The possible involvement of alternative electron transport pathways, proton gradient regulator 5 (PGR5)-dependent CEF and plastid terminal oxidase (PTOX)-mediated electron transfer to oxygen in the response of Arabidopsis plants—wild type (wt) and lut2—to treatment with these two stressors was assessed by using specific electron transport inhibitors. Re-reduction kinetics of P700+ indicated that the capacity for CEF was higher in lut2 when this was compared to wt. Exposure of wt plants to the stress conditions caused increased CEF and was accompanied by a substantial raise in PGR5 and PTOX quantities. In contrast, both PGR5 and PTOX levels decreased under the same stress conditions in lut2, and inhibiting PGR5-dependent pathway by AntA did not exhibit any significant effects on CEF during the stress treatment and recovery period. Electron microscopy observations demonstrated that under control conditions the degree of grana stacking was much lower in lut2, and it almost disappeared under the combined stresses, compared to wt. The role of differential responses of alternative electron transport pathways in the acclimation to the stress conditions that are studied is discussed.A recent investigation has suggested that the enhanced capacity for PSI-dependent cyclic electron flow (CEF) and PSI-dependent energy quenching that is related to chloroplast structural changes may explain the lower susceptibility of  Plants are frequently facing adverse environmental stress conditions such as extreme temperatures, differences in optimal light illumination, high salinity conditions, dehydration, etc., that quite often act in a simultaneous manner and seriously affect the plant’s development and distribution. The effective functioning of photosynthetic reactions is largely dependent not only on the development conditions, but as well on the ability of plants to maintain a balance between the amount of absorbed sunlight and a capacity for its utilization through biochemical reactions. The exposure of plants to the simultaneous actions of increased light intensity and low temperature can lead to the over-reduction of the electron carriers of the photosynthetic electron transport chain ,2, to oxHigher plants have evolved a number of mechanisms that effectively dissipate the excess amounts of absorbed light for maintaining the balance between the absorbed energy and the biochemical sinks. It is general considered that the main photoprotective mechanism is the non-photochemical quenching of the excited chlorophyll states (NPQ), operating in the light-harvesting complex of the PSII (LHCII) that is dependent on the functioning of xanthophyll cycle and triggered by the buildup of the proton gradient across the thylakoid membrane ,8. Other2 fixation and the generation of ATP are limited, alternative electron transport flows around PSI are recognized as essential for the generation of the proton motive force, which is used for the induction of NPQ and photosynthetic control at the level of the cytochrome b6f (Cyt b6f) [6f that is coupled to the buildup of ΔpH is used for the synthesis of ATP [Under physiological conditions, the main electron transport pathway during photosynthesis is the linear electron transport chain (LET), but under stress conditions, in cyanobacteria, green alga and higher plants, when the COCyt b6f) ,17,18,19s of ATP ,21 or fos of ATP ,24,25,26s of ATP ,22,27. It is considered that, in angiosperms, CEF is realized by two pathways; one is sensitive to antimycin A (AntA) that involves the activity of ferredoxin plastoquinone (Fd-PQ) reductase and proton gradient regulation 5 (PGR5-mediated), and the second is related to NAD(P)H dehydrogenase-like protein (NDH-1 complex). Both pathways (PGR5- and NDH-1-dependent) of recycling electrons around PSI act in a complimentary manner , and thepgr5 mutant of Arabidopsis, indicating that the buildup of the membrane proton gradient was significantly impaired and that the PGR5/PGRL1-dependent pathway of electron recycling around PSI is essential for the dissipating of the excess absorbed light and protection of PSI against photodamage [The AntA-sensitive pathway of CEF  requirestodamage ,22. Nevertheless, the contribution of the NDH-1-dependent pathway to the overall CEF is considered to be minor but its involvement in the generation of the proton motive force under abiotic stress can be significant ,35,36. 2 for the reduction of O2 to water in chloroplasts that are in the dark. Despite this, dark chlororespiration is not directly related to the PSI-mediated alternative electron transport, as it shares electrons with PSI under light [Arabidopsis thaliana, the activity of CEF can be controlled at the level of PQ by PTOX [Arabidopsis is very low, comprising only 1% of the other electron transport components [Another enzyme that is related to CEF around PSI is the plastid terminal oxidase (PTOX), which represents a plastoquinone-oxygen oxidoreductase (diiron-containing protein). PTOX is involved in the process of chlororespiration, which is expressed in accepting electrons from PQHer light ,37. In t by PTOX . Under omponents ,39,40,41mponents ,43,44. Arabidopsis thaliana are investigated. One of the most frequently studied A. thaliana xanthophyll mutant is lut2, that, due to the inactivation of lycopene ɛ-cyclase gene, does not synthesize lutein, that is the most abundant xanthophyll in higher plants [lut1 and lut2 mutants leads to a compromised trimeric organization of the LHCII complexes, which negatively impacts the efficiency of NPQ [lut2 mutant [Carotenoids are an essential constituent of pigment-protein complexes that perform multiple functions. The oxygenated representatives of carotenoids, xanthophylls, are bound to the proteins of the LHCs of PSII and PSI, and each of them is characterized with an unique xanthophyll composition ,45. The r plants . The absy of NPQ . In the y of NPQ ,50. Rece2 mutant ,52.Arabidopsis thaliana plants—wt and lut2—to simultaneous treatment with two stress factors: a low temperature and a high light intensity, followed by a recovery period at control conditions. The importance of PTOX as a potential electron sink under the application of a double stress treatment was evaluated in parallel. Two specific electron transport inhibitors were applied—AntA for PGR5 and octyl gallate (OG) — for PTOX.The objective of the present study was to evaluate whether and to what extent the major alternative CEF, the PGR5-dependent pathway, is involved in the response and recovery of lut2 Arabidopsis thaliana mutants to the combined treatment of low temperature and high light intensity for different time periods was estimated by using specific electron transport inhibitors - AntA for PGR5-dependent CEF and OG for PTOX-dependent electron transfer to oxygen.The contribution of the alternative electron transport pathways to the photosynthetic response of PSII in wt and PSII) was observed after 6 days of treatment with two stress factors, while for lut2 discs the decline of this parameter was much stronger expressed and it occurred on the second day of the treatment  , a decrease of the quantum efficiency of the electron transport through PSII  c,d.- qP) were elevated in the wt and the lut2 over the course of treatment with two stress factors, reaching values of 0.35 for wt on the 6th day of the treatment and 0.75 for lut2 on the second day  increased in the wt discs on the 6th day of treatment and on the second day in the lut2 discs. ) were evaluated by following the far red (FR)-induced absorbance changes at 820 nm (ΔA820) that correlates with the degree of P700 oxidation to P700+. Similar to changes in the electron transport through PSII, as determined by ΦPSII and qP (700 in the wt discs that were floating on distilled water (Control) was detected on the 6th day of treatment and in lut2 discs, this was after 2 days  of the re-reduction of P700+, as a measure of CEF around PSI. The detected t1/2 in the wt discs from the non-treated plants, which were infiltrated only with water (Control), was 1.63 s and did not change considerably in the presence of AntA and OG. The t1/2 in the lut2 from the non-treated plants, which were infiltrated with water, showed a lower value of 1.05 s. The presence of both inhibitors accelerated the CEF after the treatment, with OG being more effective in the wt discs, and AntA realizing a stronger effect in the lut2 discs, with nearly 20%. The presence of both inhibitors accelerated the CEF in the lut2 discs from the non-treated plants, with nearly 20%. For the discs of the treated plants, AntA and OG were more effective in accelerating the rate of CEF in the wt than they were in the lut2 plants.lut2, non-treated plants, which were floated on water (Control), was around 30% lower than that in the wt discs  of the leaf discs of wt  for wt, while for the lut2 discs, this increase was slightly higher than the Fs . After 2 and 6 days of the treatment and in the absence of inhibitors, the transients for the wt showed a lower level of reduction, being the lowest on the 6th day of the treatment, representing 13% of the increase of Fo′ to Fs , but for the lut2 discs, the fluorescence transients demonstrated a low degree of reduction in the PQ pool (h).In the A quantitative immunoblot analysis of the thylakoid membranes that were probed with antibodies against PGR5 indicated a 132.7% and 126.6% increase of the relative abundance of PGR5 after 2 and 6 days of the combined stress treatment for the wt, respectively, followed by a decline that was close to the values of the control plants after 7 days, during the recovery period, after the stress treatment a.lut2 plants after 2 days of the treatment, but the effect was much less pronounced (114.8%) when it is compared to that of wt. Interestingly, PGR5 sharply decreased after 6 days of the stress treatment, and it remained low even after the recovery period  plants  and the ) plants a,b. The tructure c. In con2 mutant d when th2 mutant c. Thus, Arabidopsis thaliana and lutein-deficient mutant lut2 has shown that the quantum efficiency of non-photochemical quenching (ΦNPQ) and the energy-dependent component qE were lower in the mutants, in comparison with the wt at normal conditions. This could be related to the compromised three-dimensional structure and the organization of LHCII in the absence of lutein [lut2 plants showed a significant increase in the reduced pool of QA and PQ, which correlated with an enhanced intersystem electron pool size [700+) were diverted from the linear electron transport chain and recycled around PSI to provide protection to PSII against photoinhibition. It has to be pointed out that the lutein-deficient lut2 mutant demonstrated a higher capacity for CEF under the control growth conditions, in comparison with the wt [lut2 plants to a treatment of high light intensity and low temperature, specific electron transport inhibitors of the major alternative PGR5-dependent pathway (AntA) around PSI [A comparative investigation  of the pf lutein ,53,54,55ool size ,56. The h the wt ,52. To eound PSI ,57 and oound PSI ,59, werelut2 mutant of Arabidopsis thaliana have revealed that the missing lutein in the mutant was replaced by xanthophylls from the brunch of β-carotene synthesis from lycopene [lut2 plants and grown at control conditions, indicated that the fluorescence ratio F735/F685 in the mutant was higher than in the wt [lut2 was higher than it was in the wild type and/or more energy was delivered to PSI. Taken together, these results imply that in the lut2 mutant, the stroma-exposed thylakoid membranes, where the PSI complexes are located, prevailed in comparison with those in the wt. Indeed, the presented electron microscopic micrographs clearly demonstrated that the grana structures were less presented in the mutant, while the stroma-exposed thylakoid membranes prevailed in the lut2. The applied double stress differentially affected the ultrastructure of the thylakoid membranes. In the wt, the typical structural organization was not significantly altered, while in the lut2 plants, the population of the stroma-exposed thylakoid membranes in the mutant was further increased. Previous investigations of the lycopene . This sulycopene ,53,54,55n the wt ,60 suggePSII and qP) indicated that both plants, wt and lut2, suffered significant stress after the treatment with two stress factors. The negative effect was more strongly expressed and occurred earlier in the mutant. In addition, the performance of PSII was not completely restored in the lut2 plants upon their return to normal growth conditions. Application of AntA to the wt leaf discs increased the degree of ΦPSII inhibition, indicating that the function of the PGR5-dependent CEF is involved in the protection of PSII from photosynthetic activity under these conditions. The infiltration of leaf discs with AntA resulted in a significant increase in the excitation pressure of PSII in wt, emphasizing the role of the PGR5-dependent CEF for maintaining a more balanced ratio between the excitation and electron flows. In the C3 plants, the PGR5-dependent CEF is the major pathway [PSII. For the lut2 plants, the PGR5-dependent CEF does not appear to be so evident as it does in the wt whereas, the electron withdrawal through PTOX affected, more significantly, the response of qP to the high light intensity treatment, which was at a low temperature, especially after 2 days of the treatment. Higher plants rely on absorbing sunlight for performing the photosynthetic process. During exposure to abiotic stress conditions, especially at high light intensities and/or low temperatures, an imbalance can occur between the quantity of absorbed light and the temperature-dependent metabolic reactions, leading to the over-reduction of the electron carriers of the electron transport chain, photoinhibition and/or the photo-oxidative stress of PSII and PSI ,2. Indee pathway . Probabl pathway ,59. The  pathway . The imp pathway ,39,40. T700 oxidation was negatively affected by the double stress treatment and it occurred earlier in lut2 in comparison with wt. The application of both inhibitors, AntA and OG, emphasized that the PGR5-dependent CEF and the transfer of electrons from reduced PQ to oxygen by PTOX are important for the response of wt Arabidopsis thaliana to treatment with high light intensity, at a low temperature. For the lut2 plants the participation of the PGR5-dependent CEF in P700 oxidation did not seem to be so evident, but the involvement of PTOX was more strongly expressed in comparison with that of the wt. The extent of the P700 (t1/2) in the lut2 discs from the non-treated plants showed a lower value in comparison with that of the non-treated wt plants, indicating that the rate of CEF through PSI was operating faster in the mutant plants, as was shown previously [lut2. The half time of the re-reduction of Peviously ,52. The o′ was used to evaluate the extent of the dark reduction in the PQ pool [The post-illumination fluorescence increase of F PQ pool ,61. The  PQ pool . o′ in the non-treated, lut2 discs indicated that in the mutant discs, the electron pool in the intersystem electron chain was higher than it was in the wt. The alternative PGR5- and PTOX-dependent electron flows realized a small effect on the dark reduction in the PQ pool in the wt discs, while in the lut2 discs, the role of both alternative electron flows was much more strongly expressed. The extent of the PQ pool’s reduction in the lut2 discs with the blocked transfer of electrons from reduced ferredoxin to PQ (AntA-sensitive pathway) was one third of that for the non-treated and those lut2 discs which floated on water. The effect of the OG in the dark reduction of the transient increase of Fo′ was less expressed in comparison with that of the AntA. This is a clear indication that the contribution of the PGR5-dependent CEF to the intersystem electron transport chain was higher than that which was mediated by PTOX. Similar results in respect to diminishing the non-photochemical reduction in the PQ pool after treatment with AntA have been reported for Arabidopsis thaliana wt and crr2-2 and pgr5 mutants [Arabidopsis plants [lut2, but to different extents; in the lut2 discs, the decrease in the reduction state occurred earlier than it did in the wt, similar to alterations in the photochemical efficiency of PSII and P700 oxidation. The application of the AntA inhibitor decreased the reduction in the PQ pool, and this was stronger for the lut2 treated plants in comparison with the wt that were under the same conditions. For the wt, the presence of AntA led to a noticeable decline in the reduction state, while the OG increased this degree. This result clearly points out the significant contribution of the PGR5-dependent pathway for the redox state of the PQ pool.The transient increase in F mutants  and for s plants . With an2-dependent electron sink [lut2 non-treated plants, as well as in the recovered plants, was very low. This is in accordance with already published data, that under normal conditions the PTOX abundance is low and can account for less than 1% of all photosynthetic proteins [Arabidopsis [Arabidopsis plants was followed by its decrease near to the control values in the non-treated plants, and this indicated that the PTOX-mediated electron transfer to molecular oxygen was elevated under the combined action of a low temperature and a high light intensity. A similar significant up-regulation of PTOX was reported under various abiotic stress conditions [lut2 mutant was higher than it was in the wt, and it followed the same response to stress treatments, the abundance of both PGR5 and PTOX only marginally increased after 2 days of the combined stress treatment and it strongly decreased below the values of the control, non-treated, lut2 plants after 6 days of the stress treatment and remained lower even after recovery. In addition, inhibiting the PGR5-dependent pathway by AntA did not exhibit significant effects on CEF during the stress treatments and the recovery period in the lut2 plants. This suggests that the major FQR(PGR5)-dependent CEF pathway [lut2 mutant plants by an AntA-insensitive pathway [PTOX is involved in chlororespiration, carotenoid synthesis and controlling the chloroplast redox state by transferring excess electrons from a reduced PQ pool directly to the molecular oxygen, thus playing a key role in the Oron sink ,64. It wron sink . The overon sink . In agreproteins . In addiproteins  and Arabbidopsis  plants anditions ,40,41, anditions . The dyn pathway  might be pathway , and theArabidopsis thaliana, wt (Col-0) and mutant lut2, were grown on soil containing perlite at normal growth conditions—illumination with 100 μmol photons m−2 s−1 flux density (PFD) at day/night, temperature 20/18°C and humidity of 70%. The photoperiod was 12 h. After growth under these conditions for 3–4 weeks, fully developed plants were treated with low temperature (12/10 °C) and high light intensity (500 μmol photons m−2 s−1) for 6 days, followed by a period of 7 days at control conditions for the recovery period. For the whole experimental setup—plant development, treatment and recovery—plants were grown in growth chambers . Experiments for evaluation of the photosynthetic performance of PSII and PSI were performed before start of treatment, after 2 and 6 days of application of low temperature and high light intensity and after recovery period of 7 days. Two independent experiments were performed with at least 3 parallel samples at every time point. Plants of lut2 plants after the treatment with two stress factors—low temperature and high light—two different specific inhibitors of electron transport were applied: 5 µM antimycin A (AntA) for PGR5 [lut2 plants, 3 for every treatment, and were floated either on distilled water (Control), or on water containing the respective inhibitor at indicated concentration. So, prepared samples were gently shaken and illuminated with weak light intensity (70 μmol photons m−2 s−1) for 2 h before registration of photosynthetic parameters of PSII and PSIIn order to evaluate the contribution of PGR5- and PTOX-dependent pathways to the photosynthetic response of wt and for PGR5  and 10 µfor PGR5 ,59. Inhilut2 plants that were floated for 2 h either on distilled water (Control), or on water containing the respective inhibitor. Leaf discs were cut from leaves of plants that were non-treated, treated for 2 or 6 days and after a recovery period of 7 days. Measurements were performed at room (22 °C) temperature and ambient CO2 and O2 conditions using PAM 101–103 fluorometer , as described previously [o) was registered at illumination with weak (0.120 μmol photons m−2 s−1) modulated (1.6 kHz) light. To close all PSII centers and to register the maximal fluorescence in the dark-acclimated state (Fm), a saturating pulse of white light (3000 μmol photons m−2 s−1) with a duration of 0.8 s was given. Photosynthetic process was initiated with actinic light (AL) equal to the illumination of plants during growth conditions (100 μmol photons m−2 s−1). Every minute, a saturating pulse of white light was given to determine maximal fluorescence in light acclimated state (Fm′). After reaching the steady-state fluorescence level (Fs), the AL was switched off, and the minimal fluorescence in light acclimated state (Fo′) was detected. To determine the photosynthetic performance of PSII, the main fluorescence parameters were registered in the dark, acclimated for 15 min, and leaf discs of wt or eviously . The mino′ for 90 s [o′ post-illumination increase in Control and in the presence of AntA or OG, the transients were normalized from 1 (Fs) to 0 (Fo′). The alterations in dark non-photochemical reduction in the PQ pool by stroma reductants and/or by electrons from the intersystem electron pool were determined by registration of the post-illumination increase of transients of chlorophyll fluorescence level, after switching off AL and determination of Ffor 90 s ,61. For The following fluorescence parameters of PSII activity were calculated according to van Kooten and Snel (1990) . PSII = (Fm′ − Fs)/Fm′) [The photochemical efficiency of PSII was determined as ΦFs)/Fm′) .P = (Fm′ − Fs)/(Fm′ − Fo′).Coefficient of photochemical quenching was determined as qP = 1 − ((Fm′ − Fs)/(Fm′ − Fo′)) [Excitation pressure on PSII was determined as 1-qv′/Fm′) [The relative proportion of absorbed light and dissipated as heat in the antenna of PSII was calculated as 1-(Fv′/Fm′) .v′) was calculated as Fm′-Fo′.The variable fluorescence in light acclimated state  equipped with ED-800T emitter-detector unit [2 and CO2 conditions. To induce oxidation of P700, the leaf discs were illuminated using FR light . The redox state of P700 was registered as FR-induced absorbance change around 820 nm (ΔA820). When a steady state of FR-induced oxidation of P700+ was reached   on dark-tor unit  was used reached , multipl of PSII ,70, and ductants ,61,71.1/2) of the decay kinetics of re-reduction of P700+ after switching off FR illumination [700+ by stroma reductants [lut2 plant leaves.Detection of the half time . One leaf per plant and two plants per treatment were analyzed. Every leaf was cut into strips using a scalpel and were immediately fixed with 4% glutaraldehyde in 0.2 M cacodylate buffer (pH 7.2) for 1 h. The pre-treated samples were post-fixed for 2 h in 1% osmium tetroxide in 0.2 M Na-cacodylate buffer, dehydrated in a series of graded ethanol up to 100% and embedded in Durcupan  epoxide resin. Ultrathin sections (100 nm thickness) were sectioned using an ultra-microtome Reichert-Jung  and put on copper grids, stained with 1% uranyl acetate in 70% methanol and 0.4% lead citrate in dark. The sections were observed and photographed using an electron microscope at accelerating voltage of 220 kV. For every treatment, between 15 and 25 micrographs were taken.Alterations in chloroplast ultrastructure, as induced by exposure of wt and lut2 plants before start of treatment (0-day) and from plants grown under double stress for 2 and 6 days and after 7 days of recovery at control conditions were used [http://rsb.info.nih.gov accessed on 19 July 2022), as described earlier [The effect of growth at low temperature and high light intensity on PGR5 and PTOX relative abundance was evaluated by sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblot analysis. For the analysis a Laemmli SDS-PAGE system  was usedere used . Equal a earlier . The pren = 6). Statistically significant differences between the values of wt and lut2 plants in the studied variants were identified using two-way ANOVA followed by a Tukey’s post hoc test for each parameter. Prior to the test, the assumptions for the normality of raw data (using the Shapiro–Wilk test) and the homogeneity of the variances (using Levene’s test) were checked. The homogeneity of variance test was used to verify the parametric distribution of data. Values were considered statistically different with p < 0.05 after Fisher’s least significant difference post hoc test by using Origin 9.0 for data analysis and graphing software, version 9 .Two independent experiments were performed with at least 3 parallel samples per every experimental timepoint . For the wt A. thaliana, the application of the specific inhibitor AntA that blocks PGR5-dependent alternative electron flow resulted in an increased degree of PSII inhibition and an elevated excitation pressure on PSII. However, the effect of the OG that inhibits PTOX-dependent pathway was not so evident. For the lut2, the contribution of the PGR5-dependent pathway to photosynthetic performance was not so pronounced as it was in the wt, but the PTOX-mediated transfer of electrons to O2 evidently played a more significant role in the response to the exposure to two stress factors, as concluded from the more pronounced effect of the specific inhibitor, OG. The capacity for performing CEF was higher in the lut2 in comparison with the wt, and it was accelerated by the stress treatment, being better expressed in the wt. The inhibition of the PGR5-mediated pathway did not affect the kinetics of CEF in both the wt and lut2 plants. The blocking of PTOX realized a well pronounce effect on the rate of CEF, after the recovery period. The electron pool in the intersystem electron chain (reduction in PQ pool) in the non-treated lut2 was higher in comparison with that in the wt. The degree of reduction in the PQ pool was strongly affected by the growth rate under a high light intensity and at a low temperature, and it was higher in lut2. The blocking of PGR5 and PTOX by specific inhibitors further decreased the reduction in the PQ. The stronger effect that AntA had indicated a higher contribution of the PGR5-dependent pathway in the dark reduction in the PQ. The increased PTOX-mediated electron transfer in the course of treatment with the two stressors was confirmed by the increased abundance of this complex, evaluated by a western blot analysis. We suppose that the major PGR5-dependent CEF pathway in lut2 plants might be replaced by an AntA-insensitive pathway, and a withdrawal of excess electron relays on other-than-PTOX-mediated electron sink(s).The presented results have indicated that the stroma-exposed thylakoids prevailed in the


Oil pollution influences marine biology, ecology, and regional sustainable development capacity, but model uncertainties limit the ability of the numerical model to accurately predict the transport and fate of the underwater oil spill. Based on a three-dimensional underwater oil spill model validated by satellite images of the oil slick at the sea surface, the Penglai 19-3 oil spill accident in the Bohai Sea was simulated; in addition, several sensitivity experiments were set up to investigate the influence of model uncertainties in the background wind, current, start time of the oil spill, and spill site on the transport of underwater spilled oil in the Penglai 19-3 oil spill accident. The experimental results indicate that the uncertainty in the background wind has a certain impact on the simulated centroid position at the sea surface, and little effect on the simulated underwater results, while the uncertainty in the background current has a significant influence on the transport of the underwater spilled oil both at the sea surface and underwater. An uncertainty of 24 h in the start time of the oil spill can cause more than 1 time larger than the benchmark case displacement of the oil spill centroid point and sweeping area at the sea surface, as the periodic tidal current is the main constituent of the ocean current in the Bohai Sea. The uncertainty in the spill site has a large influence on the final position of the oil spill centroid point, but the oil spill trajectories do not intersect with each other within 48 h, which makes it possible to identify the oil spill platform from the actual observations. The influence of uncertainties in the important model inputs and key model parameters on the transport of underwater spilled oil in the Penglai 19-3 oil spill accident is evaluated for the first time, which is of substantial significance for improving the prediction accuracy of the transport and fate of underwater oil spills. Oil spill accidents have occurred frequently, seriously affecting the marine environment, beaches, and human activities ,2,3. In As indicated by Yang et al. , three gAn analysis of model uncertainties is required for reliable numerical simulation and prediction . ElliottAs we know, when the underwater spilled oil reaches the sea surface and is found by observation devices or by people, the oil spill accident has already started for some time. The start time and spill site of the underwater oil spill are often not known when predicting the transport of the oil spill. In addition, the background winds and currents have different characteristics in the different areas. The uncertainties in the wind, current, start time of the oil spill, and oil spill site will influence the simulated transport of the underwater oil spill, especially for serious accidents. In this study, based on the underwater oil spill model developed by Chen et al. , the PL12, situated in northeast China from 37° N to 41° N and from 117° E to 121° E [The Bohai Sea is a semi-closed shelf sea with a total area of 77,000 kmo 121° E ,24, as so 121° E .2 of sea water [As it has the second largest oil field and many oil resources, more than 50 offshore oil platforms have been established in the Bohai Sea . Severalea water  and causea water . So, theAccording to Zheng et al. , the dynThe plume dynamic model adopts the Lagrange integral technique, in which the oil spill duration is divided into several parts with the same time interval. Therefore, the spilled oil at the turbulent jet stage and the buoyant plume stage is assumed to be a bent cone, and is divided into a series of non-interfering elements according to the time interval . Each elDuring the turbulent jet and buoyant plume stages, the elements will maintain the conservation of mass, momentum, heat, salinity, and oil concentration. In addition, oil density variation due to temperature and the entrainment processes are also considered. The details of the plume dynamic model can be found in Chen et al. .bw is the buoyancy velocity of oil particle and is calculated with the oil droplet size using the methods in Yapa et al. [The advection-diffusion model adopts the Lagrangian particle-tracking method, in which the spilled oil is taken as a collection of oil particles. The distribution of oil particle size is estimated according to Delvigne and Hulsen , Wang eta et al. ; k→ is tdiffusionu, diffucionv and diffusionw are the components of the diffusion velocity in the x, y and z directions, respectively; xK, yK and zK are the components of the dispersion coefficient in the x, y and z directions, respectively, which can be calculated as described by Cao et al. [xR, yR and zR are independent random numbers ranging from −1 to 1 with uniform distribution; The diffusion velocity o et al. ; Rx, Ry windu and windv are the components of the oil drift velocity at the sea surface due to wind in the x and y directions, respectively; u10 and v10 are the components of the wind velocity at 10 m above sea surface in the x and y directions; α is the wind drift factor; β is the deviation angle, which is Besides the dynamic processes of the underwater oil spill, the spilled oil is also affected by the weathering processes. The spilled oil will be dissolved into the ambient environment and the mass is lost. In addition, the spilled oil at the sea surface will be evaporated into the air and the mass will be rapidly reduced. Besides, emulsification can also affect the mass and constituent of the spilled oil. The dissolution, evaporation, emulsification, and density variation of the spilled oil are simulated by the oil weathering model. The details of the oil weathering model can be found in Chen et al.  and Li eThe background wind and current are the important inputs of the underwater oil spill model, which are provided by Li et al. . The winmilation ,34. The milation .As shown in Xu et al. , the PL13 and the spilled oil was divided into 100,000 oil particles. For reducing the influence of the turbulent diffusion process, 100 scenarios, with different seeds when generating the random numbers in the random walk method, were conducted in Exp1.In benchmark experiment Exp1, the PL19-3 oil spill accident in the Bohai Sea was simulated using the underwater oil spill model. The PL19-3 oil spill site was located at the bottom of PL19-3 platform  with a depth of 24 m. The spill was assumed to start at 12:00 on 17 June 2011, and last for 1 h. The model duration was set to 48 h. The nozzle diameter was assumed to be 0.46 m and the initial eruption direction of the oil spill was vertically upward with spill velocity of 0.4 m/s. The oil density was set to 950 kg/msimuv is the wind speed (current speed) simulated by the numerical model as used in Exp1, r is the random number ranging from 0 to 1 with uniform distribution, e is the maximum random error, and uv is the wind speed (current speed) used in the corresponding sensitivity experiments.Model uncertainties, including the uncertainty in background wind, background current, start time of the oil spill, and spill site, are important factors affecting the simulated results of underwater oil spills. Four groups of sensitivity experiments were designed to study the influence of the uncertainties on the simulated results. As simulated by the numerical model, the background wind and current are usually inaccurate, contributing to the uncertainties of the underwater oil spill model. In the sensitivity experiments numbered with the prefix ‘SE1’, the influence of the uncertainty in background wind on the simulation results was discussed by adding artificial random perturbations in the background wind. In sensitivity experiments SE1_1, SE1_2, and SE1_3, the maximum random errors were 10%, 20%, and 30%, respectively. The artificial random perturbations were added into the background wind using Equation (4), as follows:In the sensitivity experiments numbered with the prefix ‘SE2′, the influence of the uncertainty in the background current on the simulation results was discussed by simultaneously adding artificial random perturbations in the background wind and current by using Equation (4). In sensitivity experiments SE2_1, SE2_2, and SE2_3, the maximum random errors were 10%, 20%, and 30%, respectively. It was noted that the random numbers were generated using different seeds when adding artificial random perturbations into the wind and current.In sensitivity experiments numbered with ‘SE3′, the influence of the uncertainty in the start time of the oil spill on the simulation results was discussed. Compared with Exp1, the start time of the oil spill was delayed from 1 to 24 h; in addition, the model duration was 24 h.In sensitivity experiments numbered with the prefix ‘SE4′, the influence of the spill site uncertainty on the simulation results was discussed. In sensitivity experiments SE4_1, SE4_2, SE4_3, and SE4_4, the site of the oil spill was changed from PL19-3 to BZ29-4, BZ34-1, BZN28-2, and KL10-1, respectively.EE is the error expectation, which is the average difference at time t between the simulated results of 100 scenarios in one experiment and the scenarios-average simulated result in Exp1; t; t; RU is the result uncertainty, which is the standard deviation of the 100 simulated results at time t in one experiment, and indicates the dispersion degree because of the model uncertainty; The detailed model settings of the sensitivity experiments are listed in The simulated centroid position and the oil spill trajectory in the PL19-3 oil spill accident obtained in Exp1 are shown in As shown in 2 (2.61 km2), 2.08 km2 (2.75 km2), and 2.38 km2 (3.01 km2), respectively. As the uncertainty in the background wind increases from SE1_1 to SE1_3, the average errors and uncertainty degrees of the underwater sweeping area show no significant difference (The error expectation of sweeping area (EESA) and the result uncertainty of sweeping area (RUSA) at the sea surface and underwater are calculated and shown in fference b,d.3 (0.72 km3), 0.66 km3 (0.81 km3), and 0.80 km3 (0.94 km3). With the increase of the uncertainty in wind, the average error (uncertainty degree) of the sweeping volume increases by 0.00 times (0.01 times), 0.12 times (0.14 times), and 0.47 times (0.32 times) in SE1_1, SE1_2, and SE1_3, respectively, compared with the value in Exp1.The error expectation of sweeping volume (EESV) and the result uncertainty of sweeping volume (RUSV) are calculated and shown in Overall, when the maximum random error in wind is 30%, the deviation of the simulated centroid position at the sea surface is about 0.10 km, and EECS at sea surface is 44.08 times that in Exp1; however, the other differences of underwater centroid position, sweeping area, and sweeping volume are insignificant, showing that the uncertainty in the background wind has a certain impact on the simulated centroid position at the sea surface, and little effect on the simulated underwater results.When the artificial random perturbations are simultaneously added into the background wind and current in SE2_1, SE2_2, and SE2_3, the simulated mean centroid positions in the 100 scenarios are near to those in Exp1, but the dispersion degree of those centroid positions increases, as shown in As shown in 3 (1.44 km3), 4.25 km3 (3.33 km3), and 9.36 km3 (4.97 km3), respectively. With the increase of the uncertainty in the background wind and current, the average error (uncertainty degree) of sweeping volume increases by 1.48 times (1.21 times), 6.87 times (4.10 times), and 16.33 times (6.61 times) relative to that in Exp1, which is significantly higher than the corresponding results in the experiments numbered with the prefix ‘SE1’.In summary, when the uncertainties in the background current and wind are present simultaneously, the influence on the simulated centroid position, sweeping area, and sweeping volume is significantly greater than when the only the uncertainty is in the background wind at the same error level. The influence of the uncertainty in the background current on the transport of the underwater oil spill is greater than that of the uncertainty in the background wind, which is important for forecasting the transport and fate of the underwater spilled oil.As the transport time of oil/water plume is usually just several minutes from well head to sea surface , severalAs shown in 2 at a delay of 24 h.As shown in As discussed above, the uncertainty of the oil spill start time has a significant influence on the transport of the oil slick at the sea surface. An uncertainty of 24 h can cause 1.19 times larger than before displacement of the oil spill centroid point at the sea surface, and nearly 1-time larger than before sweeping area at the sea surface. Therefore, accurately knowing the start time of the oil spill and its degree of uncertainty is of substantial significance to the simulation and prediction of the transport of marine oil spills and has a significant impact on the planning of the emergency response to the spilled oil.When the underwater oil spills occur on different oil platforms, the centroid trajectories of the oil spill at the sea surface have similar trend a, mainlyOil pollution has an adverse influence on the marine environment, ecology, and regional sustainable development capacity . The numElliott and Jones  indicateWith the progress of observation technology and data processing methods, there are many more means to obtain information on the spilled oil. For example, Dong et al.  developeBased on a three-dimensional underwater oil spill model validated by satellite images of the oil slick at the sea surface, the Penglai 19-3 oil spill accident in the Bohai Sea was simulated; in addition, several sensitivity experiments were set up to investigate the influence of model uncertainties on the simulated oil spill trajectory at the sea surface, the centroid position and sweeping area at the sea surface and underwater, and the sweeping volume. When the maximum random error in wind is 30%, the error expectation of centroid shift at the sea surface is 44.08 times of that in the benchmark experiment, while the uncertainty in the wind has little effect on the simulated underwater results, as the wind mainly drives the oil drift at the sea surface. When the uncertainty in the current is considered together with the uncertainty in the wind, the influence on the simulated transport of the underwater spilled oil is significantly greater than with only the uncertainty in the wind, showing the importance of the current for the accurate simulation of an underwater oil spill. As the tidal current is the significant constituent of the current in the Bohai Sea and its periodicity, the start time of the oil spill has a significant influence on the transport of the oil; in detail, an uncertainty of 24 h can cause 1.19 (1) times larger than before displacement of the oil spill centroid point (sweeping area) at the sea surface. The uncertainty of the spill site has little impact on the pattern of the simulated oil spill trajectory at the sea surface, and has large influence on the final position of the oil spill centroid point.When considering the numerical simulation and prediction of actual oil spill accidents, the model uncertainty should be considered adequately, which is of substantial significance in improving the prediction ability of underwater oil spills. This study concludes that the current and the start time of an oil spill have significant influence on the transport of underwater spilled oil, while wind would mainly impact the surface results; in addition, it is possible to identify the oil spill platform from the actual observations of the oil slick at the sea surface obtained shortly after the oil spill has occurred.


To evaluate the changes of rectus abdominis thickness and inter-rectus distance before and after delivery with high-frequency ultrasound.  A total of 148 pregnant women at 12 weeks of gestation who underwent prenatal examination in our hospital from January 2019 to March 2020 were selected, and 140 of them cooperated with rectus abdominis examination. According to the results of rectus abdominis examination 42 days after delivery, 97 patients were divided into the DRA group with rectus abdominis isolated and 43 patients were divided into the normal group with rectus abdominis not isolated. At 12 weeks, 24 weeks, and 37 weeks of pregnancy, 3 days and 42 days after delivery, the thickness and spacing of the left and right rectus abdominis muscle were measured by high-frequency ultrasound along the white linea at three positions: 5 cm above the navel, 3 cm below the umbilical edge, and 3 cm below the navel. P < 0.05). The space between rectus abdominis 5 cm above the navel, 3 cm below the navel, and the navel margin of the abdominal white line in the pregnant women of the two groups was gradually increased with the increase of the pregnancy cycle and gradually recovered after delivery. At 37 weeks of pregnancy, 3 days after delivery, and 42 days after delivery, the space of rectus abdominis along the umbilicus 5 cm above, 3 cm below the umbilicus, and the umbilicus border of the abdominal white line in the DRA group was significantly larger than that of the normal group (P < 0.05).  The thickness of rectus abdominis at 5 cm above the navel, 3 cm below the navel, and at the navel margin of the abdominal white line in the pregnant women of the two groups was gradually decreased with the increase of the pregnancy cycle and gradually recovered after delivery. At 42 days after delivery, the thickness of rectus abdominis in the DRA group was significantly lower than that in the normal group, which was 5 cm above the umbilicus, 3 cm below the umbilicus, and the umbilical margin of the abdominal white line ( Ultrasound can accurately measure the inter-rectus distance and rectus thickness, accurately evaluate the degree of DRA, and realize the one-stop evaluation from prenatal diagnosis and prediction to postpartum rehabilitation monitoring, so as to intervene during pregnancy and reduce the risk of postpartum DRA. The rectus abdominis of the human body fuses in the midline of the abdomen, and the distance between the rectus abdominis of both sides is no more than 1–2 cm in the normal nonpregnant state. Diastasis recti abdominis (DRA) is caused by acquired factors that cause the separation of the rectus abdominis muscle beyond the normal width along the long axis of the muscle and is characterized by abnormally increased inter-rectus distance, resulting in relaxation and bulging of the abdominal musculature . Women aAt present, the clinical diagnosis of DRA is mainly based on finger-width palpation, but this method has large subjective errors. CT and MRI are also used for DRA diagnosis and evaluation, but the cost is high, and CT has radiation and is not suitable for pregnant women, limiting its clinical application. In recent years, ultrasound has been widely used to detect rectus abdominis and pelvic floor muscles, and its accuracy has been widely recognized –11. In tA total of 148 pregnant women who underwent prenatal examination and filing in our hospital at 12 weeks of gestation from January 2019 to March 2020 were selected. Of the 148 pregnant women, 140 pregnant women cooperated to complete the rectus abdominis test, 5 pregnant stopped for fetal reasons, and 3 pregnant dropped out of the study. According to the results of rectus abdominis examination 42 days after delivery, 97 patients were divided into the DRA group with rectus abdominis separation and 43 were divided into the normal group without rectus abdominis separation.Singleton and full-term pregnancy; there are no previous reproductive history; there are no complications such as pregnancy-induced hypertension and gestational diabetes mellitus; carry out prenatal examination and delivery in our hospital, and voluntary cooperation with the rectus abdominis examination during pregnancy and postpartum; there is no history of abdominal and pelvic surgery; there is no urinary tract infection during the study.Previous history of DRA diagnosis; congenital abdominal wall muscle or nerve defects; previous history of abdominal hernia, urachal fistula, etc.; and inability to lie supine or cooperate with the study due to low back pain.GE Voluson E8 color Doppler ultrasonic diagnostic apparatus was used with a probe frequency of 7.5 to 12 MHz. Subject level Lie down, place both upper limbs flat on both sides of the body, flex the lower limbs to relax the abdominal muscles, and breathe calmly. The ultrasound probe was placed in the midline of the abdomen, and measurements were made along the white line at three positions: 5 cm above the umbilicus, at the edge of the umbilicus, and 3 cm below the umbilicus along the linea alba. The thickness and spacing of the left and right rectus abdominis were measured by high-frequency ultrasound.At 12 weeks, 24 weeks, and 37 weeks of pregnancy, 3 days and 42 days after delivery, a high-frequency ultrasound probe was used and was perpendicular to the long axis of the white line and aimed at the distance between the straight muscles on both sides The plane of the probe was perpendicular to the long axis of the rectus abdominis. At the end of each measurement, the still images are collected immediately. The inter-rectus distance is the distance between the hypoechoic medial edges of the rectus abdominis on both sides, the left and right rectus abdominis thickness is the distance between the anterior and posterior fascia of the rectus abdominis, and the maximum diameter is the rectus abdominis thickness. All rectus abdominis examinations were performed by the same attending sonographer, they were measured three times, and an averaged value was taken. Diagnostic criteria of DRA: the lesion site of DRA was divided into infraxiphoid, epigastrium, umbilicus, subumbilicus and above pubic symphysis. DRA was diagnosed when the inter-rectus distance was >20 mm at rest in any part.s). T-test was used to compare the measurement data between groups, and analysis of variance was used to compare multiple groups. The natural birth rate and other counting data are expressed in (%) and compared by pairs χ2 inspection. The test level is α = 0.05, and P < 0.05 was considered statistically significant.SPSS 22.0 software was applied for processing. Age, prepregnancy BMI, postpartum BMI, weight change during pregnancy, abdominal circumference, and other experimental data were expressed by mean ± standard deviation (P < 0.05). There was no significant difference in maternal age, prepregnancy BMI, and fetal weight between the two groups (P > 0.05), as shown in Postpartum BMI, gestational weight change, and abdominal circumference were significantly greater and the rate of spontaneous delivery was significantly lower in the DRA group than in the normal group, and the differences were statistically significant (P > 0.05). In the DRA group, the thickness of the left and right rectus abdominis 5 cm above the umbilicus along the linea alba at 42 days postpartum was significantly lower than that in the normal group, and the differences were statistically significant (P < 0.05).The thickness of the left and right rectus abdominis 5 cm above the umbilicus along the linea alba increased gradually with the pregnancy cycle in both groups. Recovery began postpartum, with left and right rectus muscle thicknesses still lower at 42 days postpartum than at 12 weeks gestation. In the DRA group, there was no significant difference in the thickness of the left and right rectus abdominis 5 cm above the umbilicus along the white line at 12 weeks of pregnancy, 24 weeks of pregnancy, 37 weeks of pregnancy, and 3 days postpartum compared with the normal group (P > 0.05). The inter-rectus distance of 5 cm above the umbilicus along the linea alba at 37 weeks of gestation, 3 days postpartum, and 42 days postpartum in the DRA group was significantly greater than that in the normal group, and the differences were statistically significant (P < 0.05). Figures 1The rectus abdominis distance of 5 cm above the umbilicus along the linea alba increased gradually with the pregnancy cycle in both the groups. Recovery began postpartum, and the abdominal straight muscle distance was still greater at 42 days postpartum than at 12 weeks of gestation. There was no significant difference in the distance between the rectus abdominis 5 cm above the umbilicus along the linea alba at 12 and 24 weeks of pregnancy between the DRA group and the normal group (P > 0.05). In the DRA group, the thickness of the left and right rectus abdominis 3 cm below the umbilicus along the linea alba at 42 days postpartum was significantly lower than that in the normal group, and the differences were statistically significant (P < 0.05).The thickness of the left and right rectus abdominis 3 cm below the umbilicus along the linea alba increased gradually with the pregnancy cycle in both the groups. Recovery began postpartum, with left and right rectus muscle thicknesses still lower at 42 days postpartum than at 12 weeks gestation. In the DRA group, there was no significant difference in the thickness of the left and right rectus abdominis 3 cm below the umbilicus along the linea alba at 12 weeks of pregnancy, 24 weeks of pregnancy, 37 weeks of pregnancy, and 3 days postpartum compared with the normal group (P > 0.05). The inter-rectus distance 3 cm below the umbilicus along the linea alba at 37 weeks of gestation, 3 days postpartum, and 42 days postpartum in the DRA group was significantly greater than that in the normal group, and the differences were statistically significant (P < 0.05).The rectus abdominis distance 3 cm below the umbilicus along the linea alba increased gradually with the pregnancy cycle in both the groups. Recovery began postpartum, and the abdominal straight muscle distance was still greater at 42 days postpartum than at 12 weeks of gestation. There was no significant difference in the rectus abdominis distance 3 cm below the umbilicus along the linea alba at 12 and 24 weeks of pregnancy between the DRA group and the normal group (P > 0.05). In the DRA group, the thickness of the left and right rectus abdominis in the umbilical border of the linea alba at 42 days postpartum was significantly lower than that in the normal group, and the differences were statistically significant (P < 0.05).The thickness of the left and right rectus abdominis in the umbilical border of the linea alba gradually decreased with the increase of the pregnancy cycle in both the groups. Recovery began postpartum, with left and right rectus muscle thicknesses still lower at 42 days postpartum than at 12 weeks gestation. In the DRA group, there was no significant difference in the thickness of the left and right rectus abdominis of the umbilical border of the linea alba compared with the normal group at 12 weeks of pregnancy, 24 weeks of pregnancy, 37 weeks of pregnancy, and 3 days postpartum (P > 0.05). In the DRA group, the rectus abdominis intermuscular distance at the umbilical border of the linea alba was significantly greater than that in the normal group at 37 weeks of gestation, 3 days postpartum, and 42 days postpartum, and the differences were statistically significant (P < 0.05).The rectus abdominis interval of umbilical line of linea alba increased gradually with the increase of pregnancy cycle in both the groups. Recovery began postpartum, and the abdominal straight muscle distance was still greater at 42 days postpartum than at 12 weeks of gestation. There was no significant difference in the rectus abdominis distance between the umbilical line of the linea alba at 12 and 24 weeks of gestation in the DRA group and the normal group (The rectus abdominis is an important muscle structure of the abdominal wall, which is located on both sides of the midline of the anterior abdominal wall, separated by a white line in the middle. Therefore, the position of the abdominal organs will not be affected by the change of gravity and the pressure in the spine, plevis, and abdomen, and assisting normal physiological functions, such as breathing, defecation, and delivery , 13. TheUltrasound examination is widely recognized in clinical practice because of its stability, consistency, and reproducibility, and is considered as the gold standard for noninvasive evaluation of DRA. High-resolution probes can directly measure the inter-rectus distance and observe its maximum separation position. Double stitching, virtual wide field of vision, continuous wide field of vision and other imaging are feasible for patients with wide inter-rectus distance in the third trimester –21. UltrThere are also multiple studies on the postpartum rectus abdominis in China, but there are few studies on the postpartum period of the same individual from pregnancy. In this study, we measured the thickness of rectus abdominis and inter-rectus distance before and after delivery by ultrasound to evaluate the changes of rectus abdominis throughout pregnancy and after delivery. It was found that the thickness of rectus abdominis 5 cm above the umbilicus and 3 cm below the umbilicus along the linea alba and at the umbilical line of the linea alba in the two groups decreased gradually with the increase of pregnancy cycle and gradually recovered after delivery. However, the thickness of the rectus abdominis at 42 days postpartum in the DRA group was significantly lower than that in the normal group. With the increase of pregnancy cycle, the inter-rectus distance gradually increases, and it gradually recovered after delivery. At 37 weeks of pregnancy, 3 days postpartum, and 42 days postpartum, there was a significant difference in rectus muscle spacing between the two groups. These results suggest that the inter-rectus distance can directly reflect the situation of DRA, and has an effect on the rectus abdominis at umbilical level. Under the action of estrogen and progesterone, the collagen fibers of pregnant women will gradually expand and stretch, weakening their muscle strength to meet the needs of delivery, while the gradually enlarging fetus and uterus will press pelvic floor muscle tissue and rectus abdominis for a long time, resulting in the damage of muscle tissue, blood vessels, and nerve, and destroying the support system of their abdominal and pelvic organs –24. On tThis study is a measurement study of DRA from pregnancy to a short time after delivery, which has some limitations. The recovery time of rectus abdominis after delivery is long, and the its influencing factors need further study. At the same time, it is necessary to accumulate cases and conduct large sample, multi-center, and high-quality research.In summary, ultrasonography can accurately measure inter-rectus distance and rectus thickness, accurately assess the degree of DRA, and achieve a one-stop assessment from prenatal diagnosis prediction to postpartum rehabilitation monitoring for pregnancy intervention.


It was found that NQ at 0.1 and 1 nM and DHNQ at 0.1 nM significantly stimulated the fresh weight of the aboveground parts of the seedlings (APS), while the fresh weight of the underground parts of the seedlings (UPS) was enhanced only at 0.1 nM NQ. Interestingly, DHNQ at higher concentrations (1 and 10 nM) significantly diminished the fresh weight of the APS and UPS. When NQ and DHNQ were applied together, an increase in the fresh weight of the APS at all of the concentrations studied was observed. It was also found that NQ and DHNQ individually and in combination, at all concentrations studied, decreased the H2O2 production in the aboveground and underground parts of maize seedlings. The presence of the DHNQ at higher concentrations (1 and 10 nM) triggered an increase in the catalase (CAT) activity of the UPS and APS compared to the control. However, NQ added at 1 nM decreased the CAT activity of both the UPS and APS, while 10 nM increased the CAT activity of UPS. NQ and DHNQ applied together at 0.1 and 10 nM almost completely inhibited catalase activity in the UPS and APS. The data that were obtained for lipid peroxidation, measured as the malondialdehyde (MDA) concentration, indicated that NQ and DHNQ at all concentrations studied decreased the MDA content of the UPS, while both naphthoquinones increased it in APS. The data presented here are discussed taking into account the mechanisms via which naphthoquinones interact with biological systems.This study investigated the effects of 1,4-naphthoquinone (NQ) and naphthazarin  individually and in combination, applied at low concentrations , on growth, hydrogen peroxide (H Naphthoquinones are the products of bacterial and fungal metabolism, as well as secondary metabolism in higher plants, where they are produced and used as natural defense chemicals ,2,3,4. NBoraginaceae, Droseraceae, and Nepenthaceae families [Naphthazarin is one of the natural 1,4-naphthoquinone substances derived from the tissues of several members of the families ,14  changed the auxin-induced reorientation of the cortical microtubules from perpendicular to oblique with respect to the long cell axis, which, according to the authors, suggests that this effect may be related to the nucleophilic interaction of naphthazarin with the proteins that are associated with the cortical microtubule reorientation processes.There are two general action mechanisms of naphthoquinones; one is the covalent modification of biological molecules at their nucleophilic sites, such as the thiols in proteins and glutathione (GSH), in which quinones act as electrophiles, while the other mechanism consists of redox cycling, in which reactive oxygen species (ROS) are generated ,27,28,292O2 production and catalase activity) in maize seedlings, and (3) examining the effects of NQ, DHNQ, and NQ plus DHNQ on the malondialdehyde (MDA) content of maize seedlings. This experimental design can provide new data on the effects of naphthoquinones on the growth of plant seedlings.Bioassays that use plant growth and other physiological processes are often the primary tools for determining the biological activity of natural and synthetic compounds. The main goal of the present study was to determine the effects of 1,4-naphthoquinone (NQ) and naphthazarin  individually and in combination, both at low concentrations , on the growth of maize seedlings and shed light on the mechanism of this phenomenon. This goal was realized by (1) studying the effect of both naphthoquinones and their combination on the length of maize seedling organs  and fresh weight of the above- and underground parts of the seedlings, (2) determining the redox cycling properties of NQ and DHNQ individually and in combination  were selected taking into account our previous papers ,17,18 inThe treatment of maize seedlings with 1,4-naphthoquinone (NQ) at concentrations of 0.1, 1, and 10 nM did not significantly change the growth of the main roots and mesocotyls, while, at 0.1 nM and 1 nM, NQ stimulated the growth of the first leaves by about 40–50% . In the In order to more quantitatively verify the results obtained from the measurements of the elongation growth of the seedling organs, the fresh weight of the aboveground parts of the seedlings (APS) and underground parts of the seedlings (UPS) was also determined . In subsThe treatment of maize seedlings with NQ at 0.1 and 1 nM stimulated a ca. 40% increase in the fresh weight of the aboveground parts of the seedlings (APS) compared to the control, while 10 nM NQ did not significantly change it. In the case of DHNQ, the fresh weight of the APS was stimulated by 20% only at 0.1 nM; however, at higher concentrations, 1 and 10 nM, DHNQ did not change the fresh weight of the APS and inhibited it (by ca. 40%), respectively, compared to the control. When NQ and DHNQ were added together at 1 nM, a stimulatory effect of both naphthoquinones on the fresh weight of the (APS) was observed; however, when NQ and DHNQ were added together at 10 nM, the toxic effect of DHNQ on the fresh weight of the APS was inhibited. In the case of the fresh weight of the underground part of the seedlings (UPS), an increase was observed only in the presence of 0.1 nM NQ, while, at higher concentrations, NQ did not change this parameter. DHNQ significantly lowered the fresh weight of the UPS at higher concentrations (1 and 10 nM). However, when NQ and DHNQ were added together at 1 and 10 nM, the toxic effect of DHNQ, in contrast to the fresh weight of the APS, was inhibited only at 1 nM DHNQ.2O2) to lower values compared to the control medium  was greater than that in the underground parts of the seedlings (UPS). For example, when NQ or DHNQ was added at concentrations higher than 0.1 nM, the production of H2O2 in the APS was at least twofold greater than in the UPS. DHNQ added at concentrations higher than 0.1 nM enhanced H2O2 production, although it was still lower than in the control. The same was observed for NQ added at 10 nM. Interestingly, NQ and DHNQ added together at all of the concentrations studied almost completely inhibited the production of H2O2 in the UPS, while, for APS, the production of H2O2 was decreased to values lower by almost 60% compared to the control.In all experimental variants, the production of H2O2. The presence of NQ or DHNQ in the control medium at the lowest concentration (0.1 nM) did not change the catalase (CAT) activity in the UPS, whereas it significantly reduced it in the APS compared to the control  in the UPS and APS compared to the control. The presence of DHNQ at 1 and 10 nM triggered an increase in the catalase activity of the UPS and APS by about 20% and 100%, respectively, compared to the control. However, NQ added at the same concentrations as DHNQ at 1 nM decreased the CAT activity of both the UPS and the APS by ca. 50%, while 10 nM increased the CAT activity of the UPS. NQ and DHNQ added in combination inhibited the CAT activity at almost all concentrations tested, excluding variant NQ + DHNQ at 1 nM for APS .Lipid peroxidation in the UPS and APS treated with NQ and DHNQ individually and in combination, which was measured as the malondialdehyde (MDA) concentration, is presented in The data obtained indicate that NQ and DHNQ at all concentrations studied decreased the MDA content of the UPS, while they increased the MDA content of the APS compared to the control (seedlings not treated with NQ or DHNQ). Interestingly, DHNQ at 0.1 and 10 nM increased the MDA content of APS much more than NQ applied at the same concentrations. When both naphthoquinones were added together at 0.1 and 1 nM, their combined effect resulted in both a decrease in MDA content of the UPS to values close to 0% and a decrease in MDA content of the APS compared to the control. The combined effect of both naphthoquinones applied at 10 nM resulted in stimulation of the UPS and APS by ca. 40% and 60%, respectively, compared to the control.2O2 and catalase activity, while NQ at 10 nM only significantly changed the lipid peroxidation. NQ at all concentrations studied caused an increase in lipid peroxidation (measured as MDA concentrations). In contrast to NQ, DHNQ at higher concentrations (1 and 10 nM) decreased the fresh weight of the APS, accompanied by a decrease in the production of H2O2 and an increase in catalase activity. As in the case of NQ, DHNQ enhanced lipid peroxidation. When NQ and DHNQ were applied together, an increase in the fresh weight of the APS at all concentrations studied was observed. As in the case of NQ, the increase in fresh weight was accompanied by a marked decrease in H2O2 production and catalase activity. The combination of both naphthoquinones at 0.1 and 10 nM significantly stimulated the MDA content of the APS.Considering the data presented in 2O2 in the UPS was reduced, especially under the combined action of NQ and DHNQ, where it was practically reduced to zero. Moreover, as in the case of the APS, catalase activity in the UPS was stimulated at higher concentrations of DHNQ (1 and 10 nM) and inhibited, especially in the case of the combined action of NQ and DHNQ. In contrast to APS, the MDA concentration in the UPS was decreased at all concentrations of NQ and DHNQ individually and in combination, excluding the variant of NQ and DHNQ added together at 10 nM.Comparing the abovementioned data obtained for APS with those for UPS, it should be stated that NQ at 0.1 nM increased the fresh weight of the UPS, while, at higher concentrations, no change was observed. In turn, DHNQ at 1 and 10 nM, similar to the case of the APS, diminished the fresh weight of the UPS. NQ and DHNQ applied together at lower concentrations slightly stimulated the fresh weight of the UPS. As in the case of the APS, the production of HIn recent years, a great deal of attention has been paid to naturally occurring naphthoquinones as compounds with a high level of biological activity. The toxicity of naphthoquinones is mainly due to two major types of reaction. First, as redox cyclers, they induce oxidative stress by generating reactive oxygen species (ROS); second, as electrophiles, they react with nucleophiles, such as thiols or glutathione ,29. It i2O2 production in maize coleoptile cells, which closely contributed with an increase in catalase activity (CAT) and lipid peroxidation. Interestingly, when 1,4-naphthoquinone (NQ) and naphthazarin (DHNQ) were applied together at 1 and 10 nM, their inhibitory effect on the auxin-induced growth and proton extrusion of the maize coleoptile cells was stronger compared to the individual action of either of the naphthoquinones. In the presence of both naphthoquinones at 1 nM, the oxidative stress was significantly higher. It should be added here that the segments of etiolated maize coleoptiles (cut 3–5 mm below the tip of the coleoptiles) represent a classical model system for studies on the mechanism of auxin action on plant cell growth. In this system, the number of cells is constant, and an organ grows only through elongation. Interestingly, most of the important evidence on the molecular mechanisms of auxin action was obtained in experiments performed with segments of maize coleoptiles .The main goal of the experiments described here was to shed light on the effects of 1,4-naphthoquinone (NQ) and its derivative naphthazarin  individually and in combination, applied at low concentrations , on the growth and oxidative stress in the intact maize seedlings. Understanding these effects is important due to the allelochemical properties of naphthoquinones and their effect on plant growth and development.2O2 production, catalase activity, and lipid peroxidation in maize seedlings.Taking into account the data presented above for the fresh weight, it can be suggested that NQ at 0.1 and 1 nM and DHNQ at 0.1 nM significantly stimulated the fresh weight of the APS, while, in the UPS, it was changed only slightly. Interestingly, DHNQ at higher concentrations (1 and 10 nM) significantly diminished the fresh weight of the APS and UPS. The observed differences between APS and UPS probably resulted, among other reasons, from the fact that, in the case of the UPS, the naphthoquinones directly interacted with the root system. This means that NQ and DHNQ could change root growth by affecting two processes: cell division and cell expansion. However, in order to at least partially explain the mechanisms via which naphthoquinones induced changes in the fresh weight of the seedlings, we also took into account the redox cycling properties of both naphthoquinones, determined as the H2O2 production in the aboveground and underground parts of maize seedlings compared to the control  by others [The data presented in  control . The hig control . This oby others ,33,34  plays a key role in maintaining the balance between Hf plants ,36. The  control . Due to Reactive oxygen species (ROS) are products of membrane lipid peroxidation, and MDA content can indicate the degree of lipid peroxidation damage . Under nIn conclusion, this study sheds new light on the effects of 1,4-naphthoquinone and naphthazarin alone and in combination on the growth and oxidative stress of maize seedlings, suggesting the use of both naphthoquinones at low concentrations as growth biostimulants.Zea mays L. cv. Cosmo 230) caryopses were soaked in tap water for 2 h, sown on wet lignin in plastic containers, and placed in a growth chamber  at 27 ± 1.0 °C. Four day old seedlings with similar lengths (50–70 mm) were transferred to 2.4 L opaque plastic containers (nine seedlings in each) filled with Hoagland’s solution [All experiments were conducted on 21 day old maize seedlings grown hydroponically. Briefly, maize  were determined with a ruler. The fresh weight of the aboveground and underground parts of the seedlings was then determined using a laboratory balance .2O2) in roots and shoots of maize seedlings was determined according to Velikova et al. [w/v) trichloroacetic acid (TCA). The homogenate was centrifuged at 10,000× g at 4 °C for 10 min. Then, 0.5 mL of the supernatant was added to 0.5 mL of 0.1 M K-phosphate buffer (pH 7.0) and 1 mL of 1 M KI. The absorbance of the supernatant was measured at 350 nm. The H2O2 content, expressed in µmol/g fresh weight (FW), was calculated from a standard calibration curve prepared in 0.1% TCA at different H2O2 concentrations.Hydrogen peroxide  TCA and 0.01% butylated hydroxytoluene, or (2) +TBA solution containing the above plus 0.65% TBA. Samples were then vigorously mixed, heated at 95 °C in a boiling water bath for 20 min, cooled, and centrifuged at 10,000× g at 4 °C for 10 min. Absorbance values were read at 440, 532, and 600 nm. MDA equivalents (µmol/g FW) were calculated according to [The lipid peroxidation in maize seedlings was determined by estimating the MDA content as a TBARS concentration, described by Hodges et al.  with slirding to .p > 0.05). The statistical differences between the control and other variants were analyzed using a paired t-test (p < 0.05).The data were analyzed using Dell Statistica , version 13 . The normal distribution was evaluated using the Shapiro–Wilk test (