Saccharomyces cerevisiae and Wickerhamomyces anomalus solutions were added to Fengmi Qu (FMQ) from Fangxian, China to produce IQ, and brewing trial was conducted. High-throughput sequencing (HTS) was used to analyze the microbial community in fermentation broth of IQ (IQFB). Headspace solid-phase microextraction (HS-SPME) combined with gas chromatography–mass spectrometry (GC-MS) and sensory evaluation were performed to analyze volatile aroma compounds (VACs) in sample without Qu and both fermentation broths. The results showed that Pediococcus, Cronobacter, Enterococcus, Weissella, and Acinetobacter and Saccharomycopsis, Wickerhamomyces, and Saccharomyces were dominant bacterial and fungal groups, respectively. A total of 115 VACs were detected, and the content of esters including ethyl acetate, isoamyl acetate, and so on was noticeably higher in IQFB. The finding of sensory evaluation reflected that adding pure yeast to Qu could enhance fruit and floral aromas. Correlation analysis yielded 858 correlations between significant microorganisms and different VACs. In addition, prediction of microbial community functions in IQFB revealed global and overview maps and carbohydrate metabolism to be the main one. This study is advantageous for further regulation of the fermentation process of Huangjiu by microbial means.In recent years, intensified Qu (IQ) has been gradually applied to brewing in order to improve the aroma of Huangjiu (Chinese rice wine). In this study,  Thered minpai . It is tevisiae) ,5. The pevisiae) ,6,7.Rhizopus from glutinous rice wine starter and added it to Saccharomyces cerevisiae in different proportions to manufacture fortified Qu. Compared with common commercial koji, the saccharification power and liquefaction power of fortified Qu increase 14.8% and 20.7%, respectively. After brewing with this Qu, the total ester and alcohol contents also improve 168.9% and 92.1%, respectively. Cui et al. [Qu is the feature of Chinese brewing and plays an important role in the saccharification and fermentation process of wine . As a sai et al.  made IQ Lactobacillus, Neisseria, Staphylococcus, Thauera, and Bifidobacterium were the functionally predominant bacteria to produce higher alcohol, which were bound up with the production of 2-methyl-1-propanol, phenethyl alcohol, 2-phenoxy-ethanol, 1-triacontanol, and 1-hexacosanol, as well as provide new ideas for controlling the generation of higher alcohol. Ren et al. [Lactococcus, Virgibacillus, Sphingobacterium, and Sporolactobacillus, providing reference for developing new varieties of Huangjiu.Recently, as the important quality standards for evaluating Huangjiu, aroma and flavor were researched and analyzed extensively by a great deal of scholars. Chen et al.  studied n et al.  found thDue to the advantages of geographical location and environmental conditions, there is a superb place called Fangxian in Shiyan City, Hubei Province, China for making Huangjiu. Therefore, fuzhi Huangjiu has become one of the local specialties. The Qu of fuzhi Huangjiu is added to a Chinese herbal medicine called polygonum, also known as JIUYAO. The JIUYAO is made of japonica rice, polygonum, high-quality JIUYAO made in previous years, and water. However, the production of JIUYAO is carried out in an open environment, where different kinds of microorganisms are enriched in the JIUYAO, resulting in its unstable quality and a plethora of miscellaneous bacteria. Consequently, on the basis of previous JIUYAO, improvements were made to develop a formula of IQ, and two types of Qu were also used for Huangjiu fermentation, respectively. Then, the fermentation broths were collected during the fermentation process and their physicochemical indexes were measured according to national standards and other methods. Next, amplicon sequencing was applied to comprehensively analyze the microbial community of the fermentation broths and headspace solid-phase microextraction (HS-SPME) combined with gas chromatography–mass spectrometry (GC-MS) was used to identify their volatile aroma compounds (VACs). The fermentation broths were also subjected to sensory evaluation. Finally, by means of a correction analysis, the relationship between dominant microorganisms and VACs in fermentation broths was established, and the metabolic pathways of microorganisms were predicted. In this study, IQ was produced by adding two types of yeast to the previously made JIUYAO to strengthen the aroma of Huangjiu, which provides a reference for stabilizing the quality of Qu and enhancing the quality of Huangjiu.Rhizopus oryzae were obtained from China Industrial Microbial Strain Preservation and Management Center. Copper sulfate pentahydrate, potassium sodium tartrate, sodium hydroxide, glucose, methylene blue, hydrochloric acid, and methyl red  were purchased from Sinopharm Chemical Reagent Co., Ltd., Beijing, China. 2-octanol was purchased from Beijing Bailingwei Technology Co., LTD, Beijing, China. Edible lactic acid was purchased from Henan Jindan Lactic Acid Technology Co., Ltd., Zhoukou, China.Japonica rice was obtained from COFCO Corporation. Wuchang glutinous rice was obtained from Wuchang Yinhe Rice Industry Co., Ltd., Wuchang, China. Fengmi Qu (FMQ), polygonum, bran, and rice husk were purchased from Fangxian, Shiyan City, Hubei Province, China. Glucoamylase for brewing and CICC41435 200 g of glutinous rice were washed and soaked for 15 h. After steaming rice, 400 mL of water and 3.0 g of glucoamylase were added, then saccharified for 3–4 h at 60 °C. After filtration, the sugar content was diluted to 14 °Bx, and then sterilized at 115 °C for 15 min to prepare rice starter juice.30 g of rice husks were washed, moistened with boiling water, filtered dry, and mixed with 120 g of bran. 105 mL of hot water above 90 °C was added to moisten the grains for 1 h, and 7.5 mL of 14 °Bx rice starter juice was added. After mixing, they were placed in 10 triangular flasks and sterilized at 121 °C for 30 min to obtain mold expanded culture medium.Rhizopus oryzae was inoculated into the expanded culture medium, cultured for 3–5 days, and mold spores were washed with sterile physiological saline. After filtering, the spore count was adjusted to 107 CFU/mL to prepare mold spore suspension.Rhizopus oryzae spore solution were added. They were mixed well and left to stand for 1 h. Thirdly, 2% Saccharomyces cerevisiae solution and 1% Wickerhamomyces anomalus solution were added, and the water content was replenished to 60%. The above two strains were isolated and identified from JIUYAO in Fangxian by our laboratory. Fourthly, they were made into balls or blocks, coated with powder (1% FMQ powder), and then put into a plate with holes at the bottom, where two layers of dry gauze were placed. After adding the Qu, they were covered with a damp gauze cloth on the plate. Finally, in the fermentation chamber, they fermented for 24–36 h, and the temperature was controlled at 30–35 °C until the hyphae of Rhizopus oryzae covered the surface before removing the gauze. After being transferred to a cool and ventilated place, they were further cultured for 2–3 days to measure their moisture and saccharification power.The production method of IQ was as follows: firstly, 252 g of rice flour, 30 g of bran, 18 g of rice husk, and 1.5 g of polygonum were taken, then crushed and sieved through 60 mesh screens. Secondly, 2% FMQ, 130 mL of edible lactic acid (pH 5–5.5), and 2% The saccharification power and moisture content of Qu were determined using the QB/T 4257-2011 General Analysis Method for Brewing Daqu.After washing 1.0 kg of glutinous rice, it was soaked for 15 h. The steamed rice was cooled to 30–35 °C in an ultraclean bench . Approximately 600 mL of water was poured into the glutinous rice to disperse the rice, while adding 0.7% FMQ and IQ separately. They were stirred evenly and a nest was made for the glutinous rice, which was fermented for 5 days at 30 °C. After 600 mL of sterile water was added, the fermentation broths were stirred evenly and fermented at 15 °C for 27 days. At the same time on the 3rd, 5th, 9th, 14th, 20th, and 27th days of fermentation, samples of Huangjiu fermentation mash were aspirated out. Sterile straws were used to place fermented mash into sterile centrifuge tubes. The sample before mixing Qu was recorded as Blank. Fermentation broth brewed with FMQ was labeled as FMQFB, while that brewed with IQ was labeled as IQFB. Samples were stored in a −20 °C refrigerator for analysis.The pH value, total sugar, and total acid of Huangjiu fermentation broths were measured according to the determination methods of pH, total sugar, and total acid in the national standard GB/T13662-2018 Huangjiu.The alcohol contents of Huangjiu fermentation broths were determined by a 1260 high-performance liquid chromatograph . To begin with, the fermentation broths were filtered through 0.45 µm polyethersulfone membranes and placed in liquid phase vials for later use. Next, 1, 2, 4, 8, 12, 16, and 32% ethanol aqueous solutions were prepared through membrane filtration to be used later.The conditions of HPLC were set as follows: the mobile phase was ultrapure water with a flow rate of 0.8 mL/min. The detector was RID with a temperature of 55 °C. Hi Plex H (300 × 7.7 mm) was used as a chromatographic column. The temperature of the column temperature box was 70 °C.The approaches for DNA extraction and amplification were consistent with a literature report . A GeneAThe purified and identified PCR products were sequenced by the Majobio Institute . The MiSeq platform of an Illumina high-throughput sequencer  was used to sequence.The VACs of fermentation broths were detected by HS-SPME combined with GC-MS. 5 mL of fermentation broth was added to a 10 mL headspace extraction bottle, mixing 2.0 g of NaCl and 0.020 mL of 2-octanol (99.07 mg/L). They were equilibrated in a water bath at 50 °C for 20 min, and then the SPME extraction head  was inserted into a headspace bottle. It was pushed out, extracted at 50 °C for 50 min, and then desorbed at 250 °C at the GC injection port for 5 min [The instrument method of GC-MS  was as follows : the injQualitative analysis of VACs was conducted through the NIST 2017 spectral library (compounds with a retention matching degree greater than 700) and retention index (RI) . The difThe sensory evaluation method for the aroma of Huangjiu fermentation broths was carried out referring to a previous study . Fifteenwww.drive5.com/usearch/, accessed on 18 May 2023) software was used to extract the high-quality sequencing readings. The RDP classifier Bayesian algorithm was conducted for taxonomy analysis of 97% similar OTU representative sequences on the Qiime platform (version 1.9.1), and the confidence threshold was 0.7.Usearch  was used. A correlation network between volatile metabolites and microbial communities was established by using Cytoscape software (version 3.9.0).p < 0.05.To predict the function of microbial communities, the PICRUSt software package was utilized. The evolutionary genetics of genes: Non-supervised Orthologous Groups database (eggNOG) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) database were used for functional annotation.The statistical software SPSS (version 27.0) was used for variance analysis. The significance level was considered at FMQ and IQ were applied to the fermentation experiment of Huangjiu, and the physicochemical indicators including total acid, total sugar, pH, and alcohol content during the fermentation process were monitored. The results are shown in According to PCR amplification and Illumina MiSeq sequencing were applied to analyze the diversity of bacterial and fungal communities during the fermentation process of IQFB. After eliminating low-quality reads and chimeras, 1,380,647 high-quality sequences with an average sequence length of 428 bp could be obtained in the bacterial community. On the basis of a similarity level of 97%, 230 operational classification units (OTUs), which were classified by the Silva classification database, were obtained through clustering using the algorithm named USEARCH11-uparse. In the fungal community, 1,395,768 high-quality sequences could be obtained with an average sequence length of 284 bp. The sequences of the fungi were clustered to obtain 77 OTUs, which were classified and annotated by the UNITE classification database. There was no prominent difference between the sequence numbers of bacteria and fungi in IQFB. Yet, the total number of bacterial OTUs was distinctly higher than that of fungal OTUs, which was consistent with the analysis of JIUYAO in Fangxian .Wickerhamomyces anomalus and Saccharomyces cerevisiae, to IQ. And the diversity index of fungi was lower than that of bacteria, which was consistent with the analysis results of OTUs.The coverage, richness, and diversity of bacteria and fungi in different fermentation stages of IQFB were analyzed, and the distribution of Coverage, Sobs, Chao, Shannon, and Simpson as indices at the OUT level are displayed in Firmicutes, Proteobacteria, Cyanobacteria, Actinobacteriota, and Bacteroidota, as presented in Ascomycota, Basidiomycota, and Mucoromycota phyla were identified for fungi, as shown in unclassified_k_norank_d_bacteria was also identified in the bacterial community, which also belonged to the dominant flora but accounted for a small percentage of IQFB. Furthermore, Firmicutes had a higher percentage in IQFB than in FMQ, greater than 97% in IQFB. In the fungal community, both demonstrated the same phylum, but Ascomycota had a higher percentage in IQFB, with all being greater than 99%.By sequencing, 20 phyla, 102 families, and 156 genera and 3 phyla, 40 families, and 54 genera were identified in the bacterial and fungal communities of IQFB. The top ranked species of bacteria at the phylum level including Pediococcus, Cronobacter, Enterococcus, Weissella, and Acinetobacter were the prominent clusters. Pediococcus accounted for 79% of the blank sample, but this was far below its representation on day 3 (99%), day 5 (98%), day 9 (97%), day 14 (98%), day 20 (96%), and day 27 (98%). When the microbial community of Huangjiu produced around the Winter Solstice was evaluated, Pediococcus was recognized as the dominant genus in the later stages of fermentation [Pediococcus generates acid but not gas, which encourages the production of some specialized flavor substances during the fermentation process [Weissella and Pediococcus had a decisive role in the flavor characteristics of Huangjiu [Rhodococcus was almost only present in the blank sample and on day 20. As for fungi, the prevailing phyla included Saccharomycopsis, Wickerhamomyces, and Saccharomyces, which constituted 99.70%, 99.48%, 99.39%, 99.60%, 99.37%, 99.41%, and 98.32% on day 0, day 3, day 5, day 9, day 14, day 20, and day 27, respectively. Saccharomycopsis was observed in 69% of the blank samples, while after the addition of Qu, its percentage was higher than 70% in all fermentation broths, with a maximum of 83% on day 3. Saccharomycopsis and Wickerhamomyces were also the dominant genera identified in FMQ, with Wickerhamomyces occupying more than 80% [Saccharomycopsis, which facilitates the glycation of starch [Saccharomycopsis was identified as the main microbial community in JIUYAO of Shaoxing-jiu and exerted a vital influence on fermentation activity and flavor characteristics [Saccharomyces is one of the primary genera of yeasts that produce alcohol [Saccharomycopsis acted synergistically with Wickerhamomyces, it resulted in a significant contribution to the production of ethyl acetate as well as an improvement in the concentration of VACs such as β-phenethyl alcohol and phenethyl acetate [Wickerhamomyces is one of the essential substances involved in the synthesis of phenethyl acetate [The association between samples and species was explored to more visually analyze the percentage of species in different fermentation stages, and a Circos relationship diagram is shown in entation . It not entation . Pedioco process . There wHuangjiu . Rhodocothan 80% . Howeverf starch . Sacchar alcohol . When Sa acetate . Apart f acetate .By means of HS-SPME combined with GC-MS, VACs during the brewing process of FMQFB and IQFB were analyzed . A totalThe contents of VACs in Huangjiu fermentation broths are shown in During the fermentation of FMQFB, the content of all VACs except aldehydes showed an increasing trend in the early stage of fermentation (1–5 d); in the middle stage (6–19 d), the content of aroma substances presented an up-trend followed by a small decrease and then continued to go up; in the late stage (20–27 d), the content of alcohols, esters, and furans exhibited a rising tendency, while other VACs showed a decreasing trend. In addition, during the fermentation of IQFB, an increasing trend followed by a decreasing trend was exhibited by most of the VACs in the early (1–5 d) and middle (6–19 d) periods; in the late fermentation period (20–27 d), a significant increase was presented by all VACs except nitrogenous and sulfurous compounds.In both types of fermentation broth, the content of alcoholic compounds was highest in each stage, with 11,821.77 µg/L and 7352.84 µg/L in FMQFB and IQFB, respectively, on the 27th day of fermentation. This was inconsistent with previous studies reporting ester as having the highest content in the Huangjiu fermented from corn , which mIn general, the content of various volatile aroma compounds showed an increasing trend in the early period and a fluctuating increase in the middle and late period, reaching the highest value on the 27th day of fermentation. The content of esters such as ethyl acetate, isoamyl acetate, ethyl caproate, ethyl heptanoate, and ethyl nonoate was significantly higher in IQFB compared to FMQFB. The categories of VACs in IQFB were higher than in FMQFB and the content was lower than in FMQFB on the 27th day. Moreover, compared with esters, alcohols, and acids at the same stage, other compounds such as aldehydes and ketones had lower content in the two fermentation broths, but they also contributed to the aroma of Huangjiu. It was the difference in the type and content of VACs in FMQFB and IQFB that led to differences in their aromas.In order to further understand the aroma differences between FMQFB and IQFB, sensory evaluations on the aroma characteristics of the blank sample and two types of Huangjiu were conducted by 15 sensory evaluation members, as shown in th day and, at the same time, it also had the lowest descriptor score among the eight aroma attributes in the sensory evaluation. Based on our previous work, it was found that Wickerhamomyces anomalus could increase the content of isoamyl acetate [From  acetate , which wSaccharomyces and indole, Saccharomycopsis and ethyl (E)-4-decenoate, Wickerhamomyces and decanal, and Wallemia and ethyl nonanoate, diethyl succinate, citronellol, cedrenol, and 4-hydroxyphenethyl alcohol. Wallemia had been detected from a Maotai-flavor liquor brewing environment, being the absolute dominant fungal genus in each round and having a strong positive correlation with the growth metabolism of Aspergillus [Saccharomycopsis, which had the highest abundance, also showed a negative correlation with decanal. Meanwhile, Wickerhamomyces, which was much more abundant, was associated with ethyl (E)-4-decenoate and acetoin. The high predominance of Cronobacter was negatively correlated with most compounds as well as highly correlated with the metabolism of long-chain fatty acid esters such as ethyl tetradecanoate, methyl hexadecanoate, ethyl hexadecanoate, ethyl octadecanoate, and ethyl oleate. The unclassified_f__Metschnikowiaceae in fungi was also strongly associated with long-chain fatty acid esters.To further explore the relationship between microbial communities and aroma, the correlation coefficients between microorganisms with > 0.1% abundance and VACs were calculated using Spearman correlation analysis. There were 352 positive correlations and 506 negative correlations, and a total of 858 correlations were obtained. Correlations with correlation coefficients |r| > 0.7 were visualized using Cytoscape, yielding 42 associations of microorganisms and VACs, shown in ergillus . This prergillus ,47. AmonA total of 23 categories were classified by COG function classification, shown in Based on the KEGG database, 6, 45, and 340 level were classified to 1, 2, and 3 pathways, respectively . For patSaccharomyces cerevisiae solution and Wickerhamomyces anomalus solution were added to produce IQ, and brewing trials were conducted using these two types of Qu. Microbial diversity analysis of IQFB at different brewing stages revealed that Pediococcus, Cronobacter, Enterococcus, Weissella, and Acinetobacter and Saccharomycopsis, Wickerhamomyces, and Saccharomyces were the dominant bacterial and fungal groups, respectively. Compared to fungi, bacteria had a greater diversity and abundance. 92 and 74 compounds were identified by HS-SPME combined with GC-MS for the fermentation broths of the two Qu, correspondingly. At the end of fermentation, more species of VACs were found in IQFB than FMQFB and the content was lower than in FMQFB. Among them, the content of esters such as ethyl acetate, isoamyl acetate, ethyl caproate, ethyl heptanoate, and ethyl nonanoate in IQFB was significantly higher compared with FMQFB. According to the sensory evaluation, it was shown that the addition of pure yeast to Qu could enhance the fruit and floral aromas of Huangjiu. A network of relationships between important microbial communities (abundance > 0.1%) and aroma in IQFB was established, and 858 correlations could be gained. The functions of microbial communities were predicted, finding global and overview maps and carbohydrate metabolism to be the most dominant. This will provide reference data for stabilizing the quality of Qu and improving the quality of Huangjiu, which could facilitate further regulation of the fermentation process of Huangjiu by microbiological measures.In this paper, by using FMQ as seed Qu,


The co-application of biochar compost as organic amendment for crop production and soil remediation has gained momentum due to their positive effect on plant growth and soil quality improvement. The application of biochar and compost which are green and cost-effective soil remediators would promote the availability and distribution of food, planetary conservation, alleviate poverty, and enhance the attainment of Sustainable Millennium Development Goals (SDGs). A bibliometric analysis was conducted to overview research on biochar compost from 2011 to 2021. Two hundred and fifty-four research papers were retrieved from the Scopus database and analyzed using VOS viewer. Analysis revealed that 217 (85.43%) were articles, 21 (8.27%) were conference papers, and 12 (4.72%) were review papers. The results showed an exponential increase in the number of publications. The most productive countries in the investigated subject were China (49), followed by USA (36), Australia (29), Italy (28), Germany (25), and Indonesia (20). After the search terms, ‘soil,’ which had links with keywords like ‘soil fertility,’ ‘soil quality,’ ‘soil pollution,’ ‘phosphorus,’ ‘nitrogen,’ ‘maize,’ ‘greenhouse gas,’ etc., had the highest occurrences (94). From the results of the current hotspot research in the field, the effect of biochar-compost mixture and co-composted biochar on soil remediation is currently being studied by several researchers. Biochar and compost incorporation in soil reduce the uptake of pollutants by plants which consequently increase essential nutrients for plant and soil productivity. ● Biochar-compost research has been increasing for the past eleven years● China is most productive country in biochar and compost application●Biochar-compost and co-composted biochar are effective elixirs for soil amendment and nutritional enrichment  ArouThese would adversely affect the availability of agricultural land, food distribution, and climate change, which threatens the ecosystem. Together with United Nations Sustainable Development Goals (UNSDGs) and the conservation of the planet, land reclamation, protection, and nourishment are critically necessary to accommodate a rising population. Based on  safe recThe low cost of biochar promotes its multifunctional application in science and technology. When incorporated into animal feed, biochar aids digestion; feces from animals fed with biochar constituted meals have less odor and can be used as organic manure . BiocharThe combined co-application of biochar and compost to agricultural soils has been increasing in research trends, including the application of co-composted biochar as a final product from the addition of biochar to raw composting materials for plant growth and improved soil quality ; 25, andThe current bibliometric analysis critically examined the biochar-based compost for agricultural purposes. Bibliometric analysis is an important tool for quantitative analysis that has been used in various fields to effectively analyze field development or the overall trend of a subject . To unde2.2.1.−1 on soil nutrient profile and maize (Zea mays) yield. The feedstock used was cow manure for both biochar and compost. The authors reported an 80.52% increase in soil organic matter, a 0.96%, 12.53 mg kg−1, and 208.33 mg kg−1 increase in total N, soil available P, and extractable K content respectively; in the treatment with both biochar and compost. The pH of the calcareous soil used in their study declined from 7.8 to 6.8, and the decline was associated with the increasing availability of P in the soil after combined application with biochar and compost due to changes in soil pH and amelioration of Ca2+. Compared to the control treatment, agronomic effects of biochar and compost on maize yield were reported on the increased 1000 grain weight (14.70%), plant height (43.23%), harvest index (9.90%), including an increase in biological and grain yield (13.44% and 46.29%). While a significant increase in grain N (51.2%), P (48.14%), and K (3%), and the highest shoot P-value of 3.20 g kg−1, and a K content of 16.80% compared to the control treatment were reported.For many years, compost has been used as a natural soil amendment to enhance crop yield . Based oCarica papaya cv Sunrise), and Chinese cabbage biomass . The incorporation and surface application of the mixture resulted in higher extractable P, N, Ca, and Mg. The authors also reported increased plant biomass in both tested crops. Between the two soil types, a greater response in terms of improved soil pH, root biomass, and plant nutrient uptake was reported in acidic Andisol than the Oxisol. The results of these studies [ using An studies ; 41, sho studies  and calc studies  soils ca studies , reporte studies  co-appli studies , the mix2.2.−1), and under different irrigation regimes ) resulted in increasing the productivity, leaf area, leaf number per plant, dry weight, the stem of the diameter of eggplant, including increased chlorophyll content, stomatal conductance, and photosynthetic efficiency of water-stressed eggplant at 80% ETc. The authors associated their findings with increased soil properties such as hydraulic conductivity, pH, EC, bulk density, water-holding pores%, useful pores%, fine capillary pores%, available water, and soil biota after the application of co-composted biochar to salt-affected soils. Interestingly, the utilization of biochar can improve animal waste composting when done at an application rate of 5–10% [3 by 30% and 44%. The highest application rate could induce desiccation in composting due to excess aeration [In numerous studies, the effect of co-composted biochar on the fertility of soil and crop productivity has shown a great potential of the material to enhance soil fertility and crop productivity. For example, in the study of  the effeof 5–10% ,52. In tof 5–10%  the applaeration . It is waeration  since th2.3.Even though compost has high nutrient content , increasing microbial activity and soil enzymes , howeverThe hyphenated application of biochar and compost for heavy metals contaminated wetland has been reported by . Differe2+ in the soil and compost. The uptake of toxic Cr (VI) is preceded by attraction to Fe3+ in the compost, followed by reduction by zero-valent iron Fe (O) to less toxic Cr (III) which is cleared via ligand exchange by N-H and C-O functional groups. Benzene and phenyl compounds are adsorbed via pi-pi bond interaction. Cu2+ would readily form chelate or complex with the OH groups on the biochar and compost , and Cd (II) are readily exchanged with elemental Ca compost .3.The data on the biochar-based compost was retrieved from Scopus, a citation and abstract academic database launched in 2004 by Elsevier. Scopus is an exhaustive summary and citation database with millions of records from journals, conference proceedings, and books. Based on  the data3.1.A bibliometric analysis was used for the literature review. In the present study, 254 publications were selected after data refinement, and based on  the bibli) of cited publications in a countable set i number of citations. The h-index, which integrates the number of citations , and the number of publications , was used to assess the performance and impact of publications in a qualitative view [where h-index is equivalent to the maximum number , followed by conference papers. An increase in the number of publications by 97.40% between 2011 and 2021 was observed, with the highest number  of publications in 2021. In 2021, the highest number were published under the discipline of Agricultural and Biological Sciences  and environmental Sciences  and published under the journal of IOP Conference Series Earth and Environmental Science , and journal of Science and Plant Nutrition . The results of the growth rate of annual citations showed a zigzag pattern, with a peak reached in 2017, with a corresponding increase in the h-index in 2017. It is worth noting that, after a decline from 2011 to 2014, a sharp increase from 2014 and 2015 in both the number of citations and h-index was observed.To assess the academic influence of authors, the number of publications and citations is measured , while t4.2.The map in 4.2.1.Environmental Science and Agricultural and Biological Sciences (n = 130). The third category is the field of Earth and Planetary Sciences, with the contribution of 7% (n = 27) publications, followed by Biochemistry, Genetics, and Molecular Biology . All the categories had more or less than 10 publications, with the lowest top three under the subject of Chemical Engineering, Engineering, and Multidisciplinary. An annual publication output showed an exponential trend from the top two subject categories with 130 documents each (R2 = 0.8943 and 0.868). It should be noted that, compared to the Environment Science field, the growth trend in the number of publications under the Agricultural and Biological Sciences field showed a continuously increasing trend from 2011 to 2021, respectively, with a sharp increase from 2020 to 2021.The topic of biochar and compost has been discussed in 20 disciplines, with the top 10 displayed in 4.2.2.Chemosphere, Science of the Total Environment, Journal of Soil Science and Plant Nutrition, Journal of Environmental Management, Waste Management, Agriculture, Ecosystems and Environment, Applied Soil Ecology, and Journal of Hazardous Materials) as shown by the total contribution of 22.44% (57 publications), respectively. After Elsevier, the second-ranking publisher was mdpi (Agronomy and Sustainability). Moreover, regarding the number of citations, sources followed the order science of the Total Environment  > Journal of Hazardous Materials  >, Agriculture, Ecosystems, and Environment  >, Chemosphere , etc. Notably, 80% of the sources were in the quartile in category 1 (Q1), which shows that many papers in biochar compost research were published by high-level journals, while 60% of the sources had a Cite Score above 5.The results showed that the top four most productive sources with the contribution of 48 publications (18.90%) on biochar and compost are from different publishers . Among t4.3.S) and Google Scholar (CGS) databases, including the year of publication and sources/journals. Based on [Based on  the analBased on . The papBased on , ‘BiochaBased on , ‘BenefiBased on , ‘EffectBased on . All the4.4.Zea mays,’ ‘greenhouse gas,’ etc.. In the green cluster, the keyword ‘soil,’ is also associated with ‘soil pollution,’ ‘soil pollutant,’ ‘soil remediation,’ ‘heavy metal,’ ‘contaminated soil,’ ‘adsorption,’ ‘bioremediation,’ etc.. The four clusters show the research hotspots on the topic of biochar-based compost, and these can be divided into four; red cluster: application of biochar and compost as an organic soil amendment for enhanced soil quality, fertility, and crop production; green cluster: co-application of biochar and compost in soils contaminated with heavy metals; blue cluster: the influence of biochar and compost on the microbial community; yellow cluster: the influence of biochar and compost on organic carbon, organic matter, and heavy metal content under certain pH conditions.A keyword analysis is a bibliometric method used to present development trends and research hotspots ; 92 of aTo help researchers learn the main research trends in the topic, it is important to understand the evolution process of keywords used over the past decades. Hence, the overlay visualization in 4.5.As an important form of co-authorship analysis, country co-authorship reflects the communication between countries, including the most influential countries. The co-authorship network and density visualization of countries in Interestingly, some European countries like Italy, France, Austria, Norway, and Switzerland were in the same cluster (green color), with Ghana, an African country. The density visualization map in 5.Hibiscus sabdariffa L.), and soil quality and reported a significant increase in soil microbial biomass carbon and N, increased N (16%), P (38%), and K (15%) phosphatase, and dehydrogenase activity compared to the control treatment with soil alone. Also, the combined application of biochar and compost in their study, increased the roselle shoot uptake of P, N, and K, by 31, 20, and 25%, including plant height, dry weight, fresh weight, chlorophyll content, etc., compared to the control treatment, and therefore, concluded that biochar and compost mixture increased soil and roselle quality. Using co-composted biochar under saline soil conditions to investigate its effect on soil quality and plant growth [Solanum melongena L.) growth, increased soil organic matter, N, P, K, water-holding capacity, field capacity, useful pores, available water, and several bacteria in the soil, including reduced bulk density, pH, and electrical conductivity (EC) at different co-composted biochar application rates , and irrigation regimes .Out of 16 subject categories, the mainstream research on the research topic was in Environmental science and Agricultural and Biological Sciences, and based on the analysis of all keywords (author and index keywords), the mainstream research was centered on biocharcompost, and co-composted biochar as a soil amendment to improve ‘soil fertility,’ ‘soil quality,’ and for remediation of soil contaminated with inorganic contaminants, with more recent research hotspot on ‘soil pollutants’ (e.g. lead and arsenic) . Investit growth , reporteZea mays), as a keyword it appeared more than 10 times in several studies on the topic [−1), under different light intensities and biochar-compost mixtures . The authors reported increased yield and vegetative growth of maize under reduced light intensity. These studies show that biochar and compost mixture is a promising soil organic amendment that can be exploited to produce maize under unfavorable and degraded soils.The most studied crop was maize  and concluded that biochar-based compost application to acid soils can enhance soil nutrient status, soil organic carbon, soil water content, crop yield, and can help reduce GHG emissions. Also, the effect of biochar and compost on peanut yield and greenhouse gas (GHG) was investigated in Australia on a Ferralsol [Another keyword that was linked to ‘soil,’ in the network visualization shown in erralsol . It was −1) and increased water extract organic carbon.Out of 62 countries, involved in biochar-compost research, and 67 co-authorship countries based on VOSviewer analysis, China was the most influential country on the topic as shown in 6.The properties of the Bibliometric analysis were employed in the present study to investigate current and research trends on the topic of biochar-based compost and co-composted biochar. From the analysis of the 254 publications retrieved from the Scopus database, the number of publications significantly increased between 2011 and 2021, with an anticipated increase in 2022. With China’s collaboration with the USA and other European countries being the top productive country, more collaborations, and funding with African countries, will also help increase the knowledge and benefits of using biochar-based compost and co-composted biochar in most degraded African soils.7.Like other bibliometric studies, the current bibliometric review has associated limitations. One database (i.e. Scopus) was used to retrieve data, in which a limited number of keywords were used. Therefore, future studies on the topic are recommended to explore different databases, and keywords to avoid missing important publications that are indexed in other databases.


With the development of high-power fuel cell vehicles, heat dissipation requirements have become increasingly stringent. Although conventional cooling techniques improve the heat dissipation capacity by increasing the fan rotating speed or radiator dimensions, high energy consumption and limited engine compartment space prevent their implementation. Moreover, the insufficient heat transfer capacity of existing coolants limits the enhancement of heat dissipation performance. Therefore, exploring novel coolants to replace traditional coolants is important. Nanofluids composed of nanoparticles and base liquids are promising alternatives, effectively improving the heat transfer capacity of the base liquid. However, challenges remain that prevent their use in fuel cell vehicles. These include issues regarding the nanofluid stability and cleaning, erosion and abrasion, thermal conductivity, and electrical conductivity. In this review, we summarize the nanofluid applications in oil-fueled, electric, and fuel cell vehicles. Subsequently, we provide a comprehensive literature review of the challenges and future research directions of nanofluids as coolants in fuel cell vehicles. This review demonstrates the potential of nanofluids as an alternative thermal management system that can facilitate transition toward a low-carbon, energy-secure economy. It will serve as a reference for researchers to focus on new areas that could drive the field forward. Nanofluids were first proposed by Professor Choi at Aragon National Laboratory, the United States of America in the early 1990s. They are composed of nanoparticles, metal, or polymer particles of sizes ranging from 1 nm to 100 nm and the base liquid. This is shown in The heat transfer enhancement mechanism of a nanofluid involves the addition of nanoparticles to the base fluid, increasing the thermal conductivity of the mixture and thereby improving the heat transfer capacity of the base fluid. In addition, particle-to-particle and wall-to-wall collisions and the mutual motion between the particles and liquid improve the heat transfer characteristics. However, this is a complicated process in microchannels. Molecular dynamics have been used to analyze the heat transfer mechanism at solid–liquid interfaces. Frank ,3 investNanofluids can be used as coolants in systems with high heat-dissipation requirements. For example, nanofluid applications have been investigated in food processing, solar energy collection, nuclear reactors, and oil recovery. In food processing, nanofluids can be used to increase heat exchanger efficiency and reduce energy consumption. Nanofluids can shorten the thermal processing time and improve the nutritional content of food ,12,13,14In 2004, the U.S. Department of Energy used nanofluids in fuel cells, and the heat dissipation capacity of cooling systems could be significantly improved . ParticuNanofluid application in oil-fueled vehicles’ cooling systemsNanofluids can be used to improve the heat-dissipation capacity of radiators in cooling systems. Many researchers have investigated the mechanism of heat transfer enhancement in radiator tubes using different nanofluids. 2O3, TiO2, CuO, SiO2, Fe3O4, Fe2O3, ZnO, SiC, GnP, CNC, MWCNT, CNT, and graphene have been analyzed in detail. In addition, the thermophysical properties and heat transfer performance of different types of nanofluids have been analyzed in the past. The influencing factors, including concentration, flow rate, temperature, and radiator dimensions, have been compared. Using a nanofluid instead of a conventional coolant has been demonstrated to improve the heat dissipation performance of thermal management systems [According to the literature content in  systems ,106,107.Nanofluid applications in oil-fueled vehicles’ air conditioner systems2/PAG nanofluid was selected as the nano-lubricant. A vehicle air conditioner test rig was used to measure the heat absorption and coefficient of performance (COP). The test results indicated that the maximum increase in the COP was 24% when using the SiO2/PAG nanofluid. Redhwan [2O3-SiO2/PAG nanofluids and compared the heat transfer performance and tribology enhancement. Zawawi [2O3-SiO2/PAG nano-lubricant and analyzed its effect on the performance improvement of vehicle air conditioner systems through experimental tests. The test bench is illustrated in Sharif  chose a  Redhwan  prepared. Zawawi  used an Nanofluid applications in oil-fueled vehicles’ lubrication systems2O3-TiO2/lube nanofluid. Lue [2O3/lube nanofluid. The densities and viscosities of the nano-lubricant were measured at various temperatures. The test results indicated that the 1.5 wt % Al2O3/lube nanofluid was the optimal choice for future research. Esfe [The main reason for the failure of lubricating oils at high temperatures is thermal degradation. Nanofluids can delay the oxidation and burnout of the lubricating oil, which results in a wider temperature range and higher fuel economy. Ali  used a tuid. Lue  used a tch. Esfe  proposedNanofluid applications in oil-fueled vehicles’ exhaust power generation systems2/water nanofluid using a two-step method to increase the efficiency of conversion units in thermoelectric generators (TEGs). The experimental results demonstrated that the TiO2/water nanofluid effectively enhanced the conversion efficiency and output power of the TEG. Karana [Researchers have investigated the possible utilization of waste heat in exhaust pipes because of the large amount of waste heat produced during engine operation. Based on the thermoelectric conversion unit, waste heat can be converted into electricity. For example, Hilmin  prepared. Karana  prepared2-CLPHP nanofluid as the coolant. The heating performances of different strategies were investigated. The test results indicated that the 2% TiO2/CLPHP nanofluid exhibited the best heat transfer performance. Wu [2O nanofluid with a lattice Boltzmann (LB) model to simulate natural convection in a battery. The problem of a uniformly distributed heat boundary was validated to ensure the accuracy of the model. The results proved that the Cu/H2O nanofluid could improve the cooling performance and reduce the temperature difference in a battery thermal management system. Abdelkareem [Batteries are the most important power source for electric vehicles. However, maintaining an ordinary operating temperature using a thermal management system is crucial. Nanofluids have been used as a coolant to replace existing coolants, and their performance in improving heat-dissipation capacity has been analyzed. For example, Chen  construcance. Wu  used a Celkareem  comparedelkareem  proposedelkareem  designedelkareem ,129,130.2/WEG nanofluid at 0.05–0.5%. The experimental results indicated that the thermal conductivity coefficient increased by 10% with the 0.5% TiO2/WEG nanofluid. Zakaria [2O3/WEG nanofluids in PEMFCs. The experimental results showed that the heat-dissipation power increased by 13.87% with a 0.5% Al2O3/WEG nanofluid. Zakaria [2O3/WEG nanofluids with different base fluid ratios. The test results indicated that the heat transfer performance of the 0.1% Al2O3/WEG nanofluid at a 60:40 ratio was better than that at a 50:50 ratio. Zakaria [2O3 nanofluids. The testing results showed that the thermal conductivity was reduced when the ethylene glycol proportion was increased. They also showed that the electrical conductivity was reduced when the ethylene glycol proportion was increased. In order to assess the significance of Al2O3 nanofluids in an electrically active thermal device, the thermoelectrical conductivity (TEC) ratio was established.In 2004, the U.S. Department of Energy first selected nanofluids as coolants for thermal management systems in fuel cell vehicles. The preparation cost of the nanofluid can be reduced and is close to that of the traditional coolants used in oil-fueled vehicles. Nanofluids can be used as coolants to reduce the radiator dimensions. They can also effectively improve the heat-dissipation capacity of cooling systems . Many re Zakaria  investig Zakaria  analyzed Zakaria  also anaThe aforementioned research studies were mainly based on metallic nanoparticles. Boron nitride (BN) is an insulating ceramic with high stability, high thermal conductivity, and good chemical inertness. For example, Ilhan  preparedThe thermal management system of a fuel cell vehicle (FCV) comprises heat dissipation and air conditioning systems. The heat-dissipation system includes heat-generating  and heat-dissipation  components, as shown in A proton-exchange membrane fuel cell (PEMFC) engine comprises an air supply system, control system, cooling system, fuel cell stack, and fuel supply system. Among these, the fuel cell stack is composed of a single PEMFC, which converts chemical into electrical energy. The cooling system of the PEMFC includes a radiator, fan, pump, intercooler, thermostat, and water tank. The gas temperature at the cathode inlet is adjusted using the coolant temperature and flow rate inside the PEMFC. The coolant can be circulated in a cooling loop, and heat is transferred from the interior of the fuel cell. Many researchers have investigated methods for optimizing the cooling techniques of heat-dissipation components to maintain the ordinary operating temperature of PEMFCs in fuel cell vehicles. As mentioned earlier, the heat-production components of fuel cell vehicles include fuel cells, power batteries, motors, motor controllers, air compressors, and DC/DC converters.Cooling techniques for fuel cellsAccording to the different output powers of fuel cells, their cooling methods can be classified as cathode air, separate airflow, phase change, and liquid cooling. The results are presented in The heat-dissipation performance can be improved by optimizing the internal flow channel structure and adjusting the fuel cell control strategy. The design optimization of fuel cells includes the use of flow channels, bipolar plates, and polymer film cooling channels. The structure of the flow channel includes serpentine, parallel, wave, and spiral flow channels, as shown in In addition, a reasonable control strategy can precisely control the rotational speed of the pump and fan of the cooling system under different operating conditions, ensuring that the fuel cell engines are in the normal operating temperature range to prevent overheating or overcooling. This is helpful in enhancing the operating efficiency of fuel cells ,145.Cooling techniques for batteries2.The cooling techniques for batteries include air and liquid cooling. Air cooling can be classified into natural or forced convection. The forced air-cooling technique has the advantages of a simple structure and low manufacturing cost. Moreover, it can increase the heat transfer coefficient on the air side and improve the heat-dissipation capacity relative to natural convection . Figure Currently, many researchers have investigated the effect of different air-cooling structures and arrangements on cooling performance using theoretical analyses and numerical simulations. For example, the effect of a conical structure on the cooling performance of a battery was investigated. Considering the influencing factors, including the discharge rate, fin arrangement, inlet air temperature, and inlet air flow rate, a prismatic air-cooled module and an aluminum-finned radiator were designed ,149,150.Cooling techniques for other componentsCooling techniques for the motor and motor controllers include natural, liquid, air, and oil cooling. The cost of air cooling is lower than that of other cooling techniques. However, the heat-dissipation capacity is not as high as that of liquid cooling. The cooling techniques for air compressors include air and liquid cooling. The former cooling method can be improved by optimizing the structure of the air compressor. This cooling method can be improved by enhancing the thermophysical properties of the coolant. Liquid cooling has a better heat-dissipation performance than air cooling. Liquid cooling is also used for DC/DC converters; the cooling can be improved by enhancing the thermophysical properties of the coolant or by improving the structure of the DC/DC converter. The heat-dissipation components of thermal management systems include radiators, pumps, and fans. For the radiator, heat-dissipation-capacity improvement is mainly achieved by structural and material optimization, where the structure includes fin and flat tube structures ,166,167.Pumps and fans are the main heat-dissipation components of thermal management systems, and they can be improved by optimizing the structure and control strategy. For example, Liu  used a cIn summary, the radiator, pump, and fan were improved via structural optimization. Structural optimization is more complex and expensive than improving the thermophysical properties of coolants. According to the above literature, there is considerable potential for investigating nanofluids as coolants to improve the heat-dissipation performance of thermal management systems in fuel cell vehicles.The thermophysical properties of nanofluids, including density, specific heat, and thermal conductivity, have been extensively investigated. The equations for the thermophysical properties are as follows .Nanofluid density can be calculated as shown in Equation (1):nfC is the nanofluid specific heat, bfC is the base fluid specific heat, pC is the nanoparticle specific heat, and φ is the nanofluid volume fraction.Nanofluid specific heat can be calculated as shown in Equation (2):nfk is the nanofluid thermal conductivity, bfk is the base fluid thermal conductivity, pk is the nanoparticle thermal conductivity, φ is the nanofluid volume fraction, and n is the shape factor.Nanofluid thermal conductivity was calculated by Hamilton & Crosser as shown in the following Equation (3):2O3, TiO2, and BN nanoparticles can be added into the base fluid to prepare a new coolant for fuel cell vehicles. The thermophysical parameters of four nanofluids can be seen in According to the literature above, ZnO, Al2 nanofluid > Al2O3 nanofluid > BN nanofluid. The specific heat of the nanofluid decreases as the volume fraction increases. The specific heat of BN nanofluid > Al2O3 nanofluid > TiO2 nanofluid > ZnO nanofluid at the same volume fraction. The nanofluid thermal conductivity also increases as the volume fraction increases. At the same volume fraction, the thermal conductivity of BN nanofluid > ZnO nanofluid > Al2O3 nanofluid > TiO2 nanofluid. The calculation results are presented in nfK is the effective thermal conductivity of the solid–liquid suspension, pK is the thermal conductivity of the nanoparticles, bfK is the thermal conductivity of the base fluid, and φ is the volume fraction of the nanoparticles.Thermal conductivity is one of the most important thermophysical properties used to calculate the heat-dissipation power of a nanofluid. It is affected by different factors, including the nanoparticles’ thermal conductivity, concentration (volume fraction or mass fraction), shape, and size, as well as the base fluid’s thermal conductivity. Several classical thermal conductivity models have been considered. For example, the Maxwell model, which is a thermal conductivity model for solid–liquid and two-phase coolants, was the first proposed. It is a static model for determining the thermal conductivity of monodisperse, low-volume-fraction, and spherical nanoparticles, as shown in Equation (4) . Subsequpk is the thermal conductivity of the nanoparticles, φ is the volume fraction of the nanofluid, and bk is the thermal conductivity of the base fluid.Bruggeman  proposedn is the shape factor:Ψ is the sphericity degree, which is the ratio between a sphere’s surface area and a particle’s surface area at the same volume.The Hamilton and Crosser model is extended from the Maxwell model by introducing sphericity, which allows the calculation of the thermal conductivity of solid–liquid suspensions formed by spherical or non-spherical nanoparticles . This mon − 1) factor with the parameter k in the equation [bfk is the base fluid thermal conductivity, φ is the nanofluid volume fraction, and pk is the thermal conductivity of the nanoparticles.Yamada and Ota modified the Hamilton and Crosser model by replacing the k is the thermal conductivity within the adsorption layer, bfk is the base fluid thermal conductivity, pr is the nanoparticle size, t is the adsorption layer thickness, pk is the nanoparticle thermal conductivity, and lk is average thermal conductivity of the nanofluid adsorption layer.The thermal conductivity curve for the adsorption layer includes linear and exponential distribution curves. The model for the linear distribution law of the thermal conductivity within the adsorption layer is shown in Equation (14):However, there is no unified mathematical equation for calculating the thermal conductivity of nanofluids in fuel cell vehicles. The adsorption layer theory must be used to calculate the nanofluid thermal conductivity more accurately.2O3 and MWCNTs using experimental analysis. The results showed that heat transfer coefficient of the hybrid nanofluid was slightly higher than that of the Al2O3 nanofluid at the same flow velocity. The hybrid nanofluid had the highest heat transfer coefficient at a given pumping power. Arif [2O3 nanofluid, a CNT nanofluid, and a graphene nanofluid. The analysis results indicated that heat transfer rate of the hybrid nanofluid was increased by 33.67%. Tlili [2O3 nanoparticles and C2H6O2 as the base fluid and analyzed the heat transfer performance of the hybrid nanofluid. Devi [2O3 nanofluid. The results proved that the heat transfer rate of the Cu/Al2O3 nanofluid was higher than that of the Cu nanofluid under a magnetic field environment. According to 2O3-TiO2, TiO2-SiO2, MWCNT-SiO2, Al2O3-CuO, Fe3O4-CQD, CuO-CQD, Al2O3-Ag, Al2O3-Cu, Al2O3-SiC, etc. However, current research on hybrid nanofluid applications in PEFMCs only regard the Al2O3-SiO2 hybrid nanofluid. For example, Johari [2O3-SiO2 hybrid nanofluid with different percentages and analyzed its heat-dissipation capacity using an experimental method. The results proved that the thermal conductivity of the Al2O3-SiO2 nanofluid was higher than that of a single nanofluid. Khalid [2O3-SiO2 nanofluids. The experimental results showed that the thermal and electrical conductivity of the Al2O3-SiO2 nanofluid decreased as the percentage of Al2O3 increased. Meanwhile, the viscosity of the Al2O3-SiO2 nanofluid increased. The test results indicated that the optimal percentage of Al2O3-SiO2 nanofluid as a coolant for PEMFC was 10:90. In recent years, two or more nanoparticles have been mixed to prepare multi-mixed nanofluids. Subsequently, the heat-dissipation performances of PEMFCs were investigated. For example, Huang  investiger. Arif  investig%. Tlili  preparedid. Devi  used a n, Johari  prepared. Khalid  preparedIn addition, many researchers have investigated different types of hybrid nanofluids in electric vehicles ,184,185.Research on nanofluid applications in fuel cell vehicles has been focused on the heat-dissipation performances of PEFMCs. Although current research has proven that nanofluids can effectively enhance the heat-dissipation capacity of thermal management systems, the stability problem is inevitably considered when nanofluids are applied in fuel cell vehicles. In addition, nanofluid cleaning is a significant challenge, as nanofluid coolants must be replaced at the end-of-life cycle because nanoparticle sediments accumulate in microchannels and corners.Nanofluid stabilityAccording to the theory of electrostatic stabilization (Derjaguin Landau Vewey Overbeek), if the van der Waals attraction force is greater than the electrostatic repulsion force, the nanoparticles will collide with each other. This causes the suspended nanoparticles to agglomerate and form clusters. The nanoparticles gradually settle under the influence of gravity. They even block the microchannels and affect the thermal conductivity and viscosity of the nanofluid. From a macroscopic perspective, different influencing factors, such as temperature, concentration, pH, and particle size, lead to destabilization and precipitate formation. Hence, it is necessary to analyze the nanofluid stability from macroscopic and microscopic perspectives.Nanofluid preparation includes the one-step method and the two-step method. The one-step method involves incorporating nanoparticles directly into the base fluid without intermediate steps, such as nanoparticle drying, storage, and dispersion. This method produces nanofluids with good stability; however, it is only suitable for preparing small quantities of nanofluids because of its high cost. The two-step method refers to the preprocessing of the nanoparticle, which is followed by the mixing of the nanoparticle and the base fluid. This method is suitable for preparing large quantities of nanofluids at a low cost. The stability of this method is not as good as that of the one-step method, and a dispersant is required to maintain good stability . The preTo solve the nanofluid settlement problem, many researchers have used auxiliary dispersion techniques to improve nanofluid stability. These techniques include physical and chemical dispersions. Physical dispersion involves ball milling, ultrasonic vibration, and magnetic stirring. Ball-milling dispersion refers to the use of planetary ball milling to reduce the average number of clusters. Ultrasonic vibration includes water bath ultrasonic vibration and probe ultrasonic vibration. The latter method is more effective in breaking particle clusters and reducing the average cluster size. A magnetic stirrer can prevent bubble generation during nanofluid preparation by adjusting the stirring speed, which keeps the nanofluid stable for a short time. Chemical dispersion involves the addition of surfactants and pH adjustment. Surfactants can be classified as ionic, nonionic, amphoteric, and complex. The dispersion results are presented in 2O3 nanofluid, TiO2 nanofluid, and Al2O3-SiO2 nanofluid. The stability of these nanofluids have been investigated by some researchers. For example, Choudhary [2O3 nanofluid’s stability. If the zeta potential was higher, the stability of the Al2O3 nanofluid was better. Witharana [2 nanofluid. In addition, the effects of sunlight on the stability of the TiO2 nanofluid were assessed. Sunlight caused the TiO2 nanofluid’s size to increase up to 45% in three days. The sedimentation velocity is a key parameter to evaluate the stability of a TiO2 nanofluid. Adil [According to the summarization in houdhary  used zetitharana  also useid. Adil  investigAddressing the stability problems of nanofluids is a fundamental requirement for their application in fuel cell vehicles. Although many researchers have elucidated the specific factors affecting nanofluid stability, it is necessary to reveal the hidden laws of these factors. A schematic diagram of nanofluid destabilization and sedimentation is shown in The main factors influencing the stability of nanofluids include the temperature, concentration, pH, and particle size. (1) Temperature: When the temperature changes, the dielectric constant of the base fluid also changes . (2) Concentration: The average separation distance of the nanoparticle decreases as the concentration increases. The van der Waals attraction also increases. If the Van der Waals attraction is greater than the electrostatic repulsion, the nanoparticles will agglomerate. Hence, a higher nanofluid concentration implies easier settling. (3) The pH affects the charge density on the nanoparticle surface, decreasing the zeta potential. (4) Particle size: This affects the van der Waals attraction and electric double-layer repulsion. When the nanoparticle size is small, the number of atoms on the nanoparticle’s surface is higher. The density increases in the active position. The tendency for aggregation is enhanced, indicating that the nanofluid settles more easily.Nanofluid cleaningAccording to the literature, the coolant must be replaced every four years in electric vehicles . BecauseHowever, the problem of nanofluid stability is the greatest challenge that prevents its use in fuel cell vehicles. If nanoparticles accumulate in corners, such as in the flat tubes of a radiator, it is difficult to completely clean them. This affects the thermophysical properties of nanofluid coolants.Nanofluids are composed of nanoparticles  and base fluids . During circulation in the cooling loop, the nanoparticles impact the surfaces of the microchannels. In the long term, nanoparticles may wear the microchannels, particularly those in the radiator flat tube and the flow channel of PEMFCs, as shown in Although many researchers have focused on the heat transfer performance of nanofluids, the erosion and abrasion effects of nanofluids on microchannel surfaces have been poorly reported. Eneren  analyzed2O3/WEG nanofluid as a coolant. The porosity of the nanofluid increased, causing the pumping power to increase. Pandey [The viscosity of the nanofluid increases as the nanofluid concentration increases. This causes the pumping power to exponentially increase. Hence, the nanofluid concentration should not infinitely increase. Karami  investig. Pandey  tested t. Pandey ,204. The2O3, ZnO, and TiO2 are metal oxides with electrical conductivity. According to the coolant requirement for PEMFCs, the electrical conductivity should be less than 5 μs/cm. Many researchers have investigated the electrical conductivities of Al2O3/WEG, ZnO/WEG, and TiO2/WEG nanofluids. For example, Khdher [2O3/bio-glycol nanofluid and measured the electrical conductivity using a measurement apparatus. The test results indicated that adding nanoparticles into the base fluid could increase the electrical conductivity to 53–154 μs/cm at 30–80 °C. Subramaniyan [2/W, TiO2/EG, and TiO2/PG. The research results showed that the TiO2/W nanofluids had higher dielectric constants than EG and PG at the same temperature and frequency, which could be adjusted by changing the volume fraction of the nanofluid. Chilambarasan [A nanofluid is a mixture of nanoparticles and a base liquid. However, nanoparticles that include Al, Khdher  investigamaniyan  analyzedmbarasan  selected2O3-SiO2/bio-glycol (BG) hybrid nanofluid as a coolant for a PEMFC. Compared to conventional ethylene glycol, green bio-glycol (BG) exhibited a better performance than the base fluid. The thermal conductivity also increased. Consequently, the electrical conductivity decreased. This is shown in 2 nanofluid had the highest electrical conductivity at 268.8 μs/cm, but the single Al2O3 nanofluid’s electrical conductivity was 227.3 μs/cm. The Al2O3-SiO2 hybrid nanofluid’s electrical conductivity was lower than that of the single nanofluid. The electrical conductivity of the other mixture ratio of the Al2O3-SiO2 hybrid nanofluid was approximate to that of the single Al2O3 nanofluid.Regarding hybrid nanofluids, Johari  investigBased on the literature review above, the challenges of nanofluid application in fuel cell vehicles include: (1) nanofluid stability (how to maintain low-term dispersibility of nanoparticles in the base fluid), (2) nanofluid cleaning (how to clean up remaining nanoparticles in microchannels), (3) erosion and abrasion by nanofluids (how to eliminate or reduce erosion and abrasion), (4) nanofluid pump transport power , and (5) nanofluid electrical conductivity .Solving nanofluid destabilization and sedimentation problems is a basic requirement for nanofluids’ use in fuel cell vehicles. It is necessary to investigate and analyze the factors influencing nanofluid stability, including the temperature, concentration, pH value, and particle size. However, previous research has only focused on one or two factors, which was difficult to summarize. The nanofluid preparation process includes physical and chemical dispersion techniques, such as ball milling, ultrasonic vibration, magnetic stirring, addition of a surfactant, and adjustment of the pH value. These auxiliary dispersion techniques can effectively improve a nanofluid’s stability. However, it is difficult to summarize the principle of the auxiliary dispersion technique.2O3-TiO2, TiO2-SiO2, MWCNT-SiO2, CuO-CNT-graphene, Al2O3-CNT-graphene, Al2O3-CuO, Fe3O4-CQD, CuO-CQD, Al2O3-Ag, Al2O3-Cu, Al2O3-SiC, and Al2O3-CuO, among others. Current research has verified that hybrid nanofluids have better heat transfer capacities than single nanofluids. However, in fuel cell vehicles, only a few researchers have explored the heat-dissipation performance of an Al2O3-SiO2/WEG nanofluid as a coolant. Hence, it would be of considerable scientific value to study more types of hybrid nanofluids for fuel cell vehicles.For oil-fueled vehicles and electric vehicles, researchers have investigated different types of hybrid nanoparticles, such as GnP-CNC, AlDuring the circulation of a nanofluid in a cooling system, the nanoparticles impact and wear the surface of the microchannels, which is particularly true for the flat tube and flow channel of PEMFCs. The characteristics of PEMFCs and radiators are affected by long-term wear. However, few researchers have investigated erosion and abrasion of the surfaces of flat radiator tubes and microchannels inside PEMFCs caused by nanofluids. It is important to analyze the impact phenomenon using numerous experiments and observations from macroscopic and microscopic perspectives. Finally, the nanofluid with the lowest impact on erosion and abrasion must be selected for comparison and analysis.The thermal conductivity of a nanofluid is an important thermophysical property. Many researchers have proposed classic thermal conductivity models, such as the Maxwell, Braggeman, Wasp, Hamilton and Crosser, and Yamada and Ota models. There is no unified theoretical model to accurately calculate nanofluid thermal conductivity. A few researchers have proposed a theory on thermal conductivity of the adsorption layer, which refers to the liquid layer formed around solid nanoparticles between the base fluid and the nanoparticles. This reflects the internal heat transfer mechanism of the nanofluid more accurately. Hence, it would be of great scientific value to explore the thermal conductivity of nanofluids using the adsorption layer theory.2O3/WEG, ZnO/WEG, and TiO2/WEG, which are metal oxides with electrical conductivity. Based on the requirements for coolants for PEMFCs, the electrical conductivity should be less than 5 μs/cm. When nanofluids flow through a PEMFC microchannel, they inevitably contact the inner surface of the BP. If a nanofluid has a high electrical conductivity, it can cause electric leakage inside the PEMFC, affecting the output power of PEMFC. Hence, exploring nanofluids with low electrical conductivities in the future is necessary.According to current research, nanofluid applications in PEMFC include AlWith the development of high-power fuel cell stacks, heat-dissipation requirements have become more stringent. Reinforcing the heat transfer performance of the coolant is simpler, has a lower cost, and increases the rotation speed than structural optimization. Nanofluids are a type of heat-dissipation-enhancement coolants that have been applied in different industrial fields. This study summarized the challenges and research directions of nanofluids as coolants in fuel cell vehicles. The main conclusions of this study are as follows.(1) Nanofluids have great potential as coolants in oil-fueled, pure electric, and fuel cell vehicles. Many researchers have verified this and have highlighted their disadvantages. Nanofluids can be used as coolants, refrigerating fluids, and heat transfer media in oil-fueled vehicle cooling systems, air conditioner systems, and exhaust power generation systems. In pure electric vehicles, nanofluids can be considered as coolants in battery-cooling systems. However, for fuel cell vehicles, the current research has only focused on the heat-dissipation performances of nanofluids in PEMFCs.(2) The thermal management system of an FCV comprises heat dissipation and air conditioning systems. Heat-dissipation systems can be classified into heat-generation and heat-dissipation systems. Based on the analyses and comparisons performed in numerous studies, the heat transfer capacity of a coolant is better than that of the heat-dissipation system. This further proves that nanofluids have considerable potential in replacing existing coolants.2O3/WEG, TiO2/WEG, and BN/WEG nanofluids were analyzed as coolants in PEMFCs. In this study, four types of nanofluids were selected along with their density, specific heat, thermal conductivity, and convective heat transfer coefficient. Moreover, this study mainly analyzed the thermal conductivity of the nanofluid and the Al2O3-SiO2/WEG nanofluid.(3) The thermophysical properties of nanofluids, including the density, specific heat, thermal conductivity, and convective heat transfer coefficient, have been extensively studies. The equations for the thermophysical properties have been analyzed in detail. ZnO/WEG, Al(4) According to a literature review, there are some challenges in using nanofluids as coolants in fuel cell vehicles, including nanofluid stability, cleaning, erosion and abrasion, pump transport power, and electrical conductivity. Among these, nanofluid stability is the biggest obstacle that prevents its use in fuel cell vehicles. Hence, it is necessary to improve the nanofluid preparation methods, which affect the stability of the nanofluid, from macroscopic and microscopic perspectives.(5) Finally, this study proposed five research directions for using nanofluids as coolants in fuel cell vehicles in the future. They include nanofluid stability improvements, hybrid nanofluid applications, reductions in erosion and abrasion by nanofluids, a nanofluid thermal conductivity model, and nanofluid electrical conductivity.


Zea mays L. It was found that herbicides in contaminating amounts stimulated the proliferation of organotrophic bacteria and inhibited the growth of fungi. Organotrophic bacteria and actinobacteria were represented by K-strategies and fungi by r-strategies. Bacteria belonging to the phylum, Actinobacteriota, represented by the genus, Cellulosimicrobium, were most abundant in the soil, while among the fungi, it was the phylum, Ascomycota, represented by the genus, Humicola and Chaetomium. The herbicides decreased urease activity while increasing arylsulfatase and acid phosphatase activity. They had a positive effect on the growth and development of Zea mays L., as evidenced by an increase in the values of the plant tolerance index (TI) and the maize leaf greenness index (SPAD). The results indicate that soil microorganisms and enzymes are suitable indicators reflecting the quality of herbicide-treated soil.Soil’s biological equilibrium, disturbed by the uncontrolled penetration of pesticides, can be restored by the activity of native microorganisms, which show abilities in neutralizing these xenobiotics. Therefore, this research is necessary in the search for new microorganisms used in the process of the bioremediation of contaminated soils. The aim of this study was to evaluate the effects of the herbicides, Sulcogan 300 SC, Tezosar 500 SC, and Sulcotrek 500 SC, applied to soil at the manufacturers’ recommended dosage as well as 10-fold higher, on the abundance of microorganisms, the diversity and structure of bacterial and fungal communities, the activity of soil enzymes, and the growth and development of  These methods involve, in particular, the use of microorganisms that convert harmful xenobiotics into non-toxic compounds . A safe The health of soil primarily entails its ability to promote plant growth and development and animal health, and to maintain or improve water and air quality . The heaAlcaligenes, Cellulosimicrobium, Microbacterium, Micrococcus, Methanospirillum, Aeromonas, Sphingobium, Flavobacterium, Rhodococcus, Aspergillus, Penecillium, Trichoderma, Streptomyces, Rhodotorula, Candida, and Aureobasidium genera [Opd (Pseudomonas diminuta), opaA (Alteromonas spp.), opdA (Alteromonas radiobacter), adpB (Nocardia spp.), pepA (Escherichia coli), hocA (Pseudomonas monteilli), pehA (Burkholderia caryophilli), Phn (Bacillus cereus), ophB (Burkholderia spp. JBA3), ophC2 (Stenotrophomonas spp. SMSP-1), OpdB (Lactobacillus brevis), Imh (Arthrobacter spp. scl-2), Mpd , Oph (Arthrobacter spp.), Mph (Arthrobacter spp. L1), MpdB (Burkholderia cepacia), opdE (Enterobacter spp.), A-opd (Aspergillus niger), P-opd (Penicillium lilacinum) [Bacillus, for pesticide biodegradation, due to the production of many extracellular enzymes , which exhibit a broad range of effects on the breakdown of many pesticide compounds [Pseudomonas genus bacteria, due to their rapid adaptation in a pesticide-contaminated environment. In addition, these bacteria secrete enzymes involved in the conversion of these chemical compounds, i.e., oxidoreductases, dioxins, cytochrome P450, carboxylases, and phosphotriesterases [β-glucosidase, arylsulfatase, and urease), and phytotoxic parameters .In soils, pesticides are degraded primarily through co-metabolism, as well as hydrolysis and photolysis. During the co-metabolism process, microorganisms use the active substances of pesticides as a source of carbon or energy, and also as a second nutrient that provides them with energy. This process is aided by many enzymes secreted by microorganisms . In turnm genera ,21,22. Tlacinum) . In turnlacinum)  have empompounds . High desterases . Mollea sterases  have clasterases . MicroorThe study has shown that the tested herbicides stimulated the growth of organotrophic bacteria, regardless of the dose used. Tezosar 500 SC applied to the soil in the RD dose caused an increase in the number of fungi, whereas Sulcogan 300 SC and Sulcotrek 500 SC, the herbicides used in the 10RD dose, reduced their number. The number of actinobacteria increased under the influence of Sulcogan 300 SC and Tezosar 500 SC administered into the soil in the RD dose and under the influence of Sulcotrek 500 SC applied in the 10RD dose, but decreased in the soil treated with Tezosar 500 SC in the 10 RD dose and Sulcotrek 500 SC in the RD dose .The significant effect of herbicides on the microbial abundance was confirmed by the values of the IF index . The stuThe colony development index (CD) presented in Herbicides also significantly affected the values of the ecophysiological diversity (EP) index of microorganisms . In the Actinobacteriota, (accounting for 43.37% to 50.01%) and Proteobacteria (accounting for 24.64% to 31.43%), with the population number of Actinobacteriota being significantly higher than that of Proteobacteria. The abundance of Actinobacteriota and Proteobacteria in the herbicide-treated soils was significantly greater than in the control soil, with the highest number of these bacteria observed in the soil treated with the Sulcogan 300 SC herbicide. The bacteria belonging to the remaining phyla were found in the soil in much smaller numbers. In PCA, they formed a homogeneous group composed of Acidobacteriota, Gemmatimonadota, Chloroflexi, Bacteroidota, Firmicutes, Myxococcota, Patescibacteria, Planctomycota, and Verrucomicrobiota bacteria.The data presented in Actinobacteria, belonging to the phylum, Actinobacteriota. Their OTU number was the highest in the soil treated with Sulcogan 300 SC herbicide  and the lowest in the control soil . In these soil types, there were also numerous Alphaproteobacteria and Gammaproteobacteria, bacteria belonging to the phylum, Proteobacteria. Their OTU number was highest in soils with the addition of Sulcogan 300 SC and reached 10,706 OTU and 9576 OTU, respectively  to 22,458 OTU (soil treated with Sul). In addition, the soil was densely colonized by bacteria belonging to the orders, Frankiales, Propionibacteriales and Gaiellales (class Actinobacteria), Sphingomonadales and Rhizobiales (class Alphaproteobacteria), Burkholderiales (class Betaproteobacteria), Xanthomonadales (class Gammaproteobacteria), and Gemmatimonadales (class Gmmatimonadetes).Dominant bacterial orders are shown in Promicromonosporaceae family of bacteria was found to predominate (accounting for 26.83% to 38.85%). In the soil treated with the Sulcogan 300 SC herbicide, their number increased by 3.67% compared to the control soil, whereas in the soil with the addition of Tezosar 500 SC and Sulcotrek 500 SC, their number decreased by 6.69% and 8.35%, respectively. The soil was also heavily populated by bacteria belonging to the Sphingomonadaceae family, with their abundance increasing under the influence of the Sulcogan 300 SC herbicide by 1.42% compared to the control soil, and decreasing by 1.08% and 2.61% in the soil treated with the Tezosar 500 SC and Sulcotrek 500 SC herbicides, respectively. The Sulcogan 300 SC and Tezosar 500 SC herbicides inhibited the growth of Gemmatimonadaceae, Acidothermaceae, and Solibacteraceae, while Tezosar 500 SC and Sulcotrek 500 SC stimulated the multiplication of Nocardioidaceae, Acetobacteraceae, Solirubrobacteraceae, and Haliangiaceae bacteria. In addition, all the tested herbicides contributed to the increasing counts of Burkholderiaceae, Comamonadaceae, and Gemmataceae bacteria and the decreasing counts of Frankiaceae  to 18,540 OTU (soil treated with Sul). The soil was also heavily populated by the Sphingomonas genus bacteria, the high population number of which (reaching 4493 OTU) was observed in the soil treated with Sulcogan 300 SC. The application of Tezosar 500 SC and Sulcotrek 500 SC to the soil reduced the counts of Mucilaginibacter and Lysobacter genus bacteria, and increased that of the Pseudarthrobacter and Candidatus_Solibacter bacteria. In turn, all tested herbicides increased the abundance of the Acidothermus, Gemmatimonas, Nocardioides, Ralstonia, Bradyrhizobium, Terrabacter, Burkholderia-Caballeronia-Paraburkholderia, Sphingobium, and Haliangium genus bacteria , followed by Mortiellomycota (accounting for 9.05% to 16.27%) and Basidiomycota (accounting for 2.55% to 6.36%). Ascomycota were the most abundant in the soil treated with Tezosar 500 SC, whereas Mortiellomycota was the most common in the soil treated with Sulcotrek 500 SC. The relative abundance of fungi belonging to the phylum, Basidiomycota, decreased significantly in the herbicide-treated soil compared to the control soil  and Leotiomycetes class fungi  in the soil. The remaining fungal classes at OTU ≥ 1%  occurred in the soil in slightly smaller numbers  A great abundance of the Thelebolales order fungi was also found in the soil treated with the Tezosar 500 SC herbicide .The most abundant fungi found in the analyzed soil samples were those belonging to the order, dariales , with thChaetomiaceae family. (Their numbers ranged from 67.49% to 56.05%.) Their abundance increased by 8.13% in the soil treated with Sulcogan 300 SC and decreased by 3.31% in the soil treated with Tezosar 500 SC. In the herbicide-treated soil, there was also a significant increase in the number of Pseudeurotiaceae fungi (by 15.67%) compared to the control soil. The abundance of Mortierellaceae fungi increased by 4.24% upon soil treatment with Sulcotrek 500 SC, and decreased in the soils treated with Sulcogan 300 SC and Tezosar 500 SC by 2.05% and 4.96%, respectively. All the herbicides tested reduced the abundance of the Didymosphaeriaceae, Orbiliaceae, Sporocadaceae, and Bulleribasidiaceae family fungi.The application of herbicides into the soil resulted in changes in the structure of fungal communities at the family level . The soiHumicola and Chaetomium genera, belonging to the Chaetomiaceae family, and slightly less by the Morierella genus fungi, belonging to the Mortierellaceae family  and that of Chaetomium fungi in the soil amended with Tezosar 500 SC . The soil treated with Tezosar 500 SC was also densely colonized by the Pseudogymnoascus genus fungi .The soil samples were most heavily populated by the fungi from the e family . The greHumicola, Chaetomium, Mortierella, Vishniacozyma, Pseudogymnoascus, Penicillium, Trichocladium, Fusarium, Exophiala, and Fusicolla. In addition, the soils containing the Sulcogan 300 SC, Tezosar 500 SC, and Sulcotrek 500 SC herbicides were colonized by the fungi of the genus Trichoderma  and Tezosar 500 SC applied in the 10RD dose (IF = −0.070).Dehydrogenases and urease turned out to be the most sensitive to the tested preparations, as evidenced by the negative values of their IFs. The IF values determined for dehydrogenases ranged from −0.079 to −0.155, whereas those computed for urease ranged from −0.233 to −0.510. The most negative effect on urease activity was caused by Sulcotrek 500 SC and on the dehydrogenases, by Sulcogan 300 SC. All IF values computed for acid phosphatase and arylsulfatase were positive, which may indicate a stimulating effect of the herbicides on their activity. The IF values computed for acid phosphatase were in the range of 0.078 to 0.298 and those computed for arylsulfatase, in the range of 0.104 to 0.486. The most beneficial effect on the activity of acid phosphatase was also exerted by Tezosar 500 SC, while on the activity of arylsulfatase, it was by Sulcogan 300 SC. It was observed that catalase responded negatively to Sulcogan 300 SC and Sulcotrek 500 SC applied to the soil in the 10RD dose ; alkaline phosphatase, to Sulcogan 300 SC added in the RD dose (IF = −0.044), and 10RD dose (IF = −0.220) and to Sulcotrek 500 SC applied in the RD dose (IF = −0.073); and Zea mays L. , Gammaproteobacteria , and Actinobacteria (represented by Rhodococcus wratislaviensis FPA 1). Petric et al. [Actinobacteriota and Bacteroidota phyla, while Romdhane et al. [Proteobacteria and Acidobacteriota to be the prevailing phyla. The results obtained in the present research confirm findings from previous studies. They show that the analyzed soil was most heavily populated by bacteria belonging to the Actinobacteriota phylum, with their numbers being significantly reduced under the influence of the Sulcotrek 500 SC herbicide. The response of these bacteria to the herbicides tested may be due to the greater stability of the mixture of its active substances and thus to their lesser availability to bacteria capable of degrading these substances [Proteobacteria, whose relative abundance increased significantly after soil treatment with the Sulcogan 300 SC herbicide. In order to survive under adverse conditions, these bacteria could produce new enzymes and undergo a number of genetic mutations, which allowed them to adapt to new conditions and modified environment [Cellulosimicrobium genus, which are probably capable of degrading these preparations. Additionally, bacteria from of the Sphingomonas and Gemmatimonas genera may exhibit high resistance to the tested herbicides, because their numbers were significantly higher in the soil treated with these preparations than in the control soil. This may indicate their adaptation to the new conditions changing in the soil environment under the influence of herbicides. According to Tonello et al. [Aspergillus oryzae and Penicillium brevicompactum, belonging to the class, Eurotiomycetes, and Lentimula edode, belonging to the class, Agaricomycetes. In the present research, the soil was mainly colonized by the fungi from the Humicola and Chaetomium genera, belonging to the Sordariomycetes class and by Mortierella, belonging to the Mortierellomycetes class. The response of the above-mentioned fungal genera varied depending on the herbicide used. The development of the Humicola genus fungi was inhibited by Sulcogan 500 SC and Tezosar 500 SC, whereas their growth was stimulated by Sulcotrek 500 SC. The soil amendment with Tezosar 500 SC had a positive effect on the development of Chaetomim, and soil treatment with Sulcotrek 500 SC, on the development of Mortierella genus fungi. An increase observed in the number of fungi under the influence of herbicides may indicate their greater resistance to these preparations and their degradation potential. The biodegradation of herbicides by fungi is based on an oxidative-hydrolytic mechanism, during which they modify the molecule by sequentially removing the aromatic ring substituent, with dealkylation as the initial stage of herbicide degradation [Mortierella genus was reduced under the influence of Sulcogan 300 SC and Tezosar 500 SC, while the number of the Chaetomium genus fungi, after Sulcotrek 500 SC application. The inhibition of fungal growth may have been due to protein overexpression associated with increased energy demand, changes in the cell wall and cytoskeleton, as well as a response to oxidative stress induced by herbicides [Dong et al.  have empegraders . In turne amines . The cone amines ,38. Accoe amines , the mosc et al.  and Merlc et al.  reportedbstances . Anotherironment . Regardlo et al. , the mosradation . It has rbicides .The use of herbicides may disturb not only the populations and microbial diversity of the soil, but also its enzymatic activity. The activity of certain enzymes can be inhibited by herbicides applied to the soil, while that of others may be stimulated. These changes may occur depending on the herbicide type and dose, as well as the duration of the deposition of these xenobiotics in the soil . Accordi−1 dose) on maize, indicated by a decrease in the dry weight of plants, the length of their shoots and roots, and chlorophyll content. The reduction in chlorophyll content in the plant by herbicides may be caused by blocking the transport of electrons in the photosynthesis process. However, in the present study, the herbicides did not impair the photosynthesis, as we noted their positive effect on the greenness index of the maize leaves, especially at the BBCH 19 growth stage. Such a positive response of the maize to the herbicides applied may be due to the secretion of antioxidant enzymes by the plants, i.e., peroxidase or glutathione s-transferase, which are some of the most important metabolic enzymes in the plant responsible for the transformation of pesticides [Maize is one of the most important crops in the world, and its growth may be inhibited by competing weeds. Its proper yielding may be ensured, foremost, by the use of herbicides that effectively eliminate weeds ,48. Accosticides ,56.Cellulosimicrobium and Nocardioides bacteria belonging to the phylum, Actinomycetota; Sphingomonas and Ralstonia, belonging to the phylum, Proteobacteria; and Gemmatimonas, belonging to the phylum, Gemmatimonadota, which were identified in all soil samples. These are soil bacteria that promote plant growth and development and live in close association with plants. These microorganisms also show the ability to combat plant pathogens, induce systemic plant resistance to disease, and have antagonistic effects on other microorganisms [Microorganisms have a great impact on the disappearance of herbicides in the soil, as they degrade organic compounds present in the environment they inhabit as a result of their metabolic properties. Their ability to metabolize these xenobiotics is related to the activity of structural enzymes and induced enzymes, which are produced by microorganism cells. This response of microorganisms to herbicides present in the soil is related to their adaptive mechanism, which allows them to survive in changing environmental conditions. In addition, microorganisms use herbicides as a source of carbon and energy during the decomposition of herbicides, which makes them multiply intensively . Such abrganisms . Plants,rganisms  and selerganisms  through rganisms . In our −1 d.m. of soil): nitrogen (in the form of CO(NH2)2)—130 mg, phosphorus (in the form of KH2PO4)—100 mg, potassium (in the form of KH2PO4 + KCl)—130 mg, and magnesium (in the form of MgSO4 × 7H2O)—25 mg. The same fertilizer doses were applied in all combinations. In the respective sites, herbicides were added once in form of an aqueous emulsion at manufacturer’s recommended dose (RD) and 10-fold higher than the recommended manufacturer dose (10RD). The soil, along with micronutrients and herbicides, was thoroughly mixed and placed in polyethylene pots (capacity 3.7 dm3). The control soil was soil without addition of herbicides. On the same day, 8 maize grains each (variety LG 32.52) were sown into pots, and soil was moistened to 50% capillary water capacity. When the maize was at BBCH 10 stage (the first leaf emerging from the coleoptile), plants were aborted and 5 plants were left in each pot. During plant vegetation (60 days), soil moisture was monitored and maintained at a constant level (50% of capillary water capacity). The moisture content of soil was checked 3 times each day, and water losses were systematically supplemented with deionized water. According to BBCH scale , when Zea mays L. was at developmental stage BBCH 19 (9th leaf stage) and BBCH 33 (stage of 3 knees), the leaf greenness index in relative units of SPAD was determined using the Chlorophyll Meter SPAD 502 Plus Product Manual . The measurements were made in 5 replicates. At maize stage BBCH 39 (stage of 9 or more knees), the length of aboveground parts of plants and dry weight of plants were determined (fresh weight samples of plants were dried for 5 days at 65 °C). Moreover, at same time, soil samples were collected for microbiological and enzymatic analyses. The experiment included seven combinations, and each combination was carried out in 5 repetitions. The experimental scheme was as follows:Soil without addition of herbicides (control);−1 d.m. Soil per active substance—sulrd);Soil with addition of herbicide Sulcogan 300 SC at a dose recommended by the manufacturer (0.150 mg kg−1 d.m. Soil per active substance—Sul10RD);Soil with addition of Sulcogan 300 SC herbicide at a dose 10-fold higher than recommended by the manufacturer (1.50 mg kg−1 d.m. Soil per active substance—tezrd);Soil with addition of herbicide Tezosar 500 SC at a dose recommended by the manufacturer (0.165 mg kg−1 d.m. Soil per active substance—Tez10RD);Soil with addition of Tezosar 500 SC herbicide at a dose 10-fold higher than recommended by the manufacturer (1.65 mg kg−1 d.m. Soil per active substance—sucrd);Soil with addition of herbicide Sulcotrek 500 SC at a dose recommended by the manufacturer (0.333 mg kg−1 d.m. soil per active substance—Suc10RD).Soil with addition of Sulcotrek 500 SC herbicide at a dose 10-fold higher than recommended by the manufacturer . Before setting up experiment, the soil was sifted through a sieve (mesh diameter of 5 mm), and then 3.4 kg of soil was weighed each and following amounts of micronutrients were added per pure component : K+—168.0; Na+—10.0, Ca2+—11.50, Mg2+—82.10. Physicochemical analyses of soil were performed according to procedure described by Wyszkowska et al. [The soil used in experiment came from Tomaszkowo , near Olsztyn in north-eastern Poland . The soil was taken from the arable-humic layer at a depth of 0–20 cm. The soil material in terms of grain size was classified as sandy loam , belonging to Eutric Cambisols . The soi−1 d.m. soil: organotrophic bacteria—0.562 × 1010, actinomycetes—0.237 × 1010 and fungi—0.405 × 108, and activity of soil enzymes in kg−1 d.m. soil h−1: dehydrogenases—8.958 µmol TFF, catalase—0.365 mol O2, alkaline phosphatase—0.499 mmol PNP, acid phosphatase—3.128 mmol PNP, β-glucosidase—0.163 mmol PNP, arylsulfatase—0.269 mmol PNP, and urease—0.601 mmol N-NH4.The soil was characterized by the following number of microorganisms per cfu kgThree herbicides were applied to the soil in the study: Sulcogan 300 SC (Sul), Tezosar 500 SC (Tez), and Sulcotrek 500 SC (Suc). The tested preparations are used to protect maize against monocotyledonous and dicotyledonous weeds.−3. The dose recommended by manufacturer is 1.5 dm3 ha−1. It is a selective herbicide with systemic action, which comes in the form of a concentrate for dilution with water.Sulcogan 300 SC was manufactured by Nufarm Polska Sp. z o. o. , and it was registered in Poland in 2012. In its composition, it contains an active ingredient called sulcotrione (a compound of triketone group) in amount of 300 g dm−3. The dose recommended by manufacturer is 1.0 dm3 ha−1. It is a selective herbicide with systemic action, which is applied foliarly. It comes in the form of a concentrated suspension for dilution with water.Tezosar 500 SC was produced by Ciech Sarzyna S.A. , launched in Poland in 2018. The active ingredient of this preparation is terbuthylazine (a compound from triazine group), present in amount of 500 g dm−3 and terbuthylazine in amount of 327 g dm−3. The manufacturer recommends using this product at a dose of 2.0 dm3 ha−1.Sulcotrek 500 SC was produced by Adama Agan Ltd. , and it was registered in Poland in 2017. It is a selective herbicide with systemic action in the form of a concentrated suspension for dilution with water. It contains two active ingredients: sulcotrione in amount of 173 g dm−1 d.m. soil. Based on abundance of above-mentioned microorganisms, colony development (CD) index [Determination of the abundance of organotrophic bacteria, actinobacteria, and fungi was performed in 4 replicates for each soil sample using the serial dilution method according to the procedure described in paper by Kucharski et al.  and BoroD) index  and ecopD) index  were calDNA was isolated from soil without herbicides and with herbicides added at doses 10-fold higher the manufacturer’s recommended, using the Genomic Mini AX Bacteria+ kit . Amplification of bacterial 16S rRNA and fungal ITS gene fragment was carried out using universal primers 1055F and 1392R. The procedure for performing DNA isolation and conditions for conducting PCR reaction are described in work by Ferris et al. . Sequencβ-glucosidase, arylsulfatase, and urease according to Alef and Nannipieri [Biochemical analyses of soil were performed in 3 replicates for each combination. Dehydrogenases activity was determined according to Öhlinger procedure , catalasnnipieri . Enzyme nnipieri  and Wysznnipieri .p = 0.05 using Statistica 13.3 software [Zea mays L.) was also calculated according to formula presented in the work of Boros et al. [The results were statistically analyzed at a significance level of software , using msoftware . If IF =s et al. .http://www.interactivenn.net/, accessed on 24 July 2023) [Metagenomic data on bacterial and fungal families were statistically analyzed using the G-test (w/Yates’) and Fisher’s test using STAMP 2.1.3 software . Dominanly 2023) . MetagenActinobacteriota and Proteobacteria, which colonized the soil most numerously. In the case of fungi, there was a decrease in number of the phylum, Ascomycota, under the influence of Sulcogan 300 SC, and an increase under the influence of Tezosar 500 SC and Sulcotrek 500 SC. The number of Mortierellomycota fungi decreased after the application of Sulcogan 300 SC and Tezosar 500 SC, and increased in soil with Sulcotrek 500 SC. All tested herbicides inhibited the growth of fungi belonging to the phylum, Basidiomycota. The maize proved tolerant to the herbicides applied to the soil, as evidenced by an increase in the tolerance index (TI) and the leaf greenness index (SPAD) values of the plants. This research is necessary in the search for better and new solutions in restoring soil homeostasis, and, thorough an understanding of the soil microbiome in the presence of Sulcogan 300 SC, Tezosar 500 SC, and Sulcotrek 500 SC herbicides, it can serve as a suitable tool for the bioaugmnetation of soils contaminated with these preparations. The microorganisms that may possess the ability to inactivate these herbicides are Lysobacter and Sphingobium, unique in soil with Sulcogan 500 SC, and Candidatus_Solibacter and Haliangium, univalent in soil with Sulcotrek 500 SC, as well as Trichaoderma fungi, present in soil with the tested herbicides.Herbicides applied both at the dose recommended by the manufacturer and at a dose 10-fold higher stimulated the proliferation of organotrophic bacteria and the activity of arylsulfatase and acid phosphatase, while causing inhibition of urease activity. They increased the number of bacteria belonging to the phyla,