Since the outbreak of COVID-19 pandemic, clinical data from various parts of the world have been reported. Up till now, there has been no clinical data with regards to COVID-19 from Bosnia and Herzegovina (B&H). The aim was to report on the first cohort of patients from B&H and to analyze factors that influence COVID-19 patient’s length of hospitalization (LOH).2 saturation upon admission.This retrospective cohort study was conducted at Tuzla University Clinical Center (UKC), B&H. It involved 25 COVID-19 positive patients that needed hospitalisation between March 28th and April 27th 2020. The LOH was measured from the time of admission to discharge. Factors analyzed induced age, BMI, presence of known comorbidities, serum creatinine and O. LOH for patients with BMI < 25 was 9 ± SE2.646 days (CI 95% 3.814–14.816) vs 14.182 ± SE .937  for ≥25 BMI. The mean LOH of patients with normal levels of O2 ≥ 95% was 11.667 ± SE1.202 , while LOH for patients with < 95% was 14.625 ± SE 1.231 CI95% 12.184 to 16.757 p = 0.042; HR 3.732 CI95%1.137–12.251 p = 0.03). Patients without known comorbidities had a mean LOH of 11.700 ± SE1.075 (CI 95% 9.592–13.808), while those with comorbidities had a mean of 14.8 ± 1.303  with HR2.552.The mean age was 52.92 ± 19.15 years and BMI 28.80 ± 4.22LOH varied among COVID-19 patients and was prolonged when analyzed for BMI ≥25, comorbidities, elevated creatinine, and O2 saturation < 95%. Furthermore, risk factors for COVID-19 patients in B&H do not deviate from those reported in other countries. In December 2019, a novel β-coronavirus emerged in the Wuhan, China, causing pneumonia-like illness. Later, this virus was identified as the severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV2), which caused a widespread flu-like respiratory disease, and was named the Coronavirus Disease 2019 - COVID-19 .The reported mortality varied from country to country, from 7.2% in Italy to 2.3% that has been reported in China . Until A2 exchange in the lungs [SARS-CoV2 is an RNA virus, from the family of the coronaviruses, that were first identified in the 1960s, and since then, seven of the coronaviruses are known to infect humans . Conjunche lungs , 12.Association of obesity (BMI > 30) and severe clinical presentation in COVID-19 infected patients is currently being investigated. The mechanism on how obesity correlates to more severe outcome could be explained by the substantial respiratory system compromise paired with increased airway resistance, impaired gas exchange, a lower lung volume and weaker respiratory muscle strength. Furthermore, obese patients are more likely to have other comorbidities such as cardiovascular disease, insulin resistance or metabolic imbalances, that put these patients at risk severe form of COVID-19 infection. The risk of mortality in obese patients was recognized before COVID-19 in other viral infections, where in the previous influenza pandemics of H1N1 and H1N5, patients with a higher BMI had higher risk of lethal outcome , 14. In According to the CDC, older age groups as well as any age group with known comorbidities were found to have a higher risk of developing severe from a SARS-CoV2 infection. Comorbidities that were reported to be associated with severe form of COVID-19 were moderate to severe asthma, chronic lung conditions, diabetes, serious heart disease, kidney disease undergoing dialysis, chronic liver disease, cancer or immunocompromised individuals .This report aims to analyze the first 25 patients who tested positive for COVID-19 in Northeaster part of B&H and whose clinical condition required hospitalization at Tuzla University Clinical Center, Bosnia and Herzegovina and the impact of different clinical characteristics and admission factors on the length of hospitalization (LOH). These patients were followed from admission to discharge. Further exploration of their clinical characteristics will be elaborated as a full understanding of the disease is still limited.2 saturation > 94%, no O2 support, as well as two consecutive COVID-19 RT-qPCR negative tests with at least 48 h in-between. Twenty-five patients were admitted to the hospital due to their clinical severity (n = 25). Data was collected from patients medical history and discharge reports, as well as from blood laboratory results that were collected upon hospitalization. Data included length of hospitalization, the eventual outcome of care, age, O2% saturation, Body Mass Index (BMI), creatinine, as well as any presence of known comorbidities. Ethical approval was granted from Ethical Committee of Tuzla University Clinical Center, Tuzla.Patients studied in this retrospective cohort study were admitted to the Infections Diseases Clinic, Tuzla University Clinical Center, Bosnia and Herzegovina, from 28th of March 2020 until the 27th of April 2020, when the last patient in this study was discharged form hospital. The diagnosis of COVID-19 infection was confirmed by oropharyngeal swab using reverse transcription polymerase chain reaction (RT-PCR) diagnostic test. Admission criteria was based on Modified Early Warning Score greater than 4 together with overall clinical presentation; while criteria for discharge were: afebrile for three consecutive days, improvement of respiratory symptoms, Op < 0.05 was reported as significant. Hazard ratio was calculated by Cox linear regression. The statistical analysis and graph creation were conducted using SPSS v.25 .Data is represented as mean ± standard deviation if not indicated otherwise. Kaplan-Meier survival analysis was used for analyzing the LOH, and the difference between two groups was analyzed using the log-rank; any n = 15) and 40% female (n = 10) The mortality rate in this study was 0%. In respect to presence of known comorbidities, 60% had one or more, while 40% had no none  needing hospitalization. Out of the 25 patients, 60% were male .p = 0.040) LOH .When the mean LOH was compared to BMI the mean duration of hospitalization for patients with normal BMI was 9 ± SE 2.646 (CI 95% 3.814–14.816) while patients with BMI ≥25 had 14.182 ± SE0.937 . On the 8th day of hospitalization 80% of patients with known comorbidities were still in hospital whereas 85% in the group without , while those with comorbidities had a mean of 14.8 ± 1.303 , while LOH for patients with ≤94% was 14.625 ± SE 1.231 CI95% 12.184 to 16.757 p = 0.042 with HR 3.732 CI95%1.137 to 12.251 p = 0.03  when compared to group of patients that had a normal creatinine level upon admission 12.727 ± SE0.846   with HR 5.148. Even though our study had no mortality, it is in line with previous studies on BMI impact on disease severity and can be used as a morbidity predictor [The effect BMI had on the LOH was significant, where patients who were categorized as overweight or obese had a longer LOH when compared to patients with a mean of 9 days for patients with BMI < 25 vs 14.18 days with BMI > 25  with HR of 2.552 but with borderline statistical significance. This could the consequence of smaller size of the study group and limited number of patients that were infected in the first wave of COVID-19 in the region.In other studies, COVID-19 patients who had comorbidities on admission to hospital or critical care had higher mortality rates, as well as severe clinical manifestation. Individuals who have comorbidities are more inclined have health with reduced capacity to withstand COVID-19 infection . When pap < 0.05) [Moreover, creatinine levels show the overall function of the kidney-urinary system; elevated levels could point to decrease in function or kidney injury. COVID-19 patients with an acute kidney injury in COVID-19 patients showed higher levels of hospital deaths than those with normal levels. When LOH was compared among the two groups of patients – those who had a normal level of creatinine to those with elevated – patients with elevated showed longer LOH with 19.67 ± 3.18 (CI95% 13.434–25.899) days, higher than those with normal levels 12.727 ± 0.85; CI95% 11.07–14.385;  < 0.05) .2 saturation levels with HR 3.732 and finally presence of known comorbidities with HR of 2.552. Despite the absence of mortality in our sample, these factors provided guidance in future planning of hospitalization of COVID-19 patients at UKC Tuzla. This is the first study regarding COVID-19 pandemic in B&H that has been written about clinical characteristics and biochemical results from COVID-19 patients. Results in this study could add to the current information about the COVID-19 pandemic in Europe. Furthermore, it provides additional information on risk factors attributed to longer hospitalization. However, this study involved a fraction of COVID-19 patients and included only a part of the B&H experience. Also, a larger sample of patients with COVID-19 is needed to make further conclusions on the nature of the pandemic in B&H. Moreover, not all biochemical and inflammatory markers that are routinely requested to monitor patients with COVID-19 were analyzed, such as d-dimer; hence, they were not included in this study. Elevated creatinine levels showed statistical difference on log-rank analyses but has not provided significant result in respect to HR, which could be verified in larger studies.Possibility of predicting LOH of COVID-19 could be one of the crucial aspects of this pandemic, by providing additional tolls in planning hospital capacity and allocation of resources. Factors that were associated with a longer LOH was BMI ≥25 with HR 5.148, followed by O


Microalgae can contribute to more than 40% of global primary biomass production and are suitable candidates for various biotechnology applications such as food, feed products, drugs, fuels, and wastewater treatment. However, the primary limitation for large-scale algae production is the fact that algae requires large amounts of fresh water for cultivation. To address this issue, scientists around the world are working on ways to reuse the water to grow microalgae so that it can be grown in successive cycles without the need for fresh water.Euglena gracilis. Firstly, the results show that E. gracilis can be only cultivated through two growth cycles with water that has been filtered and reused, and the growth of E. gracilis is significantly inhibited when the water is used a third time. Secondly, as the number of reused water cycles increases, the Cl− concentration gradually increases in the cultivation water. When the Cl− concentration accumulates to a level of fivefold higher than that of the control, growth of E. gracilis is inhibited as the osmolality tolerance range is exceeded. Interestingly, the osmolality of the reused water can be reduced by replacing NH4Cl with urea as the source of nitrogen in the cultivation water. Thirdly, E. gracilis secretes humic acid (HA)—which is produced by the metabolic pathways for valine, leucine, and isoleucine biosynthesis and by linoleic acid metabolism—into the cultivation water. Because HA contains large fluorescent functional groups, specifically extended π(pi)-systems containing C=C and C=O groups and aromatic rings, we were able to observe a positive correlation between HA concentration and the rate of inhibition of E. gracilis growth using fluorescence spectroscopy. Moreover, photosynthetic efficiency is adversely interfered by HA, thereby reductions in the synthetic efficiency of paramylon and lipid in E. gracilis. In this way, we are able to confirm that HA is the main growth inhibitor of E. gracilis. Finally, we verify that all the HA is removed or converted into nutrients efficiently by AC or UV/H2O2/O3 treatments, respectively. As a result of these treatments, growth of E. gracilis is restored (AC treatment) and the amount of biomass is promoted (UV/H2O2/O3 treatment).In this study, we present the results when we cultivate microalgae with cultivation water that is purified and reused. Specifically, we purify the cultivation water using an ultrafiltration membrane (UFM) treatment and investigate how this treatment affects: the biomass and biochemical components of the microalgae; characteristics of microalgae growth inhibitors; the mechanism whereby potential growth inhibitors are secreted ; the effect of activated carbon (AC) treatment and advanced oxidation processes (AOPs) on the removal of growth inhibitors of E. gracilis and for saving water resources. Our work may also provide a useful reference for other microalgae cultivation.These studies have important practical and theoretical significance for the cyclic cultivation of The online version contains supplementary material available at 10.1186/s13068-021-01980-4. Unicellular eukaryotic microalgae are a diverse and ubiquitous group of plants that are a promising biomass source and a biological feedstock , 2. HoweEuglena spp. algae, especially, there have been no reports of E. gracilis cultivation with reused water. E. gracilis is a unicellular flagellated alga characterized by the absence of a cell wall. It produces a wide variety of bioactive compounds such as paramylon, carotenoids, tocopherols, euglenophycin, and lipids. It has tremendous potential for metabolic engineering and commercialization [E. gracilis, and the mechanisms underlying its growth-inhibiting secretions, are urgently needed.The effects of water reuse on algae growth are different across algae taxa . The moslization . Therefo−, Na+, and K+ from NH4Cl, NaHCO3, and KH2PO4, respectively, can accumulate in the cultivation water. When these ions accumulate, the osmotic pressure of the cultivation water increases, thereby inhibiting microalgae growth [After the microalgae assimilate nutrients ions, unabsorbed counter ions such as Cle growth , 9. TherScenedesmus sp. LX1, DOM concentrations between 6.4 and 25.8 mg/L in reused water resulted in a decrease in the maximum algae cells density and the maximum growth rate by 50–80% and 35–70%, respectively [Scenedesmus acuminatus in the reused water included palmitic acid and octadecanoic acid, both of which inhibited the growth of this algae species. Although many studies have attempted to characterize the growth inhibitor present in DOM, it is unclear which major metabolic pathways within microalgae cells regulate and secrete these inhibitory substances into the cultivation water.In addition to accumulated ions, excreted metabolites, such as dissolved organic matter (DOM) from microalgae is considered to be the main cause of negative biomass growth , 6, 10. ectively . The DOMNannochloropsis oceanica moderately reduced growth inhibition and lipid accumulation. Moreover, the AC treatment significantly increased the final dry weight of S. acuminatus to 2.33 ± 0.04 g/L, which was almost the same as the dry weight obtained after growth in fresh media [3, UV/H2O2 have been successfully applied in the treatment reused water for the cultivation of Scenedesmus sp. LX1 [S. acuminatus GT-2 [E. gracilis.Recently, many researchers have tried to use traditional methods of wastewater treatment on microalgae cultivation water. Zhang et al.  reportedsh media . Advancesh media , 15. O3, sp. LX1  and S. atus GT-2 , respectE. gracilis; (2) to identify the characteristics of the growth inhibitors in reused water and uncover the mechanism whereby potential inhibitors are secreted by E. gracilis via metabolomics analysis; (3) and to evaluate the effect of AC and AOPs on the removal of growth inhibitors.In this study, the main objectives were: (1) to identify the effect of treatment with ultrafiltration membrane (UFM) on the biomass and biochemical components of cultivated p < 0.05), 28.6% , and 79.2%  compared to the control group on the last day of cultivation  compared to the control, the relative cell density was lower, algal cytochromes were almost absent, and the algae cells were elongated . However, when the concentrations of the medium were increased to above fivefold that of the control medium, the DW gradually decreased. In fact, when the concentration of the medium was increased by eight-, nine-, and tenfold, the DW of cells decreased by 81.3% (p < 0.01), 85.0% (p < 0.01), and 92.5% (p < 0.01), respectively  and found that as the concentration of urea increased, the DW of algae cells increased significantly. When the concentration of the culture medium was four to tenfold, the biomass of algal cells was stably maintained at about 2.7 g/L, and there was no significant difference among them (p > 0.05) , and the osmolality for NH4Cl was 4.3-fold greater than urea . These results show that the growth of algae cells was not inhibited with a fivefold increase in salinity (< 15.6 psu) and osmolality (< 361.1 mosm) of the medium, and we can confirm that during the UFM-R3 culture cycle, the growth of E. gracilis was not hindered by the accumulated ions in the reused water. This phenomenon has also been confirmed by the cultivation of S. acuminatus in reused water [E. gracilis had a certain tolerance range to ions. If this tolerance range was exceeded, the growth of E. gracilis was inhibited. In addition, it was determined that the traditional medium PEM with NH4Cl as the nitrogen source was not suitable for the continuous recycling of cultivation water or for batch-fed cultivation of E. gracilis (such as heterotrophic batch-fed fermentation). When urea is used, it serves as an ideal nitrogen source because it reduces the salinity and osmolality in the culture medium.To prove the above hypothesis, we used urea instead of NHed water , 19. OurE. gracilis continuously secreted DOM during the culture process. By the time UFM-R3 was reached, the DOM concentration was 189.21 mg/L, while the control group contained only 54.92 mg/L DOM, a 3.4-fold difference (p < 0.01)  for organic matter represents the content of organic functional groups that contribute to fluorescence, such as C=C bonds, C=O bonds, and aromatic rings. The importance of ultraviolet spectra for detecting pollutants in the water treatment process was described by Altmann et al. [254 of 80-fold-concentrated DOM and found that the fluorescence intensity from high to low was: HoN (0.58 AU/cm), HoA (0.53 AU/cm), HiA (0.50 AU/cm), HiB (0.44 AU/cm), HoB (0.30 AU/cm), and HiN (0.24 AU/cm) . From this result, it is obvious that HA significantly reduces the algae cell’s photosynthetic efficiency. Similarly, studies on S. acuminatus [Arthrospira platensis [The Fv/Fm ratio reflects the ability of microalgae to dissipate, absorb, and transmit light energy during photosynthesis. It is a useful parameter that indicates physiological state and growth rate, and is also an internal probe of the relationship between microalgae and their environment , 23, 24.uminatus  and ArthE. gracilis in the experimental group containing DOM were 7.1% and 12.2%, respectively, while the control group was 21.2% and 35.2%. Both of these values were significantly lower than the control group, which showed a decrease of 66.5% , and 65.3% , respectively. These results were confirmed for the TFA of Scenedesmus sp. LX1 [E. gracilis may interfere with its own photosynthesis, and that this leads to inhibition of the synthesis of organic matter in the algae cells. The mechanism behind this process is worthy of our in-depth study in the future.The paramylon content and TFA content of  sp. LX1 . These rE. gracilis cells and cultivation media, the range of metabolites detected in negative ion mode was greater than that in positive ion mode, so this study only analyzes metabolites that were observed in negative ion mode to describe the mechanism whereby algae cells secrete DOM. With this analysis, we observed 4130 metabolites  interfere with the electron transport processes of photosynthesis. How these functional groups in the compounds secreted by different metabolic pathways interfere with the photosynthetic system of microalgae requires further in-depth study.HA containing multiple functional groups can complex iron ions that are essential for photosynthesis in microalgae. However, Sun et al.  found thonfirmed . In addip > 0.05). This indicates that AC is effective at completely adsorbing and removing substances that inhibit the growth of E. gracilis. Although the reused water of cultivated N. oceanica [and S. acuminatus [E. gracilis in this study. We would like to develop recyclable AC technology, such as biological AC, to increase the utilization rate so that it can be more convenient for large-scale reuse of water resources to cultivate E. gracilis.Markiewicz et al.  have conoceanica  and S. auminatus  showed a2O2/O3 group was weaker . These results indicate that the advanced oxidation method not only eliminates the growth inhibitors, but may also oxidize these inhibitors into small organic molecules that could be absorbed by algae cells, thereby increasing their biomass. Our results show that the growth inhibitors were mainly HAs with luminescent functional groups . O3 and UV/H2O2 have been shown to work well for the treatment of Scenedesmus sp. LX1 [S. acuminatus GT-2 [3 or UV/H2O2 alone. Therefore, we believe that UV/H2O2/O3 is an ideal and efficient method for the removal of inhibitors of E. gracilis.AOPs have been widely used in the field of wastewater treatment. Oxidizers create a large number of free radicals under ultraviolet catalysis, such as hydroxyl radicals. These free radicals have strong oxidizing properties and can oxidize organic acids with unsaturated bonds . Accordiker Fig. b, follow2O2 Fig. c, indica sp. LX1  and S. atus GT-2  reused wE. gracilis is a main growth inhibitor.According to our previous research, the free radicals in the reused water after treatment with AOPs could also inhibit the growth of microalgae. Therefore, we need to optimize the AOP treatment process in the future by optimizing the treatment time, the concentration of the oxidizing agent, and the development of indicators for online detection of the concentration of free radicals in reused water . Use of AOPs is more conducive to the wide application of water reuse for algae cultivation. In addition, through these treatments, again it is clear that HA secreted by E. gracilis  and even enhanced E. gracilis growth (UV/H2O2/O3 treatment). This result provides further confirmation that HA was a main growth inhibitor. An effective model for the cyclic culture of E. gracilis was thus proposed. These studies have important practical and theoretical significance for cyclic cultivation of E. gracilis or even other species of microalgae and for saving precious water resources.Our study demonstrated that cultivation water used three times had a significant inhibitory effect on the growth of Euglena gracilis CCAP 1224/5Z was obtained from the Culture Collection of Algae and Protozoa (CCAP) and maintained in our lab at Shenzhen University [Euglena medium (PEM) according to Cramer and Myers [4Cl, 0.6 g/L KH2PO4, 1.2 g/L MgSO4·7H2O, 0.02 g/L CaCl2·2H2O, 0.55 μg/L Na2EDTA·2H2O, 2.0 μg/L Fe2(SO4)3, 0.05 μg/L CuSO4·5H2O, 0.4 μg/L ZnSO4·7H2O, 1.3 μg/L Co(NO3)2·6H2O, 1.8 μg/L MnCl2·4H2O, 0.01 μg/L Vitamin B1, and 0.0005 μg/L Vitamin B12. The pH value of the PEM medium was 3.6 adjusted with 3 mol/L NaOH and 1 mol/L HCl. The microalgae cells were grown in 2-L glass column photobioreactors with a 10-cm internal column diameter. The photobioreactors contained 1.5 L PEM medium that was stirred with 0.2 μm-filtered mixed gas  and illuminated with an LED lamp at a light intensity of 150 μmol photons m−2 s−1 (24: 0 h light–dark cycle). The temperature of the cultivation was maintained at 25.0 °C.iversity . This stnd Myers . PEM inc750 concentration of 0.2 for the microalgae suspension. After the first cultivation, water samples were cycled through this cultivation and UFM cycle three times and these subsequent cycles were named UFM-R1, R2, R3. Samples of the microalgae were taken every other day to monitor the cells’ dry weight (DW).When the algae cells were cultured on day 10, the cultivation water sample (UFM-R0) was treated with ultrafiltration membrane (UFM) harvesting equipment  that was4HCO3. The filters were weighed after drying at 105 °C for 24 h to reach a constant mass. DW was calculated using Eq.  as the nitrogen source. When the algae cells were cultured on day 10, the morphology of the algae cells was observed with an inverted microscope . In addition, the DW, Cl− concentration, salinity, osmolality, and the pigment changes of the algae cells in the culture medium were measured. Cl− concentration was measured using a capillary ion chromatograph  [T(°C), according to the following Eq.  content between the control (UFM-R0) and the reused water samples  were measured using a total organic carbon analyzer .https://www.originlab.com/origin).DOM was determined with three-dimensional fluorescence excitation–emission matrix (3D-FEEM) spectrophotometry. Briefly, 3D-FEEM spectra were obtained using a fluorescence spectrophotometer . The excitation (Ex) and emission (Em) slits were set to a bandpass of 5 nm. Ex wavelengths were scanned from 200 to 450 nm, and Em wavelengths were scanned from 220 to 550 nm. All of the 3D-FEEM spectral data were analyzed with Origin Pro 2018 software (254) of DOM in each fraction was measured by using a UV–Vis spectroscopic measurements .The method of pretreated water sample was based on Leenheer . The frac” is the DW of the control group and “i” is the DW of algae cells under the different fractionated DOM stressed conditions. Finally, the correlation analysis of UV254 and IG% were performed by using Origin Pro 2018 software.The fractionated organic acids were added to fresh medium according to the percentage of DOM. After culturing the algae cells with this medium for 10 days, the DWs of the control and the experimental groups were measured. The inhibition rate of growth (IG%) was calculated using the following Eq. :3\documeAfter the fractionated DOM was diluted according to the DOM content in the UFM-R3 reused water, as described above, fresh medium was added, and then the algae cells were cultivated under the afore mentioned culture conditions. The Fv/Fm ratio, the paramylon and the total fatty acid (TFA) content of the algae cells were measured after the 10th day of cultivation.The Fv/Fm ratio was determined by dividing the variable fluorescence (Fv) by the maximum fluorescence (Fm), according to the method of Sha et al. . The algg for 5 min, the supernatant was removed. 1.5 mL of 1% sodium dodecyl sulfate (SDS) solution was added to the tube, then the contents were transferred into a 1.5-mL centrifuge tube and heated in a water bath at 85 °C for 2 h. Again, the supernatant was removed after the centrifuge tube was centrifuged at 2000×g for 5 min. The precipitate was washed and centrifuged in 1 mL deionized water, then oven-dried at 70 °C to a constant mass. The resulting precipitate was paramylon. The paramylon content was calculated as shown in Eq. . Furthermore, the value of variable importance in the projection (VIP) of the first principal component in OPLS-DA analysis was obtained. It summarizes the contribution of each variable to the model. The metabolites with VIP > 1 and p < 0.05 (Student’s t-test) were considered to be significantly changed. In addition, commercial databases including KEGG database (http://www.genome.jp/kegg/) and MetaboAnalyst  were used for metabolic pathway enrichment analysis. From these analyses, bubble diagrams and metabolic pathways were made.u et al. . All of 2O2/O3 and UV/H2O2) were similar to the result of Hu et al. [3 with a flow rate of 3000 mg/h, was fed into a solution containing 189.2 mg/L DOM (found in reused water UFM-R3) and 1% H2O2 for 2 h under UV254 ultraviolet lamp irradiation. This is the UV/H2O2/O3 experimental group. In the other set of experiments, O3 was not used, but all other conditions were the same. This is the UV/H2O2 experimental group. After all the treated water samples were freeze-dried, fresh medium was added to each. The method of AC filtration was similar to that of Sha et al. [The methods of advanced oxidation that we used  were considered significantly different, p < 0.05 values were considered statistically different, and p > 0.05 values were considered not statistically different (NS) compared to the control groups.All DWs, ClAdditional file 1: Fig. S1. Microalgae harvesting equipment. a, microalgae suspension; b, ultrafiltration membrane; c, pre-concentrated microalgae; d, reused water. Fig. S2. PCA analysis for groups EEG and IEG. IEG, EEG represent intracellular and extracellular metabolites, respectively. Fig. S3. OPLS-DA analysis for group EEG and IEG. IEG, EEG represent intracellular and extracellular metabolites, respectively. Fig. S4. Permutation test for group EEG and IEG. IEG, EEG represent intracellular and extracellular metabolites, respectively. Fig. S5. Heatmap of hierarchical clustering analysis for group EEG and IEG. IEG, EEG represent intracellular and extracellular metabolites, respectively; * represents the metabolite can be annotated in KEGG database.Additional file 2. The data of statistical analysis results, different metabolites, and pathway analysis from the group EEG and IEG.


Setaria italica) as the main traditional crop in China, is rich in many kinds of high quality fatty acids (FAs). In this study, Ultra-high performance liquid chromatography-time-of-flight-tandem mass spectrometer (UHPLC-Q-TOF-MS/MS) was used to determine the lipids of JG35 and JG39. A total of 2,633 lipid molecules and 31 lipid subclasses were identified, mainly including thirteen kinds of glycerophospholipids (GP), eleven kinds of glycerolipids (GL), four kinds of sphingolipids (SP), two kinds of fatty acyls (FA) and one kind of sterol (ST). Among them JG35 had higher contents of diacylglycerols (DG) and ceramides (Cer), while triacylglycerols, phosphatidyl ethanolamine, phosphatidic acid, sterol, fatty acyls and pardiolipin  were higher in JG39. Meantime, the correlation analysis of lipidomics and transcriptomics was used to map the main differential lipid metabolism pathways of foxtail millet. The results shown that a differentially expressed genes (DEGs) of FATA/B for the synthesis of FA was highly expressed in JG35, and the related genes for the synthesis DG , TG (DGAT and PDAT) and CL (CLS) were highly expressed in JG39. The results of this study will provide a theoretical basis for the future study of lipidomics, improvement of lipid quality directionally and breeding of idiosyncratic quality varieties in foxtail millet.Foxtail millet ( Setaria italica) is one of the most ancient grain crops originating from China and is among the top five most important cereals worldwide , glycerolipids (GL), glycerophospholipids (GP), sphingolipids (SP), saccharolipids (SL), sterol lipids (ST), prenol lipids (PR) and polyketides (PK) . LipidomSince its development, lipidomics has been widely applied in biomedicine, human health, and other aspects . Lipids Zanthoxylum bungeanum seeds, the results of intergroup correlation and RDA analysis suggested that ENR, ECR, and SAD1 are the key genes in fatty acid synthesis. Paeonia lactiflora seeds, including MCAT, KASIII, FATA, SAD, FAD2, FAD3, and DGAT, providing more comprehensive genomic resources for understanding the transcription of genes in P. lactiflora seeds.With the completion of foxtail millet genome sequencing and the development of bioinformatics, it has provided convenient conditions for digging into the excellent functional genes of foxtail millet . Transcrper se , seeds were selected 28 days after pollination with three replicates. And the transcriptome data of seeds were obtained from the NCBI Sequence Read Archive (SRA) under accessions with the SRR number 11840494, 11840495, 11840514, 11840501, 11840502, and 11840503, which were reported by Representative whole seeds without mildew, particles, or other impurities were selected and unshelled into foxtail millet using a JLGL-45 hulling mill . Next, the seeds were ground into powder using liquid nitrogen and mixed evenly for subsequent analysis. Precisely, 50 mg sample was weighed and put into an EP tube containing 200 μL water. The sample was then blended through whirl pooling for 30 s, then ground using friction bails for 12 min at 1,000 rpm. Next, the sample was subjected to 15 min of ultrasonic processing in an ice-water bath, then 480 μL extract (MTBE: methanol = 5:1) was added, followed by 30 s of blending through whirlpooling and 10 min of ultrasonic processing in an ice-water bath. After that, the sample was left to stand for 1 h at −20°C and centrifuged for 15 min at 4°C, 10,000 rpm. A total of 380 μL supernatant was carefully extracted and added into an EP tube. The extract was dried in a vacuum concentrator then mixed with 200 μL solution (dichloromethane: methanol = 1:1) for redissolution. Next, the mixture was blended through whirlpooling for 30 s and subjected to ultrasonic processing in an ice-water bath for 10 min. The sample was then centrifuged for 15 min at 4°C, 13,000 rpm, and 180 μL supernatant was carefully extracted and added into a vial. Exactly 10 μL of each sample was taken and mixed into the QC sample for testing on the machine. All reagents used were chromatographically pure.Chromatographic separation was performed on a Waters UPLC Acquity I-Class PLUS ultra-high performance liquid chromatography system. The chromatography column used was the Acquity UPLC CSH C18  column purchased from Waters. Positive ion mode: mobile phase A: 60% acetonitrile water solution, 10 mM ammonium acetate, 0.1% formic acid; mobile phase B: 90% isopropanol/acetonitrile solution, 10 mM ammonium acetate, 0.1% formic acid. Negative ion mode: mobile phase A: 60% acetonitrile water solution, 10 mM ammonium acetate, 0.1% formic acid; mobile phase B: 90% isopropanol/acetonitrile solution, 10 mM ammonium acetate, 0.1% formic acid. The gradient elution was programmed as follows: 0–2.0 min, 40% B; 2–18 min, 43%–100% B; 18–20 min, 40% B. Flow rate: 0.4 ml/min; injection volume: 5 μL; column temperature: 55°C. The sample was placed in a 10°C automatic sampler for analysis.Primary and secondary mass spectrometric data were collected under software  controlled MSe mode using a Waters Xevo G2-XS QTOF high-resolution mass spectrometer. Data on the low and high collision energy were collected by two channels in each data collection cycle. The low collision energy was 2 V, whereas the high collision energy interval was 10–40 V. The scanning frequency was 0.2 s/mass spectrum. ESI ion source parameters: capillary voltage: 2,000 V (positive ion mode) or −1,500 V (negative ion mode); taper hole voltage: 30 V; ion source temperature: 120°C; desolvent gas temperature: 550°C; blowback air flow rate: 50 L/h; desolvent gas flow rate: 900 L/h.The original data collected by MassLynx V4.2 were put through peak recognition, peak extraction, peak alignment, and other data processing operations using Progenesis QI software. Identification was performed using Progenesis QI software’s online LMSD (Lipid Maps Structure Database) database and BMK’s self-built database, and theoretical fragment recognition was conducted. The Precursor Ion mass number deviation was within 100 ppm in each case, the fragment ion mass number deviation was within 50 ppm in each case . The extp values were adjusted using the Benjamini and Hochberg’s approach for controlling the false discovery rate. Genes with an adjusted p-value < 0.01 and | log2FC | > 1 found by DESeq2 were assigned as differentially expressed. Gene Ontology (GO) enrichment analysis of the differentially expressed genes (DEGs) was implemented by the GOseq R packages based Wallenius non-central hyper-geometric distribution  were subjected to differential analysis based on univariate analysis, where the red denotes the differential metabolites with | log2FC | > 1, and adjusted p-value < 0.05. The volcano plot of differential expression was shown in The univariate statistical methods commonly used for differential analysis of two sample groups include variation fold analysis and the 2X, R2Y, and Q2. R2X and R2Y denote the explanatory powers of the model built for X and Y matrices, respectively, and Q2 denotes the prediction rate of the model. Generally, the closer the values of R2Y and Q2 to 1, the higher the stability and reliability of the model. Under normal circumstances, the model is considered effective when Q2 > 0.5, and excellent when Q2 > 0.9. According to our results, the parameters R2Y and Q2 of the model were 0.998 and 0.832, respectively  was performed to determine the lipid micromolecules with significant changes in the JG35 and JG39 millet varieties, thus building the relational model between the expression levels of lipids and the sample. The evaluation parameters of the model mainly included Rectively , indicatly <0.05 , indicatComplete lipidomic analysis was performed on grain samples from two foxtail millet varieties using UHPLC-QTOF-MS. According to the results, 31 lipid subclasses  were identified, including 13 kinds of GP, 11 kinds of GL, 4 kinds of SP, 2 kinds of FA and 1 kind of ST . Among t8, whereas the content was 1.27×108 in JG39. Overall, the relative contents of various lipid compositions in JG35 and JG39 were similar, and GL content was the highest, accounting for 48.24 and 47.55% of the total lipid content, respectively  accounted for 13.10 and 12.31%, respectively. FA (including FA and WE) accounted for 5.46 and 5.42%, respectively. The contents of ST in JG35 and JG39 were 5.77 × 106 and 6.01 × 106, respectively. In conclusion, it was found that JG35 had higher DG and Cer, while JG39 had higher TG, PE, PA, ST, FA, and CL.Among the two foxtail millet varieties, JG35 had a relative lipid content of 1.21×10ectively . Specifi2FC | >1 and adjusted p-value < 0.05) were selected from the 2,633 lipid molecules identified through qualification  and FAs as initial substrates. Denovo synthesis occurs in both endoplasmic reticulum (ER) and plasmid pathways. Acetyl coenzyme A serves as the primer in the synthesis of FAs. malonyl-ACP provides a dicarbon unit for each step of the extension reaction. Malonyl thioester first engages in condensation reactions with acetyl-CoA and then goes through acceptor reactions with acyl-ACP. These reactions are catalyzed by KAS condensing enzymes, resulting in the formation of carbon-carbon bonds. It takes three different KAS condensing enzymes to generate one 18C fatty acid, containing KAS І, KAS ІІ, and KAS ІІІ. In the condensation reaction process, the extension of the fatty acyl group chain further requires the participation of KAR, ENR, and HAD. The long-chain acyl groups are hydrolyzed by FATB, generating 18:0-ACP. 18:0-ACP is further hydroxylated under the action of FATA/B. Finally, FAs are activated into acyl coenzyme A via LACS, which is exported into the ER.PA and DG are both important intermediate products in the metabolic process of lipids. Their FATA/B were also higher in JG35. For DG, they were exhibited a relatively high content in JG39, and the expression level of ACCase, KAS, KCS, LACS and GPAT corresponding to DG synthesis were also high in the transcriptomes. Similarly, JG39 had higher TG content, and the corresponding synthase genes DGAT and PDAT expression were upregulated. Besides, CL was higher in JG39 and the expression of CLS was also higher  by the catalysis of CDP-DAG synthase (CDP-DAGS) and then produce CL by the catalysis of cardiolipin synthase (CLS)  . Here, JChina has abundant foxtail millet variety resources; however, there is limited research on the effective utilization and mining of different resources. Also, many excellent resources remain unexplored, limiting the breeding of special foxtail millet varieties. The study of investigating the composition and change in foxtail millet by lipidomics is of great significance for the development of functional foods, the utilization of germplasm resources and the breeding of special varieties of foxtail millet.FATA/B for the synthesis of FAs was highly expressed in JG35, and the related genes  for the synthesis of DG were highly expressed in JG39. Besides, in JG39 the expression of DGAT and PDAT for the synthesis of TG was also higher, and CLS related to CL synthesis was upregulated. The two varieties contain different contents of lipids which are beneficial to human health, so the development of functional foods and the breeding of new varieties can be carried out selectively according to different needs.In this paper, UHPLC-Q-TOF-MS system was firstly used to compare the lipid content and composition of two different foxtail millet varieties containing JG35 and JG39, and combined with transcriptome data to construct the molecular regulatory network of major lipid metabolism preliminatively in foxtail millet. A total of 2,633 kinds of lipid molecules and 31 kinds of lipid subclasses were identified by lipid detection. By comparing, the lipid composition was not significantly different. However, there were differences in the contents of each subcategory, JG35 had higher contents of DG and Cer, and the contents of TG, PE, PA, ST, FA, and CL were higher in JG39. By the comparison of differential lipid, it was found that the key genes


Tagetes lucida Cav. (Asteraceae) have antioxidant and antimicrobial properties.The roots of T. lucida roots ethanol extract (TLRE) using carbon tetrachloride (CCl4)-induced hepatotoxicity in rats.This study aimed to examine the hepatoprotective effects of 4 (a single dose of 2 mL/kg) before TLRE or silymarin administration. Meanwhile, the protective group was implemented by administering CCl4 (a single dose of 2 mL/kg) after force-feeding TLRE or silymarin. Each therapeutic and protective group was divided into three subgroups: force-fed with saline, TLRE (500 mg/kg), and silymarin (25 mg/kg). The positive group was split into two subgroups that were force-fed TLRE and silymarin. Positive, therapeutic, and protective groups were compared to the negative group (untreated rats). CCl4, TLRE, and silymarin were orally administrated using a gastric tube.The active ingredients of TLRE were identified by high-performance liquid chromatography, infra-red spectrum, and mass spectrometric procedures. Ninety rats were distributed into four main groups: positive, therapeutic, protective, and negative group. The therapeutic group was implemented using CCl4-induced hepatotoxicity controls.In the therapeutic and protective groups, TLRE significantly reduced liver enzymes, i.e., aspartate aminotransferase (12.47 and 6.29%), alanine aminotransferase (30.48 and 11.39%), alkaline phosphatase (17.28 and 15.90%), and cytochrome P450-2E1 (39.04 and 48.24%), and tumour necrosis factor-α (53.72 and 53.72%) in comparison with CClTLRE has a potent hepatoprotective effect with a good safety margin. After a repeated study on another type of small experimental animal, their offspring, and an experiment with a large animal, this study may lead to clinical trials. These toxicants induce lipid peroxidation and oxidative stress. The industrial progress that facilitates the modern lifestyle has poisoned the environment through poor control of industrial waste disposal. One toxicant in such waste is CCl4, emitted from chemical industries and accumulates in the atmosphere. The CCl4-induced hepatotoxicity model is used to study the hepatoprotective effects of drugs and plant extracts. CCl4-induced hepatotoxicity is caused by a) the promotion of lipid peroxidation, b) inhibition of antioxidant enzymes, and c) induction of free radical production  activation on cytochrome P-450 2E1 (CYP2E1), b) generation of reactive metabolites such as trichloromethyl radicals (CCl3), and c) peroxy trichloromethyl radicals (OOCCl3). These radicals bind covalently to macromolecules such as lipids, proteins, and nucleic acids  is native to Central America. Its common names include sweet-scented marigold, Mexican mint marigold, Mexican tarragon, sweet mace, and Texas tarragon. T. lucida is new in Egypt. T. lucida plant reaches 18-30 inches in height, with a branched stem and linear to oblong leaves of shiny medium green. The plant’s small golden-yellow flowers appear in late summer at the ends of the stems. Traditionally, T. lucida has been prescribed to treat fever, tumours, diarrhoea, asthma, rheumatism, and flu. Also, it has been recommended to treat nervous complaints, including anxiety, irritability, and depression  demonstrated that the roots of T. lucida are cytotoxic to crustaceans, with low LC50 values due to the high content of coumarins in the roots. Many valuable components occur in T. lucida plants as essential oils, polyphenols, flavonoids, and coumarins  on CCl4-induced hepatotoxicity in rats.This work aimed to study the effect of T. lucida were imported from the Canadian branch of Johnny's Selected Seeds www.johnnyseeds.com (product ID: 2273) in 2017. Seeds were cultivated at the SEKM Company Farm at Bilbase, El-Sharkya Governorate, Egypt, during the winter (January) of 2018. Plants were collected in May at the flowering stage, and the roots were separated. The cultivated plant was verified by the National Research Centre herbarium (CAIRC) team. The plant was deposited in the herbarium under Voucher Specimen No. M132. Roots were exposed to heat shock at 100 °C for 1 min to stop enzyme activity that might convert phytochemicals, and they were then air-dried in the shade to completion in an oven at 40 °C. Powdered roots (1000 g) were exhaustively extracted with 70% ethanol (EtOH) by soaking at room temperature. The extract was lyophilised, and the resulting powder was kept at −20 °C until use  and was expressed as umbelliferone.The total phenolic content of TLRE was determined by the Folin-Ciocalteu method  analysis, according to Kim et al. , using aThe HPLC analysis did not identify the major component in TLRE, so TLRE was evaluated by 2-dimensional paper chromatography. The diluted TLRE (1 mL) was spotted in the corner of a 20 × 20 cm paper chromatography sheet (Whatman No. 1) about 2 cm from the edges. The separation was first developed with a nonpolar running system after air-drying , followed by a polar running system (acetonitrile: methanol: 3:1) the direction of the nonpolar run. The paper sheet was allowed to dry at room temperature and then was visualised with ultraviolet (UV) light at 354 nm; the major compound appeared as a pale bright yellow spot at the corner. After detecting the spot, it was cut out and re-extracted from the paper with 99% ethanol; after concentration, the residue was subjected to structure analysis by infra-red (IR) and mass (MS) spectrometry.n = 8) by the method of Bruce  weighing 140–180 g each were obtained from the central animal house of the National Research Centre, Dokki, Giza, Egypt. Animals were housed in plastic cages under laboratory conditions  at the animal facility of the animal house. The feed was a commercial chow-based diet obtained from the animal house. Water and food were available 4-induced hepatotoxicity model was used in this study according to Mir et al. , where each mL of CCl4 mixed with 1 mL of corn oil.CClr et al.  with sli), the TLRE dose was calculated as 1/10 of the LD50 that was 500 mg/kg/day. The dose of silymarin was the standard recommended dose, 25 mg/kg/day , which received a saline solution for the entire experimental period. This group was maintained as a negative control.th day of the experiment.The second main group was the positive group (20 rats), broken into two subgroups; each subgroup of 10 rats received either silymarin or TLRE, respectively. These were maintained as the positive control for the two materials. Rats in these subgroups were killed on the 104 at 2 mL/kg body weight as a single dose, and after two days, these rats were treated with either the TLRE or silymarin for 7 days. These rats were killed on the 10th day. The therapeutic group was classified into three subgroups (n = 10). Rats received saline only for 7 days and were maintained as the therapeutic control. Rats received the recommended dose of silymarin for 7 days, and rats received the TLRE 500 mg/kg body weight for 7 days.The third main group was the therapeutic group (30 rats), which was designed to evaluate the therapeutic effect of TLRE, according to Ibrahim et al. . The rat4 at 2 mL/kg body weight as a single dose. These rats were killed two days later, on the tenth day. This hepatoprotective group was divided into three subgroups (n = 10). First, rats received saline only for 7 days and were maintained as a hepatoprotective control. Rats received the recommended dose of silymarin for 7 days, and rats received the extract as 500 mg/kg body weight for 7 days.The fourth main group was the hepatoprotective group (30 rats) that was designed to evaluate the hepatoprotective effect of TLRE, according to Ibrahim et al. . Rats ing, 10 min, 4 °C, using a Sigma Laboratory centrifuge) to separate the serum. Livers were separated, washed in ice-cold 1.15% KCl solution, and freshly weighed. A piece of liver from each rat was homogenised in ice-cold Tris-HCl buffer, 0.1 M, pH 7.4, using an ultrasonic homogeniser. The homogenate was centrifuged at 4000 g for 15 min at 4 °C using a Sigma Laboratory centrifuge and kept at −20 °C until analysis of the antioxidants. Another piece of liver from each rat was removed, and the liver microsomes were prepared and were isolated according to a previous method . In addition, globulin was calculated as the difference between TP and albumin according to the method of Reinhold , albumin, total bilirubin, and aspartate aminotransferase (AST) and alanine aminotransferase (ALT), and alkaline phosphatase (ALP) activities were measured according to Henry , Dumas e1969Reinhold .Liver antioxidant enzyme activities were determined in the liver homogenate. Activities of catalase (CAT), superoxide dismutase (SOD), glutathione reductase (GR), and glutathione peroxidase (GPx) were determined according to the methods described by Paglia and Valentine , Beers a2O2) in liver homogenate was estimated according to Chance and Maehly  is a lipid peroxidation biomarker and was assayed by a method described by Ohkawa et al. . The cond Maehly .). TNF-α content was determined using an enzyme-linked immunosorbent assay kit; the instructions from the manufacturer, NOVA kit, Beijing, China, were followed to calculate the results.CYP2E1 activity was measured in the liver microsomal fraction as described by Chang et al. . TNF-α cLiver total lipid (TL), total cholesterol (TC), and triglycerides (TG) were extracted using the procedure developed by Folch et al. . The conSome of the data were expressed as an inhibition percentage; IR% = [/control value] × 100.Specimens of all animals were dissected immediately after death and fixed in 10% neutral-buffered formal saline for at least 72 h. All the samples were washed in tap water for half an hour and then dehydrated in ascending grades of alcohol , cleared in xylene, impregnated in soft paraffin at 55 °C, and embedded in hard paraffin. Serial sections were cut at 6 µm thickness and stained with haematoxylin and eosin  followed by a post hoc test.Data were analysed as mean ± SE for eight rats each. Comparisons among groups were performed by one-way analysis of variance ANOVA test at In this study, TLRE contains 12.62 ± 1.21 mg polyphenols as gallic acid/g extract, 5.64 ± 0.44 mg flavonoids as quercetin/g, 9.55 ± 0.15 mg coumarins as umbelliferone/g extract, and 238.00 ± 1.77 mg total alkaloids/g extract. The phytochemical composition of TLRE was identified from the HPLC analysis. The identified compounds include gallic acid (399.76 µg/g extract), chlorogenic acid (1426.04 µg/g extract), syringic acid (143.80 µg/g extract), ellagic acid (180.25 µg/g extract), coumaric acid (58.74 µg/g extract), ferulic acid (208.22 µg/g extract), cinnamic acid (408.94 µg/g extract), propyl gallate (670.08 µg/g extract), vanillin (198.18 µg/g extract), naringenin (1419.65 µg/g extract), and quercetin (4182.24 µg/g extract) .−1 suggested the presence of NH, OH, aromatic ring, and ester carbonyl functionalities. The C-N stretching and C-N bending vibrations of a nitro group occur near 968 cm−1 and 700 cm−1, respectively  absorption bands at 3367, 3167, 1656, 1635, and 1464 cmm/z 308 and 281, referred to as the indole-alkaloid skeleton  database, https://massbank.eu/MassBank, and the MassBank of North America database, https://mona.fiehnlab.ucdavis.edu. Structure confirmation was carried out using NIST mass search software and suggested that the structure was corynan-17-ol, 18,19-didehydro-10-methyoxy, with a molecular formula C21H24N2O4 , p ≤ 0.05.The data presented in p ≤ 0.05). In the protective control rats, TLRE prevented the weight loss and increased the total body weight gain of these rats to 14.88 ± 1.28 g, compared to silymarin treatment, which increased the body weight gain 8.02 ± 0.87 g during the experimental period (p ≤ 0.05). In addition, the total body weight gain for the positive control of TLRE or silymarin was elevated compared to that recorded in the negative control group. Thus, the effect of TLRE on total body weight gain in the therapeutic and protective rats was close to that of silymarin treatment.TLRE increased the total body weight gain of the therapeutic rats, but silymarin did not significantly change it compared to the therapeutic control group (p ≤ 0.05) . The relative weight of the liver of therapeutic rats force-fed with either TLRE or silymarin was significantly reduced to 3.71 ± 0.28 and 3.94 ± 0.35%, respectively, compared to the value in the therapeutic control 4.34 ± 0.2% (p ≤ 0. 05). Either TLRE or silymarin protected the liver of protective-group rats, where the relative weight of the liver declined to 4.78 ± 0.3% and 4.88 ± 0.25%, respectively, compared to the value for the protective control 5.25 ± 0.3% (p ≤ 0.05). Compared to the negative control, the relative weight of the liver of the positive control for TLRE and silymarin was significantly reduced.The relative weight of the liver of protective rats was raised by 5.25 ± 0.3%, significant compared with that of the negative control, 4.42 ± 0.18% ( ≤ 0.05) . The relp ≤ 0.05)  . TLRE re4 administration caused remarkable inflammation, where TNF-α content significantly increased from 15.20 ± 0.55 pg/mg protein in the negative control rats to 38.83 ± 2.80 pg/mg protein in the therapeutic control and 30.01 ± 2.34 pg/mg protein in the protective control (p ≤ 0.05) (The CCl ≤ 0.05) . Treatme4 administration . In addition, albumin in CCl4-treated rats declined considerably in comparison with the level in the negative control.This study examined liver performance evaluated by liver function, which was dramatically affected by CClstration . Hepatocp ≤ 0.05). TLRE recovered and protected TP and its fractions at levels close to the effects of silymarin.TLRE as a therapeutic agent did not cause a significant effect on TP, albumin, and globulin concentrations compared to the corresponding values in the therapeutic control. Meanwhile, TLRE as a protective agent protected hepatocytes and maintained TP and its fractions at the normal levels . Thus, Tp ≤ 0.05). The effect of TLRE on total bilirubin concentration and ALP activity was close to the effect of silymarin. The bilirubin level of the positive control for TLRE did not change significantly; however, the ALP activity was significantly reduced compared to that of the negative control.TLRE decreased the total bilirubin level of therapeutic rats, to 0.85 ± 0.07 mg/dL, compared to therapeutic control (1.19 ± 0.09 mg/dL). Also, the ALP activity of the therapeutic rats was reduced to 162.22 ± 4.33 U/L, a significant change compared to the therapeutic control. In the protective group, TLRE maintained bilirubin concentration and ALP activity lower than the protective control (p ≤ 0.05). Also, TLRE exhibited a protective effect on AST and ALT close to the effect produced by silymarin. The AST activity of TLRE positive control did not change significantly compared to that of the negative control; however, the ALT level was reduced considerably to a level lower than for the ALT of the negative control.The activity of AST and ALT enzymes in serum was significantly lowered due to the administration of TLRE as a therapeutic agent . The AST4 caused remarkable oxidative stress status that appeared as an increment in H2O2 production in the therapeutic and protective controls: 6.65 ± 0.25 and 6.49 ± 0.29 μmol/g, significant when compared to the concentration in the negative control, 3.55 ± 0.27 μmol/g (p ≤ 0. 05) (p ≤ 0.05).Treatment with CCl≤ 0. 05) . In addi2O2 production in the liver tissue was significantly reduced to 3.57 ± 0.27 and 3.06 ± 0.21 μmol/g tissue, respectively, compared to the corresponding control. H2O2 production in positive control rats did not change significantly compared to the negative control. In comparison with silymarin, TLRE exhibited potent therapeutic and protective activities. No significant difference was recorded in H2O2 production between the rats treated with TLRE or silymarin and the negative control.The TLRE, either as a therapeutic agent or as a protective agent, ameliorated the oxidative stress status. The Hp ≤ 0. 05). In the protective group, the same trend took place. TLRE caused a decrease in MDA concentration in the liver and serum, reaching 17.03 ± 0.74 nmol/g and 7.65 ± 0.33 nmol/mL, respectively, which was significant compared to the protective control . No significant difference was noticed between the levels of MDA of the serum or the liver of the positive control rats and those found in the negative control rats.The levels of MDA of the therapeutic rats, whether in the liver tissues or released into the serum, were significantly reduced to 18.06 ± 0.85 nmol/g and 8.03 ± 0.48 nmol/mL, respectively, compared to the MDA levels for the therapeutic control of 24.46 ± 1.63 nmol/g and 10.81 ± 0.48 nmol/mL . The recovery effect on CAT activity produced by TLRE was better than that from silymarin in the therapeutic and protective groups. The CAT activity of the positive control did not alter significantly compared to the CAT activity of the negative control. TLRE improved the SOD activity in the therapeutic and protective groups, where the activity increased dramatically by about 72.44 and 56.39% compared to the values for the corresponding control. The effect of TLRE on SOD activity was close to the result shown with silymarin. TLRE did not affect the SOD activity of the positive control rats.Administration TLRE attenuated CCl4-induced injury to the liver glutathione system for the therapeutic and protective groups compared to effects in each control (p ≤ 0.05). The GSH content and related enzymes GR, GST, and GPx of the therapeutic group were significantly elevated to 16.61 ± 0.99 mg/dL, 15.83 ± 1.53, 14.17 ± 1.37, and 10.30 ± 0.43 U/mg protein, respectively, as a response to TLRE treatment, compared to levels in the therapeutic control. Meanwhile, TLRE prevented imbalance in the GSH, GR, GST, and GPx levels in the protective group of rats and maintained them close to the normal levels at 17.62 ± 1.74 mg/dL, 16.99 ± 1.14, 14.70 ± 0.99, and 8.67 ± 0.37 U/mg protein, respectively, compared to the protective control group. TLRE did not alter the glutathione system of the rats in the positive control group compared to the levels of the negative control group.The administration of TLRE improved CCl4-induced hepatotoxicity taking place in the rat liver increased the lipid accumulation in the liver, shown as total lipid (TL), total cholesterol (TC), and triglycerides (TG) of the therapeutic and protective control, compared to the levels for the negative control (p ≤ 0. 05), as shown in p ≤ 0.05). TLRE exhibited an anti-hyperlipidemia effect in the protective group of rats and prevented lipid accumulation in the liver there. TLRE restored the TL, TC, and TG of the protective group to within normal levels. Thus, TL, TC, and TG parameters were significantly decreased by about 33.98, 41.90, and 34.71%, respectively, compared with the corresponding values in the protective control (p ≤ 0.05). The positive group appeared with a normal liver lipid profile, where the TL, TC, and TG levels were decreased or did not change compared to the same parameters in the negative control. The improvements in liver lipid profile recorded in the silymarin-treated groups were close to those recorded in TLRE-treated groups.The CClIn examining the liver tissue sections, the livers of the negative control group appeared to have normal morphology and histology. The liver tissues for the negative control group appeared grossly brown-red. They presented normal hepatic lobules with normal central veins, hepatic cords, sinusoids, and portal tracts . Using e4 and after two days later received saline solution exhibited massive damage to the liver tissue in the form of necrosis and vascular degeneration of cells with distortion of the general architecture of the tissue  that are generated during cytochrome P-450 action on CCl4. These radicals are responsible for lipid peroxidation, inflammation, and fatty changes of the liver. Moreover, CCl4 metabolites are linked with oxidative stress. The CCl3· and CCl3O2· radicals disrupted the liver antioxidant system by inhibiting the liver’s antioxidant enzymes as CAT, SOD, GPx, GR, and GST. In addition, the CCl3· radicals deactivate GSH via reaction with its SH groups  concentration and enzymatic antioxidant activities . The anti-inflammatory effect of TLRE is evident in the remarked modulation in an inflammatory biomarker, TNF-α content. TLRE acted as a CYT2E1 inhibitor and modulated its activity to the normal level.TLRE significantly decreased the levels of AST, ALT, and ALP enzymes in the bloodstream, moving them towards the normal level, indicating that TLRE protected the structural integrity of the hepatocellular membrane and liver cells against damage caused by CClFrom previous data, we concluded that TLRE had hepatoprotective features. The hepatoprotective effects of TLRE may be related to its antioxidant aspects, anti-inflammatory features, CYP2E1 inhibition. Also, its content from polyphenols, flavonoids, and alkaloids, and its active constituents gallic acid, chlorogenic acid, naringenin, and quercetin.4 toxicity, especially hepatotoxicity, resulting in decreased levels of released ALT and AST in the bloodstream and activation of an antioxidant enzyme  activation of antioxidant enzymes , b) protection GSH, c) inhibition lipid peroxidation, and d) reduction of hydrogen peroxide.Oxidative stress has been implicated in the pathogenesis of various liver diseases, including alcoholic liver disease, non-alcoholic fatty liver disease, and chronic hepatitis C  reported the anti-inflammatory activity of TLRE coumarin in a mouse model of 12-Ο-tetradecanoylphorbol 13-acetate (TPA)-induced auricular edoema.Healthy hepatocytes are insensitive to TNF-α action, but they become sensitive once protein, and RNA synthesis is inhibited. Liver disease or liver failure is associated with the upregulation and downregulation of inflammatory biomarkers. TNF-α and NO have a vital function in inducing inflammation in oxidative stress-related hepatic diseases, promoting programmed cell death and fibrosis. TNF-α is an important cytokine that promotes hepatic damage , flavonoids (as naringenin and quercetin), alkaloids , and coumarins (as coumaric acid).Phenolic compounds downregulate liver enzyme biomarkers; AST and ALT. They can also restrain the oxidative stress and activation of T cells, a prime instigator of inflammation , attenuating the inflammatory mediators COX-2 through NF-kB inhibition pathway. TLRE contains gallic acid as a 399.76 μg/g extract, which may help in liver protection. Chlorogenic acid has a hepatoprotective effect in CCl4-induced hepatotoxicity rats via free radical-scavenging and antioxidative activity. Chromogenic acid (1426.04 μg) occurs in each gram of TLRE. Quercetin and ellagic acid have potent hepatoprotective, antioxidant, and anti-inflammatory activities against hepatotoxicity. Administration of QR and EA showed hepatoprotective effect through reducing liver biomarkers, improving the redox status of the tissue, and hampering the expression level of fibrosis-related genes  on other organs.In this study, tagetes extract exhibited a hepatoprotective effect against CCl


We investigated 180 consecutive patients with congestive heart failure (CHF), of whom 83 had type 2 diabetes (T2DM) and 97 did not have diabetes as well as 223 controls without CHF, of whom 39 had T2DM and 184 did not have diabetes. Data was recorded by standardized interviews and by standardized examination protocols at our institution and were extracted from medical records. Here, we analyzed data on gender differences. Further, we examined the effect of CHF and T2DM on moderate albuminuria, i.e. on an albumin-creatinine ratio (ACR) of 30–300 mg/g. Table 1 shows baseline characteristics of our patients stratified by gender. Table 2 gives ACRs and prevalence rates of albuminuria separately for men and women.p = 0.002 and OR 2.38 [1.11–5.11]; p = 0.027, respectively), as well as in women . There was no significant interaction between gender and CHF or T2DM regarding the presence of albuminuria , indicating that the association of CHF and T2DM with albuminuria did not differ significantly between men and women.In logistic regression analyses adjusting for age, sex, body mass index, LDL cholesterol, history of smoking, history of hypertension, use of statins, ACE inhibitors/angiotensin II receptor blockers, aldosterone antagonists and other antihypertensive medication CHF and T2DM predicted the prevalence of albuminuria in a mutually independent manner in men ; for men the corresponding F-values were 2.70; p = 0.102 and 3.12; p = 0.079, respectively. There was no significant interaction between gender and CHF or T2DM regarding ACR , indicating that the association of CHF and T2DM with the ACR did not differ significantly between men and women.In multivariate analysis of covariance, CHF and T2DM proved to be independent predictors of ACR in women after adjustment for age, sex, body mass index, LDL cholesterol, history of smoking, history of hypertension, use of statins, ACE inhibitors/angiotensin II receptor blockers, aldosterone antagonists and other antihypertensive medication (Regarding moderate albuminuria, both CHF and T2DM predicted moderate albuminuria adjusted in a mutually independent manner after the adjustments described above, with ORs of 4.75 [95% CI 2.16–10.45]; p< 0.001 and OR 2.08 [1.13–3.83]; p=0.018, respectively.The data set presented here could be reused with similar patient cohorts for pooled analysis. Albuminuria predicts cardiovascular morbidity and mortality in T2DM •The data benefit researchers as well as health care professionals in the fields of diabetology, cardiology, nephrology and general internal medicine.•Our data should stimulate the development of study protocols to gain further insight into the interplay between albuminuria, congestive heart failure and T2DM, including prospective investigations and interventional studies.1•p-value of 0.05.••Dataset: Features the data of our study population including the presence of CHF and T2DM, gender, ACR and prevalence of albuminuria. The data are pseudonymized as each participant was assigned to consecutive numbers (“ID”) on the basis of date of inclusion. The variables “Diabetes_mellitus_type_2” and “congestive_heart_failure” distinguish if subjects are suffering from T2DM and/or CHF, whereas “0” encodes “no” and “1” stands for “yes”. “Sex” describes the biological sex of subjects, whereas “0” stands for “female” and “1” for “male” patients. The variable “albumine_creatinine_ratio” describes the quantity of albumin-creatinine ratio (ACR) in mg/g. Finally, the variable “moderate_albuminuria” states if subjects suffer from moderate albuminuria, which is defined as an ACR of 30–300 mg/g, and as before “0” stands for “no” and “1” for yes.•Questionnaire: The questionnaire used for the standardized interview and examination of every subject.2We ruled in consecutive patients who were admitted to the LKH Feldkirch, a tertiary care center in Austria, for congestive heart failure. Eligible patients were identified via the hospital record system. As a control group we used consecutive patients who were admitted for coronary angiography with no signs or symptoms of CHF according to the 2016 European Society of Cardiology (ESC) definition The diagnosis of type 2 diabetes (T2DM) was made according to 2020 ADA criteria ACR was measured on either cobas c501/c502, c702 or Integra 800  using fasting morning urine . Moderate albuminuria was defined as a range of ACR of 30 mg/g or more and 300 mg/g or less.NT-proBNP was measured on cobas e601, e602, e801 or Elecsys 2010  using corresponding, commercially available, immunological tests. Hba1c-levels were measured on Adams HA-8160  using High Performance Liquid Chromatography.p value of 0.05. All statistical analyses were performed with the software package IBM SPSS Statistics 24.0.0.0 for Windows .Between-group differences were tested for statistical significance using Mann–Whitney–U test for continuous variables and by applying the chi-squared test for categorical variables. Analyses of covariance (ANCOVA) was performed using the general linear model approach, and logistic regression analyses were applied. Results are described as mean median [interquartile range] if not denoted otherwise. Statistical significance was defined as two-tailed We hereby state that the research has been carried out in accordance with The Code of Ethics of the World Medical Association (Declaration of Helsinki), that the Ethics Committee of the University of Innsbruck approved this study (EK-2-2008/0017) and that all participants gave written informed consent.Christoph H. Saely: Conceptualization, Methodology, Validation, Writing – review & editing, Supervision. Maximilian Maechler: Validation, Formal analysis, Investigation, Writing – original draft, Visualization. Alexander Vonbank: Writing – review & editing. Lukas Sprenger: Investigation, Visualization. Arthur Mader: Investigation, Visualization. Barbara Larcher: Investigation, Visualization. Daniela Zanolin-Purin: Investigation, Formal analysis. Andreas Leiherer: Writing – review & editing. Axel Muendlein: Writing – review & editing. Heinz Drexel: Conceptualization, Supervision, Project administration.The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.


Plastic mulching may reduce water productivity and cotton yield. HYDRUS-1D unsaturated flow model was used to access the groundwater recharge for 150 days under six treatments after model performance evaluation. Maximum cumulative recharge was observed 71 cm for the flat sowing method without plastic mulch. CanESM2 was used to predict climate scenarios for RCP 2.6, 4.5, and 8.5 for the 2050s and 2080s by statistical downscale modeling (SDSM) using historical data from 1975 to 2005 to access future groundwater recharge flux. Average cumulative recharge flux declined 36.53% in 2050 and 22.91% in 2080 compared to 2017 without plastic mulch. Multivariate regression analysis revealed that a maximum 23.78% reduction in groundwater recharge could influence future climate change. Further study may require to understand the remaining influencing factor of depleting groundwater recharge. Findings highlight the significance of climate change and the cotton sowing method while accessing future groundwater resources in irrigated agriculture.Irrigated agriculture is a foremost consumer of water resources to fulfill the demand for food and fiber with an increasing population under climate changes; cotton is no exception. Depleting groundwater recharge and water productivity is critical for the sustainable cotton crop yield peculiarly in the semiarid region. This study investigated the water productivity and cotton yield under six different treatments: three sowing methods, i.e., flat, ridge, and bed planting with and without plastic mulch. Cotton bed planting without mulch showed maximum water productivity (0.24 kg.m Meantime, a relationship of groundwater recharge and climatic parameters was also quantified by performing regression and correlation analysis.Gossypium) field, located in Rechna Doab upland of Punjab at Water Management Research Center (WMRC), Faisalabad , T2 (flat sowing without mulch), T3 (ridge sowing with mulch), T4 (ridge sowing without mulch), T5 (bed planting with mulch), and T6 (bed planting without mulch), respectively, with three repetitions. Plastic film with low-density polyethylene (LDPE) is used as mulching. The plot size was 6 m × 4.5 m, and the total experimental area was about 0.141 ha. The cotton crop was sown on June 13, 2017, and harvested on November 20, 2017, with three replications of each treatment. Cottonseeds were planted at 25 kg.ha−1 on edges by keeping up 75 cm row to row. Two seeds for every slope were dibbled to a profundity of 3 to 4 cm by separating 30 cm from plant to plant in a row to achieve the required plant population. Weeds management strategy was adopted from early season to harvesting.Randomized complete block design (RCBD), the sowing method, and land cover experimented with three sowing methods were studied, i.e., flat sowing, ridge sowing, and bed sowing with mulch and non-mulch land cover. There were six treatments, i.e., T3) and 3). In addition, climatic data of the Faisalabad station was acquired from Pakistan Metrological Department (PMD).Moisture content data was collected from the fields at 7-day intervals. The soil sampling was done at 0–30 cm, 30–60 cm, and 60–90 cm depth from each treatment plot. A fixed interval soil sample was collected using a vertical borehole (auger-hole) drilling method Fig. . The moi−1) in the form of DAP (di-ammonium phosphate), potash (75 kg.acre−1) in the form of sulfate of potash, and nitrogen (140 kg.acre−1) in the form of urea was used at the time of seeding as per standard guidelines of the Extension and Adaptive Research Wing (EARW), Punjab Agriculture Department. Nitrogen (35 kg.acre−1) in urea was applied after 30 days of sowing, during boll formation initiation, and boll formation completion. Irrigation water was applied as a rotational program followed by EARW, with fixed irrigation interval based on the sowing method. The irrigation interval is 7 days for bed and ridge sowing and 10 days for flat sowing cotton. Irrigation depth was calculated by Eq. (The recommended dosage of phosphorus (100 kg.acreQ is discharge of the outlet/tube well (cusec), T is time of irrigation (hour), A is area to be irrigated (acre), and D is depth of applied irrigation (inch).1where −3) was measured for each treatment using Eq. , t is time (day), h is the soil water pressure head (cm), z is the gravitational head as well as the vertical coordinate , K(h) is the unsaturated hydraulic conductivity (cm.day−1), and S is the soil water extraction rate by plant roots (cm3.cm−3.day−1).where θ is the volumetric soil water content , m [-], and n [-] are shape parameters. Ks is the saturated hydraulic conductivity (cm.day−1); l [-] is a pore connectivity parameter; and m = 1–1/n, when n > 1. The percentage of sand, silt, and clay and bulk density and van Genuchten parameters are given in Table where nd αr cm−, m [-], At first, the model’s domain and soil parameters were conventional taken into account by the measured particle-size distributions Table  and moni0) was estimated using the NIAB meteorological station’s recorded data by the Penman–Monteith equation. Meanwhile, the lower boundary condition was free drainage to count the root zone’s recharge flux. Water content was set as initial conditions to calculate the root water uptake (transpiration). The proposed method to include osmotic stress was applied as represented in Eq. The upper boundary condition was selected as atmospheric surface runoff that depended on evapotranspiration, irrigation, and rainfall. For this study, reference evapotranspiration  was computed by a regional crop coefficient (Kc) based on the initial season, mid-season, late season, and evaporation soil. Evaporation from the soil is predicted by the estimation of energy available at the soil surface.The governing equation is the water balance of the vertical soil domain covered by vegetation for a given period. There was no runoff during irrigation. The cotton plants’ interception of rainwater was neglected due to low rainfall amounts during the vegetation period. FAO-56 standards calculated potential evapotranspiration (ETo) by the Penman–Monteith method. Metrological parameters that include solar radiation, relative humidity, and leaf area index are used for the growth period. Meanwhile, crop evapotranspiration  and K(θ), coefficients of correlation (R2), and Nash–Sutcliffe efficiency (NSE) and calculated by Eqs. , an, anRMSE) by Eqs.  as folloX is the observed value, Y is simulated value removal, the bar (-) represents the mean value, and n is the number of samples.where https://climate-scenarios.canada.ca/, which has historical data of 26 climatic parameters from 1976 to 2005, respectively. Grid data 27 X and 44 Y of CanESM2 were downloaded from the above link. The data is accessible at a grid resolution of 2.8125° latitude × 2.8125° longitudes, respectively. Moreover, CanESM2 gives future climatic data under different representative concentration pathways (RCP) for three scenarios RCP 2.6, 4.5, and 8.5 from 2006 to 2100.Climate change scenarios are considered for the estimation of recharge flux at a regional level. Downscaled by the statistical downscaling modeling (SDSM) approach, it is a hybrid tool that supports examining the impact of climate change. Here, CanESM2 climatic variables are used and acquired from p-value, histograms, scatterplots, correlation matrix, and partial correlation with the assistance of the MLR model. A correlation matrix was preferred between predictands and CanESM2 predictors  predictands were different for each parameter and selected based on p-value and partial r-value, respectively. Therefore, it was cautiously chosen for maximum (minimum) temperature, rainfall, relative humidity, and wind speed.SDMS can be done in three steps. The first is predictor’s screening: the most relevant atmospheric parameters were chosen based on R2 (> 0.96) and low RMSE (< 0.4) for both calibration and validation period, consequently. In addition, NSE (> 0.95 ~ 1.0) is elucidated the good efficiency or better fitting of data. Evaluation performance has shown better agreement than the previous study  temperature, rainfall, relative humidity, and wind speed for the validation period from 1996 to 2005 of NCEP and CanESM2 predictors (95% confidence level). For better agreement with the results, 100 ensembles were used to obtain the average. Evaluation results Table  show thaThe third is scenario generator: it is used to simulate the results of future climate scenarios, i.e., RCP 2.6, RCP 4.5, and RCP 8.5, respectively. The future climate data of maximum (minimum) temperature, rainfall, relative humidity, and wind speed were predicted for 2006 to 2100 on a daily basis and studied for two periods, i.e., 2021 to 2050 and 2051 to 2080. The increasing trend of maximum (minimum) temperature Table . MoreoveMicrosoft Excel® 2019 add-in data analysis tool was used to develop a correlation and multivariate regression model. The linear relationship between tended groundwater recharge and climate variables was established to determine the groundwater recharge flux.2, T4, and T6) practice has proven to be more water-efficient than mulching . Cotton bed planting without mulch showed maximum water productivity (0.24 kg.m−3). Similarly, the cotton sowing method without mulching  has demonstrated high cotton yield than mulching . Meantime, cotton bed planting showed the highest cotton yield (1946 kg.ha−1), followed by ridge sowing without mulch (1886 kg.ha−1) and bed planting with mulch (1643 kg.ha−1). Although the cotton yield of the bed planting method is highest in the current study, still less than previously reported . Consequently, maximum water content was simulated at 30 cm depth, and minimum water content was simulated at 90 cm depth regardless of the mulching and sowing method. The cotton field has exhibited erratic fluctuations in water content during the season due to surface water and rainfall induction. Nevertheless, the observed data showed a swift decrease in water content than predicted by HYDRUS-1D. This discrepancy in the top layer (30 cm) would be ascribed to more rapid drainage (sandy loam soil) or higher evapotranspiration rates by cotton plants. The model was run under a ten-time step (0–10) and generated a hydraulic conductivity profile and soil layers’ hydraulic capacity. Hydraulic conductivity is a critical parameter to access the water flow into the soil stratum that increased to a certain soil depth then became constant  by HYDRUS-1D at 30 cm, 60 cm, and 90 cm depths corresponded closely to the observed moisture content (cm3.cm−3) for six treatments. Nevertheless, deviations between simulated and observed moisture content exhibited at certain depths  at treatment T6. Moreover, the observed moisture content was higher than the simulated moisture content at 30 cm depth for T4 and T6 due to the without mulching practice. The robustness of the HYDRUS-1D is evaluated based on R2, RMSE, and NSE values, as shown in Table R2 and RMSE values for the HYDRUS-1D model were ranged between 0.82 and 0.92 and 3.40E − 3 and 6.55E − 5, respectively. Overall, the average R2 (0.875) and RMSE (5.24E − 4) were reliable through the whole soil profile. In addition, NSE values (0.85–0.99) explained further the reliability of the model. These results show consistency with previously groundwater recharge flux estimation using HYDRUS-1D under agricultural fields , followed by T4 (70 cm) and T5 (62 cm). The crop’s cumulative actual root water uptake was highest under T6 (83 cm), followed by T5 (78 cm) and T4 (72 cm). The estimated cumulative infiltration was observed maximum under T2 (140 cm) and T4 (140 cm). Plastic mulching of cotton crops under T1, T3, and T5 has reduced soil water content drainage due to climate variability, i.e., rainfall. Meantime, sowing of cotton without plastic mulch has shown more cumulative recharge flux than with plastic mulch. That may have a decisive influence on a water-saving strategy to mitigate water stress in agriculture in rain-fed semiarid crop cultivation areas. Meanwhile, the cumulative water fluxes were reported more in flood irrigation over mulched drip irrigation  for the RCP 8.5 2080 scenario. Overall, the actual root water uptake, infiltration, and evaporation trend increased in 2080 than 2050, creating uncertainties in sustainable water resource management , cumulative infiltration 118–160 cm), and cumulative evaporation (1.9–2.45 cm) were observed under future changes in climate as compared to 2017  declined 36.53% for 2050 and with plastic mulch  declined 22.91% for 2080 with a comparison of 2017 . Multilinear regression was conducted to quantify the impact of climate variables on recharge for future changes in climate in Table r) value shows weak uphill linear relationships between groundwater recharge and climate variables. Sign of coefficients indicates heterogeneity of groundwater recharge versus climate variable. Results revealed that the model is only bound to elucidate the variation in groundwater recharge ranging from 5 to 23.78%  under climate variables. In T6-RCP 2.6, climate variables account for only 23.78% of groundwater recharge flux. Precipitation can only contribute up to 4% of recharge flux , T2 (flat sowing without mulch), T3 (ridge sowing with mulch), T4 (ridge sowing without mulch), T5 (bed planting with mulch), and T6 (bed planting without mulch). Water productivity, cotton yield, and the future impact of changing climate on groundwater recharge were investigated. Here is the key finding of the study:−3) and the highest cotton yield (1946 kg.ha−1). Plastic mulching may reduce the water productivity as 19.6%, 22.2%, and 14.3% and cotton yield as 19.8%, 23.3%, and 15.6% compared to without plastic film mulching in flat, ridge, and bed planting sowing method.Cotton bed planting without mulch showed maximum water productivity (0.24 kg.mR2 (0.82–0.92), RMSE (3.40E − 3–6.55E − 5), and NSE (0.85–0.99) that depict good reliability of the model for the assessment of groundwater recharge under the irrigated cotton field. HYDRUS-1D was simulated from sowing to harvesting periods that are 150 days under six treatments. Maximum cumulative recharge was observed 75 cm for T2. Cumulative actual root water uptake was highest at 83 cm for T6. Cumulative infiltration was observed maximum under T2 (140 cm) and T4 (140 cm). Plastic mulching of cotton crops under T1, T3, and T5 has reduced soil water content.The performance of the model was evaluated by Using historical data from 1975 to 2005, SDSM was conducted by CanESM2 for the 2050s and 2080s climate scenarios RCP 2.6, 4.5, and 8.5. A temperature rise is expected at 2–4 °C. Meantime, rainfall, relative humidity, and wind speed will decline significantly.2, T4, and T6). Multivariate regression analysis revealed that only a 5–23.78% reduction in groundwater recharge could influence future climate change. There is a weak (positive) relationship between groundwater recharge and climate variables.Average  cumulative recharge flux (cm) declined 36.53% in 2050 and 22.91% in 2080 with a comparison of 2017 for without plastic mulch (TAn experimental study was conducted in a semiarid region to investigate the groundwater recharge flux of irrigated cotton fields under six treatments, i.e., TTherefore, it is equally important to adapt climate change measures for sustainable groundwater resources to avoid food security risks in the future. It is recommended that the current cotton sowing shall be replaced with bed planting, and mulching practice should be prohibited for greater cotton yield to achieve sustainable development. Further study requires to understand the remaining influencing factor of depleting groundwater recharge.