased from 72.6% to 86.0%. −2, the corresponding rate constant for COD removal increased from 1.269 × 10−2 to 2.471 × 10−2 min−1; and rate constant for TN raised from 6.032 to 7.918 mg·L−1·min−1. Though increasing current density was conducive to pollutant removal, the energy consumption would increase obviously. Current density of 10 mA·cm−2 was thus selected as the optimal one.4+ generated during NO3−/NO2− reduction was circulated into anode chamber by loop operation and then oxidized to N2 on anode; meanwhile, the NO3−/NO2− as the over-oxidation by-product of NH4+ could be recirculated into the cathode chamber and continuously reduced to N2. This process could relieve the common issues of over-reduction and over-oxidation by combining cathodic NO3−/NO2− reduction with anodic COD/NH4+ oxidation and thus improve the TN removal.Considering to literatures and experimental results, a mechanistic model of electrochemical treatment of COD and TN containing wastewater in a divided electrolysis cell is proposed in −1 with the removal efficiency of 79.1%. Moreover, similar to the simulated wastewater treatment, COD removal in actual wastewater treatment followed pseudo-first-order kinetics. 4+ or NO2− was detected after 120 min treatment, while NO3− was removed from 28.33 to 15.69 mg·L−1. The concentrations of nitrogen oxides were under the detection limit. In addition, 4+, TN removal followed zero-order kinetics model; however, when NH4+ was completely removed, pseudo-first-order kinetics was fitted. After 120 min electrolysis, the dissolved concentration of Pb2+ was measured to evaluate the safety of the treated wastewater; it was found that Pb2+ concentration was 0.008 mg/L, which met the Standard for Drinking Water Quality in China (plumbum ≤ 0.01 mg/L). Beside, with removal of 79.1% COD and 87.0% TN, the energy consumption was calculated as 13.3 kWh·m−3, which was much lower than that of traditional electrochemical method reported by other authors (To further investigate the feasibility for COD and TN removal by this electrochemical method, an actual pharmaceutical wastewater was sampled and treated in this study.  authors . Meanwhi− concentration, and current density were investigated on the COD and TN removal. At an optimized pH 6, Cl− concentration of 1000 mg·L−1, and current density of 10 mA·cm−2, this method could remove 92.7% COD and 82.0% TN in simulated wastewater using Ti cathode and Ti/PbO2 anode. Besides, kinetics and reaction mechanism were also proposed to describe the electrochemical process of COD and TN removal. For treatment of an actual pharmaceutical wastewater, the extents of COD and TN removal were 79.1% and 87.0%, respectively.This work optimized the electrochemical treatment of COD and TN containing wastewater with a divided cell. The effects of cathode material, initial pH value, Cl


Prior studies found that participants overestimated both negative and positive emotional stimuli, compared with neutral emotion. This phenomenon can be explained by the “arousal mechanism.” Participants demonstrated individual differences in emotion perception. In other words, high emotional awareness resulted in high emotional arousal, and vice versa. This study extended existing findings by exploring the influence of emotional awareness on time perception in a temporal generalization task, while recording electroencephalographic (EEG) signals. The findings revealed that in the positive emotion condition, the high emotional awareness group made more overestimations, compared with the low emotional awareness group. However, no difference was observed in the neutral or negative emotion conditions. Moreover, the event-related potential (ERP) results showed that in the positive emotion condition, the high awareness group elicited larger vertex positive potential (VPP) amplitudes, compared with that of the low awareness group. However, no such differences were observed in the neutral and negative emotion conditions. Moreover, the contingent negative variation (CNV)  component showed that in the positive emotion, the amplitudes of the high awareness group were larger than that of the low awareness group; however, they did not show differences in the neutral condition. The findings of this study suggest that high emotional awareness produces higher physiological arousal; moreover, when participants were required to estimate the time duration of emotional pictures, they tended to make higher time overestimation. Thus, our results support the relationship between emotional awareness and time perception. Time perception differs from the accuracy of the clock, and it is a subjective temporal experience, which is influenced by emotion. A number of studies showed that emotional arousal is an important influencing factor of time perception  technique. The neural mechanisms were primarily analyzed by the vertex positive potential (VPP) and contingent negative variation (CNV) components.Vertex positive potential is differently modulated by emotional and neutral faces; it is a positive deflection detected at the frontocentral electrode, with peak latency between 140 and 200 ms , compared with the low awareness group; however, no differences would be observed in the neutral condition. Furthermore, we generated the following hypotheses related to the ERP result are as follows: (1) for the VPP component, the emotional (positive/negative) amplitude of the high awareness group would be significantly larger than that of the low awareness group; (2) for the CNV component, the emotional amplitude of the high awareness group would be significantly larger than that of the low awareness group. Moreover, for both VPP and CNV, there would be no differences between the high and low awareness groups in the neutral condition.*power  = 22.24, standard deviation (SD) = 0.69], with 21 participants each in the high  and low awareness groups . The LEAS score of the high awareness group  was significantly higher than that of the low awareness group  . All of them had normal or corrected-to-normal vision and also no neurological or psychiatric disorders.First, we estimated our sample size with the effect size of 0.25 and power of 0.8 , selected from the standard Chinese Facial Affective Picture System , and the screen resolution was 72 pixels per inch. All stimuli were displayed at the center of the screen (17-inch).The experiment was administrated in a quiet chamber. Further, the participants were at a distance of ~50 cm from the computer monitor. We recorded the participants' responses using the E-Prime software . Participants performed a temporal generalization task . The statistical method of “underestimation respond ratio” is the same.The EEG was recorded from 64 scalp sites using Ag/AgCI mounted in an elastic cap . The EEG and electrooculography were amplified using a direct current (0.05 Hz) with ~100 Hz bandpass and continuously sampled at 500 Hz/channel. All interelectrode impedance was maintained below 5 kΩ. ERP averages were computed offline; trials with artifacts were rejected with a criterion of ± 80 μV. ERP waveforms were time locked to the onset of the stimulus, and the average epoch was 1,000 ms, including a 200 ms prestimulus baseline. Recording electrodes were referenced to the left mastoid and recalculated offline to an average mastoids reference. The vertical electrooculogram signal was measured both above and below the left eye, whereas horizontal electrooculogram signal was measured on both sides at the external canthi. Eye-blink artifacts were rejected automatically using vertical ocular correction. All channels were filtered at 0.1–30 Hz offline. Incorrect response trials were removed.Similar to previous studies  = 13.16, p < 0.001, η2= 0.25. Post hoc analysis with Bonferroni correction showed that time overestimation of the positive condition (0.184 ± 0.01) was greater than the neutral condition (0.166 ± 0.01), p = 0.002, whereas that of the negative emotion condition (0.190 ± 0.01) was greater than the neutral condition, p < 0.001. There were no significant differences between the positive and negative emotion conditions (p = 0.51). The interaction between emotion valence and awareness group was significant, F = 8.58, p < 0.001, η2 = 0.18. Simple-effect analysis showed that in the positive emotion condition, the high awareness group overestimated time more than the low awareness group, F = 4.86, p = 0.03, η2 = 0.11. There were no significant differences between high and low awareness groups in the neutral or negative emotion conditions (ps ≥ 0.98); emotional awareness had no main effect (p = 0.41).For the overestimation response ratio as dependent variable, a mixed rmANOVA with the between-factor awareness group (2: high/low) and the within-factor emotion valence  revealed a main effect of emotion valence, ps ≥ 0.36) or interactive effects (p = 0.99).For the underestimation response ratio as the dependent variable, a mixed rmANOVA with the between-factor awareness group (2: high/low) and the within-factor emotion valence  did not reveal either significant main effect  = 26.51, p < 0.001, η2 = 0.40. Bonferroni post hoc test showed that the amplitudes of the negative emotion (7.21 ± 0.50 μV) were larger than that of the positive emotion condition (6.42 ± 0.47 μV), p = 0.01, and that of the positive emotion condition was larger than that of the neutral condition (5.80 ± 0.52 μV), p = 0.01. The electrode factor also resulted significant, F = 12.72, p < 0.001, η2 = 0.24; VPP amplitudes at Fz (6.68 ± 0.51 μV) were larger than F1 (6.39 ± 0.47 μV) and F2 , whereas F1 and F2 showed no significant difference (p =1.00). The interaction between emotional awareness and emotion valence was significant, F = 5.50, p = 0.008, η2 =0.12. Simple-effect analysis showed that in the positive emotion condition, VPP amplitudes in the high awareness group (7.24 ± 0.66 μV) were larger than in the low group (5.60 ± 0.66 μV), F = 3.06, p = 0.048, η2 = 0.07. There were no significant differences between the high and low awareness groups in the neutral or negative emotion conditions (ps ≥ 0.12). The interaction between emotional awareness and electrodes was significant, F = 6.81, p = 0.003, η2 = 0.15. Simple-effect analysis showed that for the low group, VPP amplitudes at Fz (5.98 ± 0.71 μV) were larger than at F2 (5.68 ± 0.67 μV), p = 0.002; however, no other significance differences were observed. For the high awareness group, VPP amplitudes at Fz (7.38 ± 0.71 μV) were larger than F1 (6.86 ± 0.67 μV), p < 0.001; VPP amplitudes at Fz were larger than F2 (7.05 ± 0.67 μV), p = 0.001. No other significant main or interactive effects were observed (ps ≥ 0.09). As shown in The mixed rmANOVA with the between-factor awareness group (2: high/low) and the within-factor emotion valence  and electrodes  revealed a main effect of emotion valence, F = 30.98, p < 0.001, η2 = 0.44. Post hoc analyses using Bonferroni correction showed that the CNV amplitudes for negative emotion (1.88 ± 0.59 μV) were larger than in the positive emotion condition (1.07 ± 0.58 μV), p = 0.001, whereas the CNV amplitudes (200–300 ms) in the positive emotion condition were larger than in the neutral condition (0.26 ± 0.53 μV), p < 0.001. The interaction between emotion valence and awareness group was significant, F = 11.25, p < 0.001, η2 = 0.22. Simple-effect analysis showed that in the positive emotion condition, the high awareness group (2.33 ± 0.82 μV) elicited larger CNV amplitudes (200–300 ms), compared with the low awareness group (−0.19 ± 0.82 μV), F  = 11.25, p = 0.035, η2 = 0.11. The high and low awareness groups showed no differences in the neutral or negative emotion conditions (ps ≥ 0.34); moreover, no main effects of awareness group were observed (p = 0.21). As shown in Average amplitudes of CNV components , measured by a mixed rmANOVA with the between-factor awareness group (2: high/low) and the within-factor emotion valence , revealed a main effect of emotion valence, F = 90.05, p < 0.001, η2 = 0.69. Post hoc analyses showed that CNV amplitudes (300–490 ms) in the negative emotion condition (0.66 ± 0.49 μV) were larger than in the positive (−0.32 ± 0.48 μV) or neutral conditions (−0.26 ± 0.49 μV), ps < 0.001. The interaction between emotion valence and awareness groups was significant, F = 14.61, p < 0.001, η2 = 0.27. Simple-effect analysis showed that in the positive emotion condition, CNV amplitudes (300–490 ms) of the high awareness group (−1.23 ± 0.68 μV) were larger than that of the low awareness group (0.51 ± 0.68 μV), F = 2.87, p = 0.038, η2 = 0.07. In the negative emotion condition, the high awareness group (1.19 ± 0.66 μV) elicited larger CNV amplitudes (300–490 ms), compared with the low awareness group (0.13 ± 0.66 μV), F =11.87, p = 0.049, η2 = 0.06. In addition, the high and low awareness groups showed no difference in the neutral emotion condition (ps = 0.93), and the awareness group had no main effect (ps = 0.33). As shown in A mixed rmANOVA with the between-factor awareness group (2: high/low) and the within-factor emotion valence  and CNV components  revealed a main effect of emotion valence, This study used a temporal generalization paradigm to explore the relationship between emotional awareness and time perception, through behavior and ERP components. The behavior results revealed that the time overestimation of the high and low awareness groups had no difference in the neutral and negative emotion conditions, but showed significant difference in the positive emotion condition. The ERP results showed that in the positive emotion condition, the high awareness group elicited larger VPP amplitudes, compared with that of the low awareness group. The high and low awareness groups had no difference in the neutral and negative emotion conditions. The CNV  results showed that in the positive emotion condition, the high awareness group elicited larger amplitudes, compared with that of the low awareness group. However, for the negative emotion condition, the high awareness group elicited larger CNV (300–490 ms) amplitudes, compared with that of the low awareness group. Whereas, the high and low reported no significant differences in the neutral emotion condition of the CNV components, our results indicated that the high and low awareness groups had a time overestimation difference on emotional pictures.Our behavioral results show a time overestimation difference in the emotion (mainly positive emotion) conditions between the high and low awareness groups. These findings are consistent with our hypothesis and can be explained by the arousal-based time perception hypothesis. Arousal and attention are the main factors affecting time perception  condition. The VPP component is an index for distinguishing emotional and neutral expression; the amplitude of the emotional stimuli is larger than that of neutral stimuli  showed that in the emotion (mainly positive) condition, the amplitudes of the high awareness group were larger than that of the low awareness group. This may suggest that CNV amplitude was not only an index of emotional arousal, but also of the accumulation mechanism and time overestimation, especially at the FCz electrode site .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.