Carassius auratus) as a suitable experimental model, this study tested this as a null hypothesis. Under laboratory conditions, the deterrence thresholds of individual goldfish exposed to 120 ms tones at six frequencies (250–2000 Hz) and four Sound Pressure Levels (SPL 115–145 dB) were quantified. The deterrence threshold defined as the SPL at which 25% of the tested population startled was calculated and compared to the hearing threshold obtained using Auditory Evoked Potential and particle acceleration threshold data. The optimum frequency to elicit a startle response was 250 Hz; different from the published hearing and particle acceleration sensitivities based on audiograms. The difference between the deterrence threshold and published hearing threshold data varied from 47.1 dB at 250 Hz to 76 dB at 600 Hz. This study demonstrates that information obtained from audiograms may poorly predict the most suitable frequencies at which avoidance behaviours are elicited in fish.Deterrents that use acoustics to guide fish away from dangerous areas depend on the elicitation of avoidance in the target species. Acoustic deterrents select the optimum frequency based on an assumption that highest avoidance is likely to occur at the greatest sensitivity. However, such an assumption may be unfounded. Using goldfish ( Freshwater vertebrates, which constitute a third of the world’s total4, are suffering population declines at twice the rate of those observed in marine or terrestrial ecosystems3. There are multiple explanations for the rapid3 and widespread deterioration of freshwater environments, including habitat degradation and loss, such as wetland drainage5; channelization6; invasive species7; climate change8; changing land use ; reduced river flows10; declining water quality11; novel pollutants ; and the cumulative effects of multiple stressors15. Centuries of river engineering, in particular, has severely damaged freshwater ecosystems, with many rivers channelised and impounded, reducing longitudinal, lateral and vertical connectivity and fragmenting fluvial habitat16.As the most threatened ecosystems on the planet17. Focusing on technology, there are a range of environmental impact mitigation solutions that may partially ameliorate some of the worst effects of river engineering16. For example, fish passes are designed to enable fish to negotiate river infrastructure, such as weirs and dams18, while physical screens are intended to prevent entrainment of fish into water intakes (e.g. at hydropower stations or cooling water systems), and in some instances to guide them to alternative routes, such as bypass systems19. The efficacy of environmental impact mitigation technology can be highly variable with site, context, and species20, as illustrated for both fish passes21 and physical screens, which themselves can be damaging if poorly designed23. Recently, efforts have been directed at improving the effectiveness of this technology. This has included a return to fundamental first principles and a reductionist approach to better understand the mechanisms that determine efficiency . At the same time, some have sought to take a more applied approach to combine technologies with the hope that efficiency will be improved if synergistic effects are realised (e.g. acoustics and light27). In some cases, behavioural deterrents have been developed with the view to using them in combination with traditional physical screening systems to enhance overall screening and guidance efficiency .Regeneration of freshwater ecosystems requires the integration of social, financial, legal and technological approaches30), elicited by fish in response to a stimulus resulting in modification of the movement trajectories of a target species. A wide variety of stimuli, mostly abiotic, have been employed as deterrents, including electricity31, strobe lights32 and acoustics . Sound can be advantageous in some contexts because the stimulus is omnidirectional, and so can simultaneously reach a number of target individuals within a locality and is not affected by changes in illumination or turbidity as are those that are mediated through vision. As a consequence, acoustic deterrents have been widely employed to reduce fish mortality at water abstraction points35, or to control the spread of invasive species .Behavioural guidance systems take advantage of the innate behaviours, typically aversive  and from 1141 to 87.9%35 for European Sprat (Sprattus sprattus). There are many proposed explanations for such inconsistent efficiencies, including the use of acoustic cues with frequencies outside the hearing range of the target species42, or because the ambient sound levels were not accounted for39. Alternatively, the results may reflect a lack of consideration of the relationship between hearing abilities, defined by the frequency range of the stimulus and hearing sensitivity of the individual, and the behavioural response of the target species39. Whilst it is impossible to know the hearing and deterrence thresholds of every individual fish, if we could quantify the intraspecific variation observed in the population, we could design deterrents that meet conservation targets43. In the past it was assumed that, for different frequencies, the difference between the hearing threshold and the sound pressure level (SPL) that elicits a behavioural response is uniform, and that this difference may be expressed in dB above the hearing threshold (dBht)44. This logic suggests that, for a fixed stimulus level, the frequency that is most likely to engender a response is the one at which the animal’s hearing is most sensitive, i.e. where the sensory threshold is lowest. Such an assumption is not evidence-based and has been dismissed among some researchers46. Nevertheless, some industry practitioners utilise the most sensitive hearing level to predict the behavioural response of fish when developing behavioural deterrents, including the use of audiograms in the design of deterrents for invasive carp47. To advance the next generation of acoustic deterrents that may be used in combination with other, perhaps more traditional environmental impact mitigation technology, there is a need to return to fundamental first principles.A limitation of acoustic deterrents is that their effectiveness can be highly variableCarassius auratus) were selected as the model species due to the large amount of physiologically derived information available on their hearing capabilities49 and their ease of maintenance under laboratory conditions. This study used the startle response exhibited by fish when exposed to an acoustic stimulus as a proxy for a deterrent response, enabling definition of a threshold to inform acoustic deterrence .This experimental study examined the relationship between frequency of hearing sensitivity and the deterrence thresholds of fish. Goldfish (This study determined the: (1) presence and absence of startle responses to determine the proportion of the population that exhibited avoidance and, then, the relationship between probability of startling and frequency  and SPL ; (2) deterrence threshold for each frequency defined as the SPL at which at least 25% of the sampled population elicited a startle response, a proxy for deterrence; and (3) relationship between the deterrence threshold, the hearing threshold, and the particle acceleration (PA) threshold at each frequency based on existing data obtained from audiograms for the subject species.z = 6.084, p ≤ 0.001), and within each frequency treatment increased with SPL for 250 Hz , 400 Hz  and 600 Hz , but not for 800 Hz , 1000 Hz  or 2000 Hz .Startles were observed in all frequency treatments. The probability Fig.  of start−2, respectively.At least 25% of the test population startled at frequencies that ranged from 250 to 800 Hz. A higher SPL  . The small sample size for the PA ΔThreshold meant only visual inspection was conducted. Visual inspection of the plot , and the ΔThreshold at 400 Hz was lower compared to 600 and 800 Hz (p < 0.001).The 47. Using a well-studied model species (goldfish) for which considerable information on hearing capability is available49, and the startle reaction as a proxy measure of deterrence, we found no evidence to support this assumption. Instead, one-quarter of all fish tested in the 250 Hz treatment startled in response to an acoustic stimulus of 123 dB re 1 μPa. This frequency is lower than that at which hearing is most sensitive, i.e. approx. 600 Hz  and deterrence thresholds indicated that the two ran parallel when plotted against frequency on a logarithmic dB scale52, in contradiction to the results of our study. In humans, however, the thresholds of hearing and loudness discomfort, a measure of behavioural intolerance, often occur at dissimilar frequencies53 i.e., the usable dynamic range (the dB difference between the hearing threshold and the threshold for some adverse reaction) is not independent of frequency. Towards the high frequency upper limit of human hearing, there is evidence that this usable dynamic range becomes so small that adverse effects are seen in some individuals at the lowest SPLs at which they can perceive the sound58. Additionally, humans sometimes finding irritating noise  at frequencies lower than those at which hearing is most sensitive.Comparisons between hearing and deterrence thresholds in fish are limited. There are few examples of attempts to define deterrence thresholds, with the exception of those used to determine the hearing ability of larval fishes60. In agreement with our findings, these authors also observed that the difference between hearing and reaction thresholds varied with frequency. However, they found that the eight subject species they studied reacted very differently, making generalisations difficult, and perhaps unwise. In our study, we selected a freshwater species with specialised hearing and a high sensitivity to sound62. Goldfish may be more reactive at the lower end of the hearing range because they are more sensitive to particle motion at 125–250 Hz63. At frequencies below 400 Hz64, there may be functional overlap of the neuromast and the otolith65. As both the otolith and the neuromast detect particle displacement, the relative contribution of the two to hearing is challenging since the inputs for auditory and lateral line nerves lie in close contact66. This means the electrophysiological techniques to measure hearing are likely to detect both inputs67. However, we would expect this same mechanism to be contributing to the electrophysical audiogram as well as the behavioural methods applied in this study. The importance of particle motion associated with low frequency sound fields has previously been considered in the development of behavioural deterrents, particularly in respect to the protection of European eel (Anguilla anguilla)68.The fish observed in this study were more responsive at the lower frequencies within their hearing range. Our findings complement those of another study that investigated the behavioural reaction threshold of fish in situ, focusing on marine species in the context of impacts of anthropogenic noise46 was contradicted by the results reported here, the implications for future fish deterrent design should be considered. It is no longer valid to accept the premise that the frequency of highest probability of reaction corresponds with the greatest sensitive of hearing or that the difference (ΔThreshold) between the two remains constant independent of frequency. The observation that ΔThreshold varies with frequency supports the consensus within the scientific community45 that suggest that the continued use of the concept of dB above the hearing threshold (dBht)43 used to inform infrastructure projects70 is inappropriate. The use of an arbitrarily defined fixed level above the basic hearing threshold has been used as a convenient method to set criteria for acceptable sound levels for other animals, such as marine mammals71 when knowledge is limited. Our findings support the arguments of others74 that the logic that underpins the use of dBht may be flawed, at least in the context described here, and may provide an explanation, at least in part, for why the efficiency of acoustic deterrents can be highly variable39.As the current assumption about the relationship between detection of a sound and response to it75 is reiterated, rather than developing design criteria based on data obtained from physiological methods alone. Quantification of thresholds of reaction is more appropriate from a fisheries management perspective, as avoidance is the often desired response in fish guidance system. Therefore, approaches such as those based on Acoustically Evoked Behavioural Response (AEBR)61 as used in this study, rather than AEP derived audiograms, are most appropriate because they determine the lowest SPLs over a range of frequencies at which a reaction is elicited. Second, once appropriate frequencies are identified there is a need to select SPLs that evoke the response desired accounting for ambient environmental conditions at the site of interest, i.e. considering appropriate signal-to-noise-ratios. Furthermore, other acoustic parameters should be characterised, including spatial distribution and temporal patterns. Third, this study adopted a novel experimental approach that resolves several of the challenges associated with small tank experiments76. By submerging the test cylinder in which the fish were constrained in a large tank, a more homogeneous sound field was created with less boundary reflections that result in unnatural heterogeneity in particle motion and sound pressure. A reductionist experimental approach such as that described here is valuable when there is a need to quantify fine-scale behaviours of the fish in response to rigorously mapped acoustic stimuli while controlling for confounding variables before moving on to field tests of prototype devices. Finally, we recommend that future research is directed at quantifying variability as a result of abiotic (e.g. the hydrodynamic environment77) and biotic factors, particularly between species and developmental stage or size60 with the view to identifying appropriate representatives guilds of commercial and conservation concern. Likewise, greater understanding is needed of how response to acoustic signals may differ between species that are typically solitary and those that occupy positions within groups .Moving forward, this study highlights factors that should be further considered in advancing the design of acoustic fish deterrents. First, the importance of behavioural studies in understanding the response of fishes to sound3 of aerated, filtered and dechlorinated water under an artificial photoperiod matching the light levels at the time of year (10:14 h light:dark) and fed once daily . Ammonia (0.10 [0.22] ppm), nitrites (0 [0] ppm), nitrates (40.0 [0.0] ppm), pH (pH 8.20 [0.00]) (API Freshwater Master Test Kit) and temperature (12.3 [1.2] °C) were monitored daily. Fish were acclimated in the holding tank for at least four days before 12 individuals were selected and moved to the experimental facility (< 700 m) the night prior to the start of trials where they were maintained in a pre-test tank  containing ≈ 0.29 m3 of aerated and dechlorinated water for a further 13 h to acclimate to the temperature of the experimental tank (temperature: 13.15 [0.49] °C). On completion of each trial, the subject fish  were placed in a post-test tank and returned to a separate holding tank at the ICER Facility at the end of the day.Goldfish  were transported from Hampshire Carp Hatcheries (UK) in oxygenated water to the International Centre for Ecohydraulics Research (ICER) facility, University of Southampton, in November 2020. They were maintained in a holding tank  containing ≈ 1.2 m78. Individuals were handled with care, and handling time was kept to a minimum. There was no evidence of stress or fatigue from exposure to the acoustic stimulus in any of the treatments during the trials. The authors complied with the ARRIVE guidelines.Experiments were carried out in compliance with guidelines established by the current UK animal protection law established by the Home Office . All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. The study was reviewed by the Animal Welfare and Ethics Review Body and approval granted by the University of Southampton Ethics and Research Governance committee (ID: 54900.A1). We reduced the number of goldfish used in our study by using the 25% threshold and we minimised distress by using a relatively low stimulus SPL−1; Logitech Pro, Switzerland) installed directly above the water surface, ensuring that the entire cylinder was maintained within the field of view. The room was lit by fluorescent lighting that provided sufficient illumination for video recording.Trials were conducted in a white medium density polyethylene cylindrical tank  suspended from a bespoke metal frame into a large water filled tank  at the A. B. Wood Laboratory, University of Southampton  and four SPLs  (6 × 4 = 24). Prior to the start of each trial (10–19 November 2020), a single fish (N = 80) was acclimated in the experimental cylinder for 30 min. Fish experienced a total of six exposures (one at each test frequency selected at random) at one of the four randomly assigned test SPLs  using a laptop connected to a DAQ , transmitting the signal through an amplifier , and was emitted via the UW30 underwater transducer. Acoustic stimuli were standardised such that the desired SPL was reached in the centre of the experimental arena. Use of artificial stimuli allowed for control of the specific acoustic components tested.a, was calculated as:ρ is the ambient density and P is the pressure80.Prior to exposing fish to stimuli, the acoustic environment of the experimental arena was quantified. A total of 246 measurements were made using a calibrated hydrophone (Brüel & Kjær 8105) to produce a 3D representation of the SPL in the experimental arena . Following this the PA was represented in maps  of the pressure difference was calculated independently in three directions . The pressure gradient was obtained by dividing by the distance between measurements. The RMS PA Eq. , in eachThe measured ambient SPL  was on average less than 96 dB re 1 µPa, which was the electrical noise floor of the measurement system being used. The SPL was relatively uniform across the horizontal plane for each frequency, with greatest variation observed at the highest frequency (shortest wavelength) at 2000 Hz . The initial GLMM contained all predictor variables with exposure (order of stimulus exposure) and trial included as random effects, and manual backwards selection using variable significance (significance at p < 0.05) was undertaken as model simplification. No random effects were detected . Logistic regression was performed using general (GLM) and generalized linear models (GLMM) with a binomial error structure and a “logit” link function. To determine whether external factors may have confounded the results by influencing the probability of startling, a reductive model was developed. Factors included in the model were: tank days (minimum number of days in the holding tank); time ; experimental tank temperature (°C); and size  of the population startled at 600 Hz. The SPL and PA at which 25% of the population startled at each frequency was predicted using the GLM. The output stated the probability that a startle occurred for a stated SPL (or PA). Both the SPL and PA were chosen and refined until the output was within 0.001 or 0.25.63.The proportions obtained by prediction using the GLM, quantified as startles per total number of individuals tested at each treatment, were represented on a plot of SPL against frequency and a plot of PA against frequency. Both 1000 Hz and 2000 Hz were omitted since extrapolation was unfeasible due to an insignificant logistic relationship, and predictive models estimated the threshold to be at a level damaging to the ear. The 25% deterrence threshold was plotted on one figure alongside the hearing threshold for goldfish, obtained by averaging AEP sound pressure hearing threshold data from 23 studies (Supplementary Table ΔThreshold was defined as the difference between thresholds for hearing and 25% startle response. The sound pressure ΔThreshold was calculated by subtracting published AEP hearing threshold values from the 25% deterrence threshold at each frequency. The same method was used for the PA ΔThreshold such that the published PA hearing threshold values were subtracted from the 25% deterrence threshold at each frequency. To assess the influence of frequency on threshold range, a GLM with Gaussian error structure was applied for both SPL and PA ΔThreshold. Planned orthogonal comparisons were used to determine whether the SPL ΔThreshold at 250, 400, 600 and 800 Hz differed from each other. This was not undertaken for PA, as there was only a single data point for each frequency since the PA threshold was based on a single study.The difference between the deterrence and the hearing thresholds were calculated and followed a Gaussian distribution, determined by visual inspection of the qq curve. The Supplementary Information.


Propyrisulfuron is a novel pyrimidinylsulfonylurea herbicide with good activity for controlling annual weed in rice fields. To evaluate the economic performance of propyrisulfuron, a field study was conducted in 2021 and 2022 on a farm of the Jiangsu Academy of Agricultural Sciences, China. Eight different herbicide treatments were employed, including CB , Py (propyrisulfuron), CBPy , PrBe 3, PrBe 10, and PrBe 3+PrBe 10 (pretilachlor plus bensulfuron applied at different times [at 3 (PrBe 3) and 10 (PrBe 10) d] or sequentially, respectively), 2PrBe+PeCBBz , 2PrBe+MeCBBz , along with weed-free and nontreated weedy check treatments. Herbicide treatments did not cause visual phytotoxicity to rice, and bending and leaf rolling were not observed. Only the two propyrisulfuron treatments had temporary negative effects on rice height, but rice recovered quickly. Compared with the weed-free treatment, CBPy did not affect rice tiller number or dry matter accumulation. Compared with the nontreated weedy check, herbicide treatments reduced total weed density by 29.4% to 99.1% and dry biomass by 32.2% to 98.7%. The CBPy treatment provided the best weed control, reducing weed density and biomass by 96.7% and 95.9% in 2021 and 97.4% and 95.6% in 2022, respectively. Rice grain yield was not significantly different between CBPy and the weed-free treatment in either year. Economic analysis showed that CBPy provided the highest net profit, followed by that in 2PrBe+PeCBBz and 2PrBe+MeCBBz, with the lowest net profit in the nontreated weedy check. Thus, CBPy provides good weed control and could be promoted in mechanically transplanted rice fields in China. The average air temperature, precipitation, and sunshine hours during the rice-growing season across the two years were recorded at a weather station close to the experimental site, as listed in To evaluate herbicides, an experiment was conducted on a farm at the Jiangsu Academy of Agricultural Sciences, Jiangsu Province, China , during the rice growing season (May–October) in 2021 and 2022. The soil was an Entisol, Typic fluvaquent  and a sajaponica cultivar), which is currently used in local production, was grown in the paddy field in 2021 and in 2022. In both years, seedlings were raised in a seedbed with sowing date on 15–16 May and mechanically transplanted on 3–5 June at a hill spacing of 0.25 m × 0.14 m with three seedlings per hill. Heading date (50% of plants) was on 25–28 August, and plants were harvested on 15–18 October. Total N application rate was 270 kg ha−1, and N was applied as urea 1 d before transplanting and at early tillering (7 d after transplanting [DAT]) and panicle initiation (40 DAT) stages in proportions of 50%, 10%, and 40%, respectively. Phosphorus (40 kg ha−1 as single superphosphate) and K (40 kg ha−1 as KCl) were applied as basal fertilizer the day before mechanical transplanting. The experiment followed a complete randomized block design, with four replicates and a plot size of 4 m × 6 m. Plots were separated by a round 0.5 m wide alley using plastic film inserted into soil to a depth of 50 cm.The high-yielding rice cultivar Nanjing 9108  propyrisulfuron , which is a novel sulfonylurea herbicide that is widely used in controlling annual or perennial weeds and grasses in paddy fields because of its high safety, high efficiency, and broad-spectrum yet highly selective effects; (3) cyhalofop butyl+propyrisulfuron (CBPy); (4) pretilachlor+bensulfuron , which is a mixture of sulfonylurea and amide herbicides that interferes with amino acid and protein synthesis in annual weeds; (5) pretilachlor+bensulfuron ; (6) pretilachlor+bensulfuron followed by (fb) pretilachlor+bensulfuron (PrBe 3+PrBe 10); (7) pretilachlor+bensulfuron fb pretilachlor+bensulfuron fb penoxsulam+cyhalofop butyl+bentazone (2PrBe+PeCBBz), with penoxsulam  an acetolactate synthase inhibitor that is a highly efficient, broad-spectrum herbicide used in rice fields worldwide and bentazone  a POST herbicide used to treat broad-leaf weeds in a variety of crops including rice; (8) pretilachlor+bensulfuron fb pretilachlor+bensulfuron fb metamifop+cyhalofop butyl+bentazone (2PrBe+MeCBBz), with metamifop  a POST aryloxyphenoxypropionic acid herbicide used to control a wide range of annual and perennial grass weeds in rice; (9) weed-free (W0), which was manually weeded; (10) nontreated weedy check (W+). The POST herbicides (2-pressurized knapsack sprayer equipped with four “teejet 8002” (TeeJet Technologies (Ningbo) Co., Ltd, Ningbo, Zhejiang, China) flat-fan nozzles to deliver water at 450 L ha−1. Spray pressure, spraying height, boom width, and nozzle spacing were 2.0 bar, 0.5 m, 2.0 m, and 0.5 m, respectively.The experiment comprised ten treatments listed in rbicides Table 1 2.32.3.12 area of rice seedlings every 10 d until 80 DAT. In addition, aboveground rice biomass was recorded in each plot in a 0.25 m2 area at 0, 20, and 40 DAT.Symptoms of rice phytotoxicity were assessed at 3, 5, 7, 10, and 15 DAT using a visual rating scale : 1 = hig2.3.2Weed density levels were recorded in four randomly located quadrats (0.5 m × 0.5 m) in each plot 15, 30, and 60 DAT. Aboveground parts of all dominant weeds were collected at 60 DAT and then oven-dried at 70°C for 72 h to determine dry weight. Weed control efficacy (WCE) was calculated by the following formulas:2.3.32 site (except border plants and weed sampling areas) in each plot then were adjusted to a moisture content of 0.14 g of H2O g−1 fresh weight before statistical analysis.Grain yields were recorded from a 5-mNet returns for each treatment were calculated over variable costs of production Table 2.2.4P = 0.05.Analysis of variance was performed using a SAS/STAT statistical analysis package . Data from each sampling date were analyzed separately. Means were tested by the least significant difference at 33.1Echinochloa spp. and Leptochloa chinensis (L.) Nees; two broadleaf species: Monochoria vaginalis (Burm.f.) C. Presl and Ammannia multiflora Roxb; and one sedge species: Cyperus difformis L.Weeds in the experimental field were a naturally occurring mixed population. In W+ (nontreated weedy check), there were 11 weed species dominated by grass, broadleaf, and sedge weeds. The dominant weeds were two grass species: 3.2−1 had a temporary negative effect on rice height, by 60 DAT, final rice height was not affected  and bending and leaf rolling were not observed (data not shown). The herbicide treatments CB, PrBe 3, PrBe 10, PrBe 3+PrBe 10, 2PrBe+PeCBBz, and 2PrBe+MeCBBz did not affect plant height Figure 2In the two years, tillering number was not significantly different among treatments before 20 DAT, but with continued rice growth, significant differences were observed among treatments Figure 3Compared with W0, rice aboveground biomass decreased in W+ but was not significantly different in the other treatments at 20 DAT Figure 43.3Echinochloa spp., L. chinensis, M. vaginalis, A. multiflora, and C. difformis were not significantly affected in CB, Py, and CBPy with post-emergence spraying , 2PrBe+MeCBBz (98.2%), and CBPy (96.7%), followed by that in Py (81.5%), PrBe 3+PrBe 10 (81.0%), and PrBe 10 (59.8%) Table 5.3.4−1) was in W0, and different herbicide treatments strongly influenced grain yield. Compared with W+, herbicide treatments increased yields 17.3% to 135.2% in 2021 and 21.5% to 153.1% in 2022 , 2PrBe+PeCBBz (9.42 t ha–1), and 2PrBe+MeCBBz (9.58 t ha–1) were significantly higher than those in other treatments , followed by that in 2PrBe+PeCBBz , PrBe 3+PrBe 10 , PrBe 3 and PrBe 10 (600 g a.i. ha–1), CBPy (385.5 g a.i. ha–1), and CB (300 g a.i. ha–1). The lowest dose was in Py (85.5 g a.i. ha–1). Cost of weeding was in an order of 2PrBe+PeCBBz > 2PrBe+MeCBBz > CBPy > Py > PrBe 3+PrBe 10 > PrBe 3 and PrBe 10 > CB. Based on rice price and other expenditures (−1), followed by that in 2PrBe+MeCBBz , 2PrBe+PeCBBz , PrBe 3+PrBe 10 , Py , PrBe 10 , CB , and PrBe 3 , with the lowest benefit in W+ (555 Yuan ha−1)  alone or in combination reduced rice plant height by 11.3% to 17.6% at 20 and 30 DAT. These results are consistent with previous studies in which Py reduced rice plant height by 11.4% to 60.1% with application rates of 12.5–200 g a.i. ha−1 , Schoenoplectus juncoides, C. serotinus (sedge), and M. vaginalis (broadleaf) at 70 and 140 g a.i. ha−1 with good selectivity to rice. It also has a wide insecticidal spectrum, a long efficacy period, and high biosafety for human, birds, and aquatic and other organisms , suggesting some weeds still occurred at the late stage of rice growth. Notably, those weeds that failed to seed would help deplete the weed seed bank.Propyrisulfuron is widely used to control annual or perennial weeds in paddy field, dry land, and other cultivated lands due to its high safety, high efficiency, broad-spectrum and high selectivity . Ikeda erganisms . In the In rice–wheat and rice–oilseed rape rotations in China, farmers often face the challenge of rapidly establishing rice crops due to limited labor availability . TherefoThe application of CBPy reduces the number of herbicide applications by 1-2 times, addressing labor shortages and reducing herbicide application dose. With such benefits, it is curious why the herbicide has not been widely promoted in China. Determined in a previous investigation, a higher price of Py than that of PrBe might be one reason. In addition, farmers have formed a habit of using the herbicide formula of PrBe as a pre-emergent herbicide combination, which also provides good weed control. Ultimately, farmers primarily grow rice for the net profit. In the present study, compared with W0, similar yields were obtained in CBPy, 2PrBe+PeCBBz, and 2PrBe+MeCBBzs. However, the highest net profit was in CBPy, which was significantly higher than that in 2PrBe+PeCBBz and 2PrBe+MeCBBz. Hence, given CBPy’s effective weed control and its ability to yield the highest net profit, it should be actively promoted as a preferred weed control strategy.The results have demonstrated that CBPy offers effective weed control, labor and cost savings, and maintains high yield in mechanically transplanted rice fields. However, one potential issue with CBPy is its temporary inhibitory effect on rice height, although rice plants recover quickly from this. This temporary setback in rice height may not be acceptable to some farmers. Therefore, finding a solution to address this concern is crucial. Switching from spraying to broadcasting the herbicide may be a viable approach to prevent rice damage while still benefiting from effective weed control.In conclusion, the field experiments showed that in mechanically transplanted rice, the herbicide treatments effectively reduced average weed density by 74.2% and average weed biomass by 74.6%. Treatments CBPy, 2PrBe+PeCBBz, and 2PrBe+MeCBBz had the highest total weed density control efficacy and total weed dry weight control efficacy. In addition, compared with W+, CBPy, 2PrBe+PeCBBz, and 2PrBe+MeCBBz also displayed relatively higher yields. Furthermore, no significant difference was observed when these treatments were compared with W0. The highest net economic gain was in CBPy, because it reduced herbicide dose and number of applications and thus weeding costs and also provided good weed control.The original contributions presented in the study are included in the article/ZZ: Investigation, Writing – original draft, Writing – review & editing. HW: Investigation, Methodology, Conceptualization, Data curation. TG: Investigation, Methodology, Data curation, Writing – original draft. JC: Conceptualization, Data curation, Methodology, Writing – original draft. YL: Formal Analysis, Conceptualization, Investigation, Writing – original draft. GL: Formal Analysis, Software, Writing – original draft.


Fibrinogen and homocysteine (HCY) are molecules known to play a role in vascular homeostasis, and their blood levels are often elevated in patients with metabolic syndrome. Recent evidence suggests that sudden sensorineural hearing loss (SSHL) may have a vascular origin. This has led many authors to advocate that fibrinogen, homocysteine, and metabolic syndrome (MetS) may play a direct role in SSHL. The aim of this brief review is to examine the role and influence of these molecules and MetS on the mechanisms of SSHL. Elevated fibrinogen levels have been associated with a worse prognosis in SSHL, possibly due to increased blood viscosity and decreased blood flow. Similarly, HCY has been associated with vascular damage, particularly in hyperhomocysteinemia, although the exact association with SSHL remains controversial. MetS has been demonstrated to function both as a causative factor and as a contributor to poorer recovery in cases of SSHL. However, although some studies suggest a possible role for these biomarkers and MetS in the prognosis and treatment of SSHL, specific therapeutic and preventive strategies based solely on these factors have yet to be developed. Given their potential role in prognosis and treatment and the global epidemic of metabolic syndrome, this issue needs to be analyzed comprehensively. Thus, further quality studies need to be conducted, even though it is difficult to determine the actual impact of MetS on the development of SSHL, as it is a multifactorial disease affecting multiple organs. Sudden sensorineural hearing loss (SSHL) is defined as hearing loss occurring within 72 h with a loss of at least 30 decibels in three adjacent frequencies (The American Academy of Otolaryngology—Head and Neck Surgery) . The incNumerous treatment options have been explored for idiopathic SSHL, including anti-inflammatory agents, antimicrobial drugs, calcium antagonists, vitamins, essential minerals, vasodilators, volume expanders, defibrinogenating agents, diuretics, hyperbaric oxygen therapy, and bed rest , but theThe current Clinical Practice Guideline for Sudden Hearing Loss recommends that systemic steroids and/or hyperbaric oxygen therapy (HBOT) may be considered as treatment options if administered within two weeks of symptom onset. However, it is important to note that the efficacy of these treatments may vary greatly from person to person, possibly due to the different causes of SSHL. Therefore, the routine use of antiviral drugs, antithrombotics, vasodilators, vasoactive agents, and similar therapies is not recommended for the standard treatment of SSHL . It is iAccording to recent evidence, fibrinogen and homocysteine play a role in sudden sensorineural hearing loss (SSHL). Also, MetS, with its intrinsic biochemical alterations, has been shown to have the potential to cause vascular damage, even within the cochlea. Consequently, the inflammatory mechanisms within the vascular network may lead to cochlear cell damage, contributing to the development of SSHL. Understanding these factors may improve our understanding of the vascular and circulatory aspects of SSHL, which may allow the development of more targeted diagnostic tools and therapeutic interventions. Such insights may ultimately lead to better outcomes for patients with SSHL, giving them a better chance of preserving their hearing function and overall quality of life.In this brief narrative review, we aim to examine the role and influence of these factors on the mechanisms of SSHL. Investigating the role of fibrinogen, homocysteine, and MetS in sudden sensorineural hearing loss (SSHL) is of paramount importance because of the potential impact on both the diagnosis and treatment of this debilitating condition.“fibrinogen” AND “Sudden sensorineural hearing loss (SSHL)”;“homocysteine” AND “Sudden sensorineural hearing loss (SSHL)”;“Metabolic Syndrome” AND “Sudden sensorineural hearing loss (SSHL)”.To collect relevant scientific literature on our research topic, we conducted a comprehensive search through the PubMed database. The search included articles published within the last 25 years. The following strings and keywords were used in our search strategy:The search was carried out to identify studies and articles that explored the relationships and interactions between these keywords, as they pertain to our research objectives.-Published within the last 25 years (from 1998 to 2023);-Written in English;-Focused on the association between fibrinogen, homocysteine, metabolic syndrome, and sudden sensorineural hearing loss;-Available as full-text articles.The included articles had to meet the following criteria:Excluded articles were those that did not meet these criteria or were duplicates of other retrieved publications.Sudden sensorineural hearing loss (SSHL) is a medical emergency that requires immediate ENT medical attention. Much of the literature indicates that 32% to 65% of cases of SSHL can recover spontaneously. However, clinical experience indicates that these numbers may be overestimated . VariousAccording to a review of the evidence regarding the etiology of sudden sensorineural hearing loss (SSHL) in adults conducted by Chau et al. , they diSeveral lines of research have focused on the concept that altered blood flow and/or oxygenation lead to cochlear hair cell damage and hearing loss . AccordiGiven the theory of microvascular occlusion, it is worthwhile to investigate hypercoagulability in patients with idiopathic SSHL, especially when thrombophilia-related abnormalities or predominant venous factors such as oral contraceptive use, pregnancy or puerperium, and cardiovascular factors such as arterial hypertension, hyperlipidemia, diabetes, and cigarette smoking are present. Thrombophilia abnormalities can be divided into two categories: inherited and acquired. Inherited thrombophilia includes conditions such as deficiency of natural anticoagulant proteins such as antithrombin, protein C, and protein S, and gain-of-function mutations in the genes of factor V and prothrombin. On the other hand, acquired thrombophilia is due to factors such as the presence of antiphospholipid antibodies. In addition, there are two other thrombophilia-related disorders for which there is no clear evidence of inheritance. These are elevated plasma levels of coagulation factor VIII and mild to moderate hyperhomocysteinemia (HHcy). Finally, circulating microparticles, tiny phospholipid-rich particles released by various blood cells such as platelets, endothelial cells, leukocytes, and erythrocytes, are also considered potential risk factors for sudden sensorineural hearing loss (SSHL) because they are associated with an increased risk of thrombosis  Figure .Fibrinogen is a glycoprotein normally present in human blood plasma. It is mainly produced by the liver and released into the bloodstream. It plays a central role in several biological functions, including hemostasis and inflammation . AlthougIn a study in guinea pigs, an association was found between elevated serum fibrinogen levels and a decrease in blood flow in the cochlea. It was also observed that administration of a defibrinogenating drug improved blood flow in the cochlea . In a prIndeed, fibrinogen levels may indicate increased blood viscosity, which in turn could lead to decreased blood flow in the affected regions. In 2018, Oya et al. performed a literature review on the relationship between SSHL and fibrinogen and analyzed 1577 patients. They found that a high serum fibrinogen level correlated with a poor prognosis . These rAs noted by Hira et al., plasma fibrinogen levels increase susceptibility to atherosclerosis due to two main factors: increased plasma viscosity and thrombotic activation triggered by fibrin during its degradation . NumerouHowever, it is important to note that the pathophysiology of SSHL at both low and high frequencies is not yet confirmed to be due to only thrombosis. Consequently, the role of fibrinogen remains unclear, and its impact on prognosis is currently not fully understood. To date, it is not possible to definitively determine whether hyperfibrinogenemia directly causes SSHL. In fact, it remains unclear whether hyperfibrinogenemia is the cause or the consequence of SSHL .Homocysteine (HCY) is an amino acid produced in the body as a byproduct of the metabolism of another amino acid called methionine. It is also a homolog of cysteine . SeveralA laboratory study showed that when homocysteine levels are elevated, a condition known as hyperhomocysteinemia (hHCY) can be associated with an increased risk of neurovascular disease, dementia, migraine, developmental disabilities, and epilepsy . MechaniAs mentioned above, HCY is a risk factor for increased vascular injury, and its abnormal increase impairs endothelial function, reduces vascular flexibility, and leads to microvascular dysfunction, resulting in ischemia and hypoxia-related changes . CurrentA retrospective case–control study by Passamonti et al. showed that HCY was associated with an increased risk of idiopathic SSHL, and hyperhomocysteinemia was reported as a negative prognostic factor for hearing recovery . In addiNiu et al. conducted a literature review on the association between SSHL and HCY in 2023 . The stup < 0.05). Huang et al. attempted to administer folic acid, vitamin B6, and B12 to patients with SSHL, but their results showed no improvement in clinical outcomes. Nevertheless, they suggest that maintaining low levels of HCY in daily life may prove beneficial for the clinical prevention and treatment of hearing loss [There are also some authors who do not believe in the role of HCY in SSHL. Cadoni et al. found no correlation between these two factors . Also, Bing loss .To date, there is insufficient evidence to definitively clarify the exact relationship between fibrinogen and HCY levels and their effects on hearing loss. Although many studies postulate this hypothesized relationship, this is not sufficient to develop therapeutic and preventive strategies based solely on these assessments.While metabolic syndrome (MetS) itself does not involve single molecules like fibrinogen and homocysteine (HCY), it plays a significant role in causing various molecular dysfunctions that can potentially lead to vascular damage, which, in turn, affects the cochlea. As a condition with a high global prevalence, it is valuable to comprehend its implications, including a comprehensive analysis of its individual components. In this study, our objective is to assess the influence of metabolic syndrome (MetS) and the accompanying molecular alterations on the pathophysiology of sudden sensorineural hearing loss (SSHL).Metabolic syndrome (MetS) is a set of metabolic disorders that include central obesity, insulin resistance or impaired glucose uptake and utilization, dyslipidemia, and hypertension . MetabolIts multisystemic pattern arises from a confluence of factors including pro-inflammatory states, oxidative stress, hemodynamic dysfunction, and ischemia in ‘dysmetabolic’ patients. This convergence leads to the emergence of various clinical conditions, encompassing cardiovascular diseases (CVD), non-alcoholic steatohepatitis, liver dysfunctions, chronic kidney disease, numerous types of cancer, and neurodegenerative disorders .Metabolic syndrome and its associated components have the potential to lead to several comorbidities, one of which is inner ear dysfunction. The exact mechanism linking metabolic syndrome and hearing loss is not yet clear, but there is evidence that it may be related to peripheral blood vessel dysfunction . It is kMetabolic syndrome encompasses a number of disorders that include insulin resistance and abnormal fat deposition. The combination of these factors can lead to microvascular damage and endothelial dysfunction . NumerouMetS has also been shown to be associated with poorer recovery in SSHL. Indeed, metabolic syndrome had a negative impact on hearing improvement in patients with idiopathic SSNHL. Lower initial hearing threshold, absence of diabetes mellitus and hypertension, and BMI < 25 were associated with improvement in hearing after treatment, according to the results of Zand et al. . These fNumerous studies have addressed the involvement of blood parameters, particularly fibrinogen and homocysteine, in the context of SSHL. Elevated fibrinogen levels have been associated with a poorer prognosis, possibly due to increased blood viscosity and decreased blood flow. Similarly, HCY has been associated with vascular damage, particularly in cases of hyperhomocysteinemia, although the exact association with SSHL remains controversial.While recent findings have shed light on the possible involvement of fibrinogen and HCY in the pathophysiology of SSHL, concrete data confirming their direct link to cell damage are still lacking. Moreover, their role in the prevention or treatment of SSHL remains unclear. To date, they remain hypothetical factors within a complex mechanism involving several elements. Several studies have investigated metabolic syndrome (MetS) and all its molecular alterations as a possible cause of sudden sensorineural hearing loss because of its impact on the vascular network and endothelial damage, particularly in the cochlear region. Indeed, MetS has been shown to affect the recovery process of SSHL, with patients with elevated levels of MetS parameters showing poorer recovery outcomes. Further research is needed to gain a comprehensive understanding of this complicated issue.The clinical implications of HCY, fibrinogen, and molecular alterations of MetS in SSHL remain a focus of research. Given their potential role in prognosis and treatment and the global epidemic of metabolic syndrome, this issue needs to be analyzed comprehensively. The actual pathogenetic role in sudden sensorineural hearing loss is not yet fully understood, so specific therapeutic and preventive strategies based solely on these biomarkers have yet to be developed. In the future, comprehensive and multidimensional research approaches should be prioritized. First, researchers should consider conducting large-scale, longitudinal cohort studies to establish causal relationships and describe the precise mechanisms by which fibrinogen, homocysteine, and molecular alterations of metabolic syndrome affect auditory function. In addition, investigating potential synergistic effects between these factors and other risk factors, such as age, genetics, and lifestyle, would provide a more holistic understanding of their impact on hearing loss. In addition, the incorporation of advanced imaging techniques such as functional MRI or Doppler ultrasound could help visualize the vascular changes associated with fibrinogen and homocysteine, providing valuable insights. Finally, interventional studies examining the efficacy of therapies to prevent or treat hearing loss by affecting fibrinogen and homocysteine levels could provide valuable clinical recommendations for future treatment strategies. With these multifaceted approaches, future studies can contribute significantly to our understanding of the role of fibrinogen, homocysteine, and metabolic syndrome in hearing loss and potentially lead to more effective prevention and treatment strategies. In addition, as recent studies have shown, there is increasing evidence of biomarkers for the prognosis of SSHL. Heat shock protein-70, anti-endothelial cell antibodies, and prestin have shown potential in this regard. Therefore, further research on various blood molecules is needed to elucidate the specific mechanisms and prognosis of SSHL. This endeavor promises to pave the way for new targeted therapies and improve the efficacy of drugs against SSHL.


The optical properties of graphene nanodots (GND) and their interaction with phosphate ions have been investigated to explore their potential for optical sensing applications. The absorption spectra of pristine GND and modified GND systems were analyzed using time-dependent density functional theory (TD-DFT) calculation investigations. The results revealed that the size of adsorbed phosphate ions on GND surfaces correlated with the energy gap of the GND systems, leading to significant modifications in their absorption spectra. The introduction of vacancies and metal dopants in GND systems resulted in variations in the absorption bands and shifts in their wavelengths. Moreover, the absorption spectra of GND systems were further altered upon the adsorption of phosphate ions. These findings provide valuable insights into the optical behavior of GND and highlight their potential for the development of sensitive and selective optical sensors for phosphate detection. Phosphate is a ubiquitous chemical compound ,2, foundNumerous methods have been developed for detecting phosphates, including high-performance liquid chromatography (HPLC) , fluores−1) based on Au@Ag nanoparticles and GND has been developed for the determination of the activity of alkaline phosphatase (ALP) [3+) [43− due to the strong interaction between the phosphate ion and Al3+. In another investigation [3+). The detection principle was based on the observation that the oxygen-donor atoms in phosphate ions exhibit a greater affinity towards Eu3+ ions than the carboxylate groups on the surface of GND. Consequently, the Eu3+ ions tend to preferentially interact with the phosphate ions instead of the GND, leading to a restoration of the optical signal of the GND.Graphene nanodots—small fragments or islands of graphene that exhibit quantum confinement effects due to their reduced size and confinement boundaries —are promse (ALP) . Since Ase (ALP) . Chen etLP) [3+) . This ontigation , an off–The motivation behind this research arises from the necessity to enhance current phosphate detection methods and investigate innovative approaches in light of the environmental and health implications associated with phosphate concentrations. The objective of this study is to establish a theoretical basis for the design and optimization of phosphate ion sensors with specific performance characteristics. Although research on using GND for phosphate detection is underway ,40,41, i96H24, commonly known as circumcircumcoronene [96H24 serves as a reasonable and appropriate choice for this study. Vacancy-containing and metal-doped GND structures  were created using the GaussView6 tool [2PO4−, HPO42− and PO43− negatively charged ions (anions) were selected to simulate the phosphate ions (+). This favors the complete protonation of the phosphate ion (H3PO4). As the pH becomes more alkaline, the gradual dissociation of protons from the phosphate molecule occurs, leading to partial or complete deprotonation and the formation of different species such as H2PO4−, HPO42− and PO43−, depending on the number of protons released. However, it should be noted that in the current research, the specific investigation or consideration of the degree of deprotonation of phosphates under varying solution pH conditions was not conducted. The primary focus of the study was on performing DFT and TD-DFT calculations in the presence of water. In aqueous environments, the probability of phosphate ions being deprotonated and existing as dihydrogen phosphate (H2PO4−) or hydrogen phosphate (HPO42−) species is increased. However, it is important to acknowledge that the presence of calcium cations (Ca2+) can significantly reduce the barrier for hydrogen phosphate deprotonation [43−) in real water samples as well. Considering these factors, the investigated reactions in this study demonstrate a high degree of likelihood, and the findings obtained hold valuable potential for the development of technologies aimed at monitoring phosphates in water. However, it is recommended that future investigations explore this aspect further to gain a deeper understanding of phosphate behavior under varying pH conditions and its impact on adsorption phenomena.DFT calculations were performed using the Gaussian 16 Rev. C.01 program package . The neucoronene , was usecoronene ,48,49. Tew6 tool  through ate ions . It is itonation , suggestEPI-GND, PEI, and EGND are the total energies of the phosphate-GND complex, isolated adsorbed phosphate ion, and isolated GND, respectively. The optimization was performed using the long-range-corrected CAM-B3LYP functional [The adsorption energy of the phosphate ions on GND in the presence of water was calculated using the formula:nctional   until tThe CAM-B3LYP functional was chosen for its good accuracy in predicting the electronic properties of graphene-based molecules ,57,58,59−1 cm−1) [TD-DFT calculations  were car−1 cm−1) . However−1 cm−1) .The optimized geometries of GND and its modified counterparts, including vac-GND, Ca-GND, Mg-GND, and Al-GND, were obtained by performing PCM/CAM-B3LYP/6-31G(d) calculations. The top and side views of the optimized structures are depicted in z1 and Δz2 parameters are then calculated to quantify the degree of buckling and curvature   was alsoα) and spin-down (β) electrons. This phenomenon arises due to the presence of spin-dependent interactions and subsequent spin polarization within the Al-doped GND system. Notably, the partially filled d-shell of aluminum facilitates an exchange coupling between the localized aluminum spins and the itinerant carbon electron spins. Furthermore, upon immersion of the Al-doped GND molecule in water, the HOMO-LUMO gaps were observed to be 3.860 eV for the α mode and 2.954 eV for the β mode  and the lowest unoccupied molecular orbital (LUMO) . The HOMe β mode .In the context of adsorption, the behavior of phosphate ions on the surfaces of the investigated systems is worth considering. Analysis of 2PO4− molecule are directed towards the GND surface, while the phosphorus atom is positioned above a carbon atom. Interestingly, despite the initial placement of the H2PO4− molecule directly above a missing carbon atom in vac-GND, it undergoes a shift towards the edge of the GND during the optimization process , instead of one as observed in the H2PO4−@Al-GND and HPO42−@Al-GND systems. An interesting observation is the significant reduction in the bond lengths of Ca-O and Mg-O with an increase in the negative charge of the ion: from 2.50 and 2.10 Å for the H2PO4−@Ca-GND and H2PO4−@Mg-GND systems to 2.29 and 1.95 Å for the PO43−@Ca-GND and PO43−@Mg-GND systems. The intermediate parameters for H2PO4−@Ca-GND and H2PO4−@Mg-GND are 2.30–2.43 and 2.01 Å, respectively.Examining metal-doped graphene nanodots, it is evident that the metal atom forms bonds with one or two oxygen atoms c–e. In t surface f–o closeh H2PO4− g–i. Addi2PO4− adsorption on vac-GND. Physisorption arises from weak intermolecular forces, resulting in low adsorption energy and minimal electronic or chemical interactions. Conversely, chemical adsorption involves stronger chemical bonds (1–3 Å), leading to higher adsorption energy, substantial electronic and chemical interactions, and increased stability and specificity. Furthermore, when examining other GND surfaces, although the adsorption energies of phosphate ions remain below the 0.5 eV threshold, the equilibrium distance between the ions and the surface falls within the 1–3 Å range, indicating a significant interfacial electron transfer, as illustrated in 43− ions exhibiting the highest adsorption energies, followed by HPO42− ions, and H2PO4− ions having the lowest adsorption energies. Finally, as we move from pristine GND to modified GND, the remaining charges on phosphate ions decrease, becoming less negative. Indeed, the observed trends indicate that phosphate ions undergo electron loss during the adsorption process. This also suggests that the modified GND surfaces, such as vac-GND, Ca-GND, Mg-GND, and Al-GND, which exhibit significant electron transfer with relatively low adsorption energies, may possess favorable characteristics for applications requiring both efficient electron transfer and easy desorption or recovery of the phosphate ions. The modification of the charge distribution caused by the adsorbed ions resulted in the changes in HOMO-LUMO energy gaps , the energy gaps decrease progressively from H2PO4− to HPO42− to PO43− adsorption. This implies an increasing influence of the adsorbed ions on the electronic properties of these systems, leading to a more pronounced impact on the energy gap. It is worth highlighting the distinct behavior of Al-GND, where the energy gaps differ significantly between the alpha and beta modes after the adsorption of phosphate ions. This suggests the presence of spin-dependent effects and spin polarization induced by the combination of aluminum dopants and phosphate ions.Moving beyond the structural characterization of phosphate ions on both pristine and modified GND surfaces, a comprehensive analysis of the adsorption energies and the magnitude of charge transfer between these ions and the surface becomes imperative. These factors assume a pivotal role in defining the sensing performance, making their evaluation vital. More specifically, the adsorption energy determines the strength of binding between the phosphate ion and the GND surface, consequently influencing the recovery time. A higher adsorption energy leads to a prolonged recovery process. Similarly, the degree of charge transfer between the adsorbed phosphate ions and the GND surface governs the sensor’s response. Hence, to achieve an optimal balance between rapid response and swift recovery, it is advantageous to have a substantial electron transfer coupled with a minimal adsorption energy . DFT resrgy gaps . Upon adThe energy gap of GND systems demonstrates a profound correlation with the size of the adsorbed phosphate ions on their surface, implying that the interaction between GND and phosphate ions can lead to significant modifications in their absorption spectra.This holds great significance, particularly in the realm of optical sensor development for monitoring phosphate presence in water. Consequently, the following sections will discuss the results of TD-DFT investigations, shedding light on the diverse aspects of the impact of GND-phosphate ion interactions on the absorption spectra of GND. Through the elucidation of these findings, valuable contributions can be made towards advancing knowledge in the field of optical sensing of phosphates. In this regard, it is crucial to conduct an in-depth analysis of the intrinsic optical behavior exhibited by both pristine and modified GND prior to phosphate ion adsorption. This analysis will serve as a fundamental reference point for interpreting any subsequent alterations observed in the optical response after the adsorption of phosphate ions.69,70 ← S0, and S3,4 ← S0 with high oscillator strength), while the third one is related to S15,16 ← S0 transitions with a much lower oscillator strength of 0.48  in the system when vacancy is introduced. Conversely, the UV band shows a noticeable redshift, characterized by a shift towards longer wavelengths.The absorption spectrum of pristine GND is dominated by two spectral bands with peaks at 252 nm and 462 nm, but a weak absorption band at 343 nm is also distinguishable . Since ttine GND . The abs3, S4, and S9 excited states, respectively. Moving to the Ca-GND system, it exhibits relatively similar spectral characteristics to pristine GND and vac-GND, with significant contributions from the S5, S6, and S7 excited states.A quick look at the evolution of the absorption spectra of metal-doped GND systems shows that the dopant-induced symmetry breaking leads to broadening of the main absorption bands of pristine GND and causes their shift, as can be clearly seen in the case of low-symmetry systems (Mg-GND and Ca-GND). Simultaneously, the Al-GND system demonstrates a prominent absorption band within the visible range, accompanied by a subtle shoulder in the shorter wavelength region of the spectrum. A comprehensive analysis of each spectrum reveals intriguing insights into the excited states of the Mg-GND, Ca-GND, and Al-GND systems. In the Mg-GND system, the most prominent and intense band spanning from 350 nm to 760 nm can be attributed to the S98 state. On the other hand, the absorption spectrum of Al-GND is dominated by a broad band centered at 434 nm, with the primary contributions originating from the S23 ← S0 and S24 ← S0 transitions. Additionally, a weak shoulder around 334 nm can be attributed to the S66 ← S0 transition. Notably, in the singlet excited state (S23), the promotion of electrons occurs from occupied molecular orbitals with a spin-down configuration to unoccupied molecular orbitals , resulting in a singlet state characterized by parallel spins. In contrast, the S24 state involves the promotion of electrons from both α and β occupied molecular orbitals to an unoccupied molecular orbital with a similar spin configuration. Furthermore, the transitions between molecular orbitals with spin-up configurations contribute significantly to the S66 state. A summary of the most probable electronic transitions involved in these excited states can be found in The UV absorption baId in the Ca-GND system is primarily influenced by the S43− adsorption, the band completely disappears.It becomes pertinent to inquire about the alterations occurring in the absorption spectra of GND systems when phosphate ions are introduced. The absorption spectrum of the GND exhibits notable modifications, mainly in the middle-ultraviolet region, following the phosphate ion adsorption, as illustrated in In the context of this study, the successive interaction between vac-GND and each of the three ions under consideration leads to the emergence of three distinct absorption spectra a. These 2PO4− and HPO42− ions, the adsorption of PO43− gives rise to a distinct set of excited states and a noticeable reduction in oscillator strengths for the primary electronic transitions. These characteristics, however, do not preclude the utilization of Al-GND for monitoring the presence of PO43−.Turning our attention to the Al-GND system d, it is 43− adsorption serves as a clear signal of phosphate presence. These spectral modifications can be employed as optical signatures for the detection and quantification of phosphates, enabling the development of sensitive and selective optical sensing platforms. Considering the information given, GND, vac-GND, Ca-GND, Al-GND, and Mg-GND present themselves as viable options for constructing a sensing configuration that incorporates these elements into the sensor design. By harnessing the sensitivity of sensing elements’ UV–vis absorption to phosphate ions, along with employing suitable excitation sources, photodetection systems, and data analysis techniques such as multivariate analysis or pattern recognition algorithms, it becomes feasible to differentiate the three phosphate ions based on their unique spectral characteristics. In this manner, their distinctive spectral fingerprints can be utilized to discriminate between the phosphate ions effectively. Although the literature on this particular topic is still relatively scarce, it is evident that the phenomenon of pronounced property alterations in GND caused by the presence of PO43− ions extends beyond GND alone. Comparable behavior has been observed in other graphene-based materials, including negatively charged molybdate-mediated nitrogen-doped graphene quantum dots (Mo7O246− mediated N-GQDs) [43−, showcasing their superior adsorption capacity for PO43− in comparison to hydrogen phosphate and dihydrogen phosphate ions.The distinctive changes observed in the absorption spectra of GND, Mg-GND, and Ca-GND upon phosphate ion adsorption, particularly in the middle-ultraviolet region, offer potential for optical sensing of phosphates. The stable spectral characteristics in the near-ultraviolet and visible regions, with minimal intensity variations, provide a reference baseline for comparison. By focusing on the absorption band within the middle-ultraviolet range, it is possible to utilize the gradual reduction in band amplitude and the redshift of the peak as indicators of phosphate ion concentration. The disappearance/quenching of the absorption band entirely after PO N-GQDs) . Remarka43− ions detection. Although the existing literature primarily revolves around fluorescence-based techniques for detecting phosphate ions (in contrast to the absorption spectra discussed in this study), it is reasonable to deduce that performing a comparative analysis of parameters that characterize the alterations in optical signal amplitude of the quantum dots in the presence of phosphate ions would prove valuable. This analysis would provide insights into the effectiveness and sensitivity of sensors utilizing the investigated quantum dots, irrespective of the specific optical detection method employed. 43−. This finding substantiates the effectiveness and performance of the proposed approach. Moreover, this method boasts a multitude of advantageous characteristics, such as the capability to detect individual phosphate ions, potential for high sensitivity, and the versatility and tunability offered by the graphene quantum dot system as well as its cost-effective fabrication process.The research focus of this manuscript lies in exploring the interaction between graphene nanodots and phosphate ions for the purpose of optical sensing, demonstrating its significant impact on GND absorption spectra and identifying potential GND configurations for sensitive and selective phosphate detection. The introduction of vacancies and metal dopants in GND systems led to modifications in the absorption bands, broadening, and shifts in the wavelengths of the major electronic transitions. The adsorption of phosphate ions further modified the absorption spectra, particularly in the middle-ultraviolet region. The disappearance of the absorption band after phosphate ion adsorption served as a clear indication of phosphate presence. Based on the distinctive changes observed in the absorption spectra, GND, vac-GND, Ca-GND, Al-GND, and Mg-GND were identified as viable options for constructing optical sensing configurations. By utilizing the sensitivity of GND’s UV–vis absorption to phosphate ions, along with suitable excitation sources, photodetection systems, and data analysis techniques, it is feasible to differentiate and quantify phosphate ions based on their unique spectral characteristics. These findings contribute to the advancement of knowledge in the field of optical sensing and pave the way for the development of sensitive and selective optical sensing platforms for phosphate detection.


Sensorineural hearing loss has been a global burden of diseases for decades. However, according to recent progress in experimental studies on hair cell regeneration and protection, clinical trials of pharmacotherapy for sensorineural hearing loss have rapidly progressed. In this review, we focus on recent clinical trials for hair cell protection and regeneration and outline mechanisms based on associated experimental studies. Outcomes of recent clinical trials provided valuable data regarding the safety and tolerability of intra-cochlear and intra-tympanic applications as drug delivery methods. Recent findings in molecular mechanisms of hair cell regeneration suggested the realization of regenerative medicine for sensorineural hearing loss in the near future. Hearing loss has been a global disease burden for decades . ApproxiCurrently, no pharmacotherapeutic options are available for stable SNHL. Stable SNHL is the condition of SNHL showing no changes in hearing levels at any frequency by standard audiometric measures for six months, which includes acute SNHL previously treated with no response to corticosteroids . The maiSound stimuli are converted into neural signals in the cochlea of the inner ear. The vibration of otic ossicles, which transmit sound vibration from the tympanic membrane to the cochlea, generates sound waves in the cochlea. Sound waves cause a vibration of the cochlear sensory epithelium, in which sensory hair cells (HCs) form four rows along the cochlear coil. Cochlear sensory hair cells are divided into two types: inner hair cells (IHCs) and outer hair cells (OHCs). The IHCs are distributed in a single row and play a central role in the transmission of sound stimuli to the auditory primary neurons, namely the spiral ganglion neurons (SGNs), which are located in the modiolus of the cochlea. The OHCs are distributed in three rows on the outside of the IHC row and amplify cochlear sensory epithelium vibration ,15,16. Thttps://hereditaryhearingloss.org/ (14 February 2023)]. Several deafness genes are associated with hair bundle development and functioning and synaptic transmission . Cisplatin is widely used for the treatment of malignant neoplasms as well as pediatric cancers. Children are at greater risk of developing hearing loss than adults are, with dire consequences for speech development and social integration. Among patients with SNHL, the proportion of cisplatin-induced hearing loss is comparatively small, but over 50% of patients who undergo cisplatin therapy acquire SNHL . Mo. Mo24]. https://www.novctrd.com/ctrdweb/patientsummary/patientsummaries?patientSummaryId=680 (14 February 2023)]. Intracochlear application was used as the drug application route. No serious adverse events were found in 22 participants, but slight hearing loss was identified in 32% (7/22). In hearing assessments using pure tone audiometry, no meaningful increase in hearing was identified. Based on these results, the trial was suspended but not canceled. The most important finding of this trial is that the intracochlear delivery of adenovirus vectors is tolerable, which may aid the progression of future clinical trials using either intracochlear application or adenovirus vectors.The human adenoviral vector serotype 5 was used for ATOH1 gene transfer to SCs in a clinical trial (NCT02132130). A results summary is available on the website [The scientific background of this clinical trial probably included two experimental studies: Izumikawa et al. (2005)  and Krafhttps://www.clinicaltrialsregister.eu/ctr-search/trial/2016-004544-10/results (14 February 2023)], which show that trans-tympanic injection  at the highest dose of 250 micrograms of LY-3056480 was safe and well tolerated. A phase 2 double-blind, randomized, placebo-controlled trial for this compound was registered as NCT05061758; however, the present status for recruitment is “withdrawn.” The outcomes of hearing recovery in these clinical trials have not yet been reported.Two-phase 1/2 clinical trials of γ-secretase inhibitors aimed at inhibiting Notch signaling were completed  aiming for HC regeneration are underway, in which the main machinery is not the direct conversion of SCs to HCs, but the gain of stem cell potential in SCs. A set of seven small-molecule compounds, namely valproic acid , FSK , CHIR , 616452 (a TGF-β receptor inhibitor), Tranyl (a histone demethylation inhibitor), DZNep , and TTNPB , could replace all four Yamanaka transcription factors to successfully reprogram mouse somatic cells into pluripotent stem cells . Among tMcLean et al. (2017) demonstrated the efficacy of the combination of VPA and CHIR (FX-322) in enhancing the SC potential for sphere formation and HC generation using cell cultures and for SC conversion to HCs in explants of neonatal mouse cochleae . HoweverClinical trials for the topical application of VPA and CHIR99021 (FX-322) are underway to investigate its potential for hearing restoration in patients with age-related hearing loss, noise-induced hearing loss, and sudden SNHL . The outRecent studies have revealed that forced ATOH1 expression alone is not sufficient to induce HC regeneration in mature mammalian cochleae ,62. ThesThe age-dependent decline in the conversion ability of ATOH1 has led to the search for additional transcription factors to convert older cochlear SCs into HCs . In geneIsl1, Ikzf2, POU4F3, LIN28B, Sall1/3, and Aff3 as lower-expressed genes in converted HCs than in mature OHCs [Isl1 but did not demonstrate the induction of converted HCs expressing mature OHC markers, including prestin [Yamashita et al. (2018) performed RNA sequencing of newly converted HCs and mature OHCs after a conditional overexpression of ATOH1 in specific types of SCs, pillar and Dieters cells , which cure OHCs . These t prestin . Sun et  prestin .ATOH1’s function is highly context dependent. During inner ear development, ATOH1 can only promote HC differentiation at specific developmental stages ,70. GFI1Successful direct conversion using co-transfection of pioneer factors in other systems has been reported ,81. TogeMany mammalian organs lack stem cell pools similar to mature cochleae but sometimes exhibit regenerative capacity. Differentiated cells in organs accomplish the regeneration of not only their own cells but also different cell types via direct conversion ,82. ReceDoetzlhofer and colleagues published a series of publications stating the roles of LIN28B and follistatin (FST) in the ability of mouse SCs to HC regeneration ,84,85. AEDNRB2, followed by the acquisition of HC identity [In the avian cochlea, HC regeneration spontaneously occurs after HC damage through SC-to-HC conversion and mitotic proliferation of SCs , similaridentity . In addiidentity . ElucidaIn the last decade, research and development of pharmacotherapy for SNHL have rapidly progressed. The major mechanism for HC protection against cisplatin is the reduction of cellular stress, which is a common cause of cochlear degeneration due to various causes. Therefore, we can expect to explore therapeutics that are commonly used for cochlear protection based on clinical trials of cisplatin-induced hearing loss. The progression of cochlear cell damage, particularly of the SCs in the sensory epithelium, can diminish the opportunities for HC regeneration. In fact, cisplatin also damages SCs . Hence, Recently, clinical trials for stable SNHL have rapidly emerged ,24. The Conversely, significant hearing recovery has not been reported in clinical trials for HC regeneration, although McLean et al. (2021) reported some beneficial effects . In addi