Biostimulants combination enhance the metabolites content of Brassica oleracea var. botrytis curds cultivated in the desert region of Basrah governorate

Biostimulants combination enhance the metabolites content of Brassica oleracea var. botrytis curds cultivated in the desert region of Basrah governorate | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Biostimulants combination enhance the metabolites content of Brassica oleracea var. botrytis curds cultivated in the desert region of Basrah governorate Wurood Hantoosh Neamah, Zainab A. Al-sudani, Fatimah Ali Hasan, and 1 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7614364/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 04 Mar, 2026 Read the published version in BMC Plant Biology → Version 1 posted 12 You are reading this latest preprint version Abstract Among Brassica vegetables, cauliflower ( Brassica oleracea var. botrytis ) is widely recognized as a consumer favorite due to its rich and healthy content of nutrients and bioactive compounds. The enhancement of productivity and quality of preferred crops has gained highly importance in recent decades, driven by increasing customers demand for affordable and healthy food. Away from chemical materials and their risky side effects, biostimulants and biofertilizers could be a suitable alternative for traditional fertilizers under desert conditions. In a current study the extract of Liquorice root LRE and bread yeast BYE applied both with 0, 10, and 20 g/L on Brassica plant to explore their influence in curds metabolites. Obtained results showed that while carbohydrates reduced in curds of treated plants, protein increased with both stimulants application. The non-significant increasing in phenols and flavonoids content was observed in curds of LRE and BYE-treated plants with an elevation in antioxidant activity in their methanolic extracts. Both LRE and BYE applications promoted the curds content of macronutrients and caused a notable alteration in active compounds profile. Noteworthy, the values of acquired outcomes increased or decreased according to applied concentrations of individual or combination of both biostimulants. Based on our finding, the foliar application with LRE and BYE enhanced the primary and secondary metabolites of Brassica curds. However, the combination of 10 g/L of LRE and 20 g/L of BYE is recommended due to its act in promoting of flavonoids content, antioxidant capacity, potassium content and active compounds profile including organosulfur, troponoid, methyl 9-cis,11-trans-octadecadienoate, 1H-Indole-3-acetonitrile and 9,12,15-Octadecatrienoic acid. Biostimulants Liquorice root extract Bread yeast extract Cauliflower curds Metabolites. Desert region Figures Figure 1 Figure 2 Figure 3 1- Introduction To achieve sustainability in the agricultural sector, it is necessary to eliminate synthetic fertilizers and apply instead biostimulants (BIOs) to enhance both the quantity and quality of crops. Biostimulants are substances or/and microorganisms that optimize plant growth through enhance nutrition efficiency, soil fertility, abiotic stress tolerance, and crop quality without direct effect on the nutrients content (Du Jardin, 2015 ). BIOs are commercially available in numerous forms and could be designated as product contain mixture of both substances and microorganism. Patrick du Jardin divided biostimulants into seven categories: humic and fulvic acids, protein hydrolysates (PHs), oligopeptides and polypeptides, seaweed and botanical extracts, biopolymers, and beneficial fungi and bacteria (Du Jardin, 2015 ). BIOs derived from plants have received a notable attention in the recent years and researchers highly investigated their potential effect on plants growth under normal and stress conditions (Han et al., 2024 ; keya Tudu et al., 2022 ). Among plants that used their extracts as BIOs, Liquorice ( Glycyrrhiza glabra ) root extract (LRE) is used particularly due to the rich content of osmoprotectants, phytohormones and antioxidants (E. Desoky et al., 2019 ). LRE has been reported as a promising natural biostimulants due to its role in enhancement the plant ability to uptake nutrients (Salih et al., 2021 ), promote a good growth and yield (Elrys & Merwad, 2017 ), and improve abiotic stress tolerance (E.-S. M. Desoky, A. S. Elrys, et al., 2019; E.-S. M. Desoky, A. I. ElSayed, et al., 2019). Numerous active compounds exist in LRE extract such as antioxidants, phenols, flavonoids, triterpenoid and saponins and help to stimulate its favourable role when applied on crops (Mamedov & Egamberdieva, 2019 ). Likewise, recent utilization of bread yeast Saccharomyces cerevisiae extract (BYE) as biostimulants have garnered attention for their potential to enhance plant growth and contribute to sustainable agriculture (Vargas et al., 2024 ). BYE derived from yeast cells is rich in effective components such as low-molecular-weight organic substances, amino acids, nucleotides, peptides, nitrogen, phosphorus, and trace elements with absence of chemical hormones and toxic ingredients (Vieira et al., 2016 ). The foliar application of BYE on plants can stimulate plant metabolism and enhance the vitality, leading to boost nutrient uptake, increase biomass, improv biochemical traits and consequently optimize the high value productivity in cabbage and tomato crops (Dima et al., 2023 ; Dima et al., 2020 ). White cauliflower Brassica oleracea var. botrytis is annual plant belongs to the Brassica family, is an edible and highly desirable crop among Iraqi consumers. The edible parts of the plants are immature inflorescences, heads, or curds. Curds are rich with a privileged content of nutrients and active constituents such as fibres, soluble sugars, minerals, carotenoids, vitamins, glucosinolates, polyphenols, and flavonoids (Fouad A Ahmed & Rehab FM Ali, 2013). White cauliflower is one of crops that are voracious for fertilizers special the nitrogen. Achieving high and good productivity of crop could be required massive amounts of fertilizers that are costly, polluted, and not ensuring the sustainable requirements. Thus, exploring suitable alternatives such as biofertilizers and biostimulants is necessary to reduce the agriculture cost, optimize the quality and quantity of yield, minimize global carbon footprint and protect our environment (Díaz-Pérez et al., 2024 ). Current study is aimed to investigate the effect of LRE and BYE combination as biostimulants on nutrient value and active component profile and their antioxidant activity in Brassica oleracea var. botrytis curds that grown in desert area in Basrah. 2- Materials and methods 2.1. Cultivation and treatments The verity cauliflower hybrid F1 Alnahar seeds from Monarch seeds, Noord-Scharwoude, Holland, cultivated in trays with total peat moss substrate. After 40 days of cultivation, seedlings transferred to permanent field, treatment with biostimulants started after 30 days of transferring. Biostimulants treatments were performed weakly with LRE and BYE at three concentrations (0, 10, 20) g/L, the period between each treatment was 2 weeks. 2.2. Biochemical traits of curds The cruds of treated plants were dried at room temperature and used to estimate the content of carbohydrates, protein, phenols, flavonoids, and elements as well as their active constituent’s profile 2.2.1. Carbohydrates content Carbohydrates content was determined by Phenol-Sulphuric Acid method according to (DuBois et al., 1956). From dried samples 0.5 g homogenized with 70 ml of distilled water at 70 °C for 1 hr. Then, 5 ml of filtrated extract placed in a volumetric flask and made up to 30 ml by distilled water. From diluted extract, 1 ml placed in a new tube and homogenized with 1m of 5% phenol and 5 ml of pure sulfuric acid by vortex. After brining the mixture to the room temperature, the absorption recorded by spectrophotometer at 490 nm and carbohydrates concentration was valuated from glucose standard curve and expressed as mg of glucose equivalents per gram of sample dry weight (mg GU/g dw). 2.2.2. Protein content Protein content was determined in dried plant samples following Kjeldahl method. From dried samples 0.20 g was digested with 5 ml of pure sulfuric acid at 350 °C for 30 minutes. After the samples cooling, 3 ml of the mixture of 96 ml sulfuric acid and 6 ml perchloric acid was added to the sample solution and placed on digestion plate at 350°C until the solution being clear. The volume of sample solution was made up to 50 ml by distilled water. From diluted sample 10 ml was mixed with 10 ml of 6 N of sodium hydroxide in the tube and placed in Kjeldahl analyzer instrument. The distillate was collected in a conical flask containing 2 % of Boric acid and 2% of bromocresol green and methyl red indicator solution (0.099 g + 0.066 g in 100 ml Ethanol). The collection was titrated with 2 % HCl until the solution has a slightly violet color, the percentage of nitrogen was calculated through the equation: % %N= Percentage of nitrogen V= Volume of titration acid (HCl) N= Normality of HCl MW= Molecular weight of nitrogen TV= Total volume of digestion solution SW= Weight of plant sample VS= Volume of digestion solution The percentage of protein was calculated following the question: (conversion factor) 2.2.3. Total phenols content Cruds content of phenols estimated by Folin–Ciocalteu method according to (Porter, 2012). With 5 ml of 80% methanol, 0.25 g of dry sample was homogenized and kept 48 hrs in dark, 24 hrs at room temperature and 24 hrs at 4 °C. To 0.5 ml of the methanolic extract, 0.5 ml of Folin–Ciocalteu reagent was added and followed with 8 ml of distilled water. After 5min of incubation, 1ml of 20% sodium carbonate (Na2CO3) was added. After 30 mins incubation at ambient temperature in dark, an intense blue color developed, and its absorbance was measured at 760 nm. Gallic acid was used as a standard by preparing stock solution at concentration 1 mg/ml and used for the calibration. The results were expressed as mg of gallic acid equivalents per gram of the dry weight sample (mg GAE/g dw) 2.2.4. Total flavonoids content Total flavonoids content in cruds of control and treated plants was evaluated following (Samidha Kamtekar, 2014). To 0.5 ml of extract that placed in 10 ml tubes, 0.5 ml of 80% methanol added and followed by the adding of 10 ml of distilled water and 0.3 ml of 5% sodium nitrite (NaNO3). After incubation for 5 mins, 0.3 ml of 10% aluminum chloride (AlCl3) and 2 ml of 1M sodium hydroxide (NaOH) were added. The volume was made up to 10 ml with distilled water and vortexed, the absorbance was recorded spectrophotometry at 510 nm after orange-yellowish color development. Quercetin was used as a standard by preparing stock solution at concentration 1 mg/ml and used for the calibration. The results were expressed as mg of quercetin equivalent per gram of the dry weight sample (mg QE/g dw) . 2.2.4. Elements composition Potassium, phosphors, and sulfur were determined by the dry ash method (Akinyele & Shokunbi, 2015). In porcelain crucibles, 1 g of cruds samples were placed and incinerated at an initial temperature of 200 ◦ C for 1h on a hot plate until the smoke disappears. Then, samples kept in Nabertherm muffle furnace at 556 °C for 5 hrs for total incineration. Next, ashes were dissolved by a mixture of 4 ml of nitric acid (65%) and 10 ml of distilled water on a hot plate. The solution was filtered and made up to 25ml with distilled water and used for elements (P, K , S) analysis by atomic absorption/emission spectroscopy. The calibration curves were generated using the standards of each element, and the results were expressed as mg of element per gram of the dry weight sample. 2.2.5. GC-MS analysis 2.2.5.1. Samples preparation From dried cruds, 0.250 g homogenized with 5 ml of 80% methanol, vortexed to achieve a good mixing and incubated in dark at room temperature overnight. Next day, supernatant collected carefully by micropipette and sent for injection in GCMC instrument 2.2.5.2 GCMC conditions GC-MS analysis was carried out by using an Agilent Technologies, 7890B GC system coupled to an Agilent Technologies 5977A MSD with EI Signal detector, using HP-5ms 5% phenyl, 95% methyl siloxane (30m*250um*0.25). The oven temperature was set at 40 °C hold for 5 mins then raised to 8 °C/min to 300 °C for 20 mins, Helium carrier gas flow rate was 1 ml/min and purge flow of 3 ml/min. The injection mode was pulsed Splitless with injection temperature 290 °C and the injection sample volume was 0.5 µl. The mass spectrometer used Ion Source Temperature 230 °C, with scan speed 1562(N2), and the mass range 44-750 m/z. Compound identification was implemented using mass spectral data and retention indices, in comparison with authentic standards and spectral libraries, including the NIST 2014, 2020 database. 3- Results Figure 1 illustrates the effect of biostimulants spray on curds content of carbohydrates and proteins, the spray with LBE and BYE at different concentration caused reduce the carbohydrates content of curds compared to the control plants, the range of values between 19.32 mg GU/g dw and 22.86 mg GU/g dw. On the other hand, protein proportion increased remarkably in curds of 10 g/L of BYE-treated plants (15.89 %) and plants that sprayed with 20 g/L of LRE and 10 g/L of BYE combination (14.87 %) (Fig.1A&B). Total phenolic content and total flavonoids content in curds exhibited non-significant increase due to foliar application of both extracts on Brassica plants. The increase of phenols content was in curds of 20 g/L of BYE-treated plants (17.03 mg GAE/g dw), while flavonoids content increased in curds of combination of 10 g/L of LRE and 20 g/L of BYE-treated plant (16.54 mg QE/g dw) (Fig. 2A&B). Antioxidants activity of curds extracts against 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals reinforced in curds methanolic extract of LRE-sprayed plants and BYE-sprayed plants and their combination-sprayed plants recorded 75.00 %, 77.59 %, and 74.09 % respectively (Fig. 2C). Elements composition including nitrogen, phosphors, calcium and sulfur displayed the effect of foliar application of biostimulants on macro elements content in Brassica curds. Nitrogen portion increased in curds of 10 g/L BYE-treated plants and 20 g/L LRE and 10 g/L BYE-treated plants (2.54, 2.38) % (Fig. 3A). Phosphor content was impacted with individual and combination applications of LRE and BYE, P content promoted in curds of 10 g/L of LRE and BYE-treated plants (5.67 mg/g), 20 g/L LRE-treated plants (6.00 mg/g) and 20 g/L of LRE and BYE-treated plants (6.10 m/g) of dry weight (Fig. 3B). Furthermore, potassium content increased subsequently the foliar spray with LRE and BYE. Curds of 20 g/L BRE-treated plants (28.83 mg/g), 10 g/L of LRE-treated plants (23.31 mg/g) and the combination 10 g/L of LRE and 20 g/L of BRE-treated plants (28.52 mg/g) of dry weight (Fig. 3C). Sulfur content in Brassica curds was boosted in curds of plants that treated with 10 g/L of LRE and the combination of 20 g/L of LRE and 10 g/L of BRE that both recorded 3. 60 mg/g of dry weight (Fig. 3D). GC-MS profile of methanolic Brassica curds extract of treated plants represents alteration in the active constituents compared to control plants (Tab. 1). The alteration may involve increase or decrease or disappearance of detected compounds area percentage according to applied treatments. For example, dimethyl trisulfide (compound is responsible for off-flavour in cooked Brassica vegetable) increased in curds of 10 g/L-treated plants and plants that sprayed with combination of 10 g/L and 20 g/L of LRE and BYE respectively. While compounds like phenol decreased in curds of sprayed plants with both biostimulants comparing to untreated plants. Saturated fatty acid Hexadecanoic acid and polyunsaturated fatty acid 9,12,15-Octadecatrienoic acid increased substantially in plants curds due to foliar application with biostimulants particularly with combination applications. Methyl ester compound such as Methyl 9-cis,11-trans-octadecadienoate was inexistent in untreated plants and gradually increasing with individual or combinated applications according to GC-MS profile. However, several compounds such as 4-Ethoxy-2-(methylamino) tropon, Furan-2-carboxaldehyde, 5-(1-piperidyl)- and gamma.-Sitosterol vanished in methanolic extract of Brassica curds due to influence of individual or combination applied treatments. Table (1): GC-MS analysis profile of Brassica methanolic extract R.T. Compound name Chemical class L0B0 L0B1 L0B2 L1B0 L1B1 L1B2 L2B0 L2B1 L2B2 9.26 Dimethyl trisulfide Organosulfur 3.68 3.58 3.13 4.06 2.87 4.98 2.81 2.79 2.89 18.43 Phenol, 4-ethenyl-2,6-dimethoxy- Phenolic compounds 1.47 0.72 0.75 1.00 0.83 0.60 0.52 0.58 0.64 19.5 4-Ethoxy-2-(methylamino) tropone Furan-2-carboxaldehyde, 5-(1-piperidyl)- Troponoid Volatile compound 8.48 - 5.79 - 6.16 - - 7.23 - 6.37 4.10 - 5.29 - 5.84 - - 6.54 20.72 d-Proline Amino acid 0.97 1.46 1.55 2.19 1.86 - 3.31 2.69 - 21.26 1H-Indole-3-acetonitrile Glycosides 1.65 1.53 1.55 1.53 1.68 1.83 1.15 1.10 1.03 22.19 Hexadecanoic acid Sat. Fatty acid 1.23 3.26 2.67 2.27 4.21 5.07 4.59 7.44 4.43 23.82 Methyl 9-cis,11-trans-octadecadienoate Methyl ester - 2.59 2.04 1.56 2.032 4.65 3.18 4.73 2.89 23.89 9,12,15-Octadecatrienoic acid Unsat. Fatty acid 1.24 10.12 9.35 7.42 10.83 21.10 13.11 15.30 9.99 27.33 Glycerol monopalmitate Fatty acid 1.33 1.18 1.51 1.57 2.18 1.38 2.18 1.73 0.86 33.29 .gamma.-Sitosterol Sterol 0.45 0.51 0.76 2.11 - - 1.93 2.00 1.53 4- Discussion The utilization of botanical extract as biofertilizer or biostimulants to enhance plant growth or plant tolerance to stress conditions has gained considerable attention in recent years. Obtained results showed that carbohydrates content in curds decreased when foliar spray applied on Brassica plants which could be attributed to exploiting carbohydrates to increase the growth parameters of sprayed plants including plant high, leaves number, leaf area and biomass (data not shown). Furthermore, total carbohydrates were utilized also by treated plants to increase the productivity traits such as yield, diameter, biomass, and weight of Brassica cruds (data not shown). Under similar conditions, instead of carbohydrates accumulation in induvial organs, plant direct total carbohydrates to promote its growth and yield (Van den Ende, 2014 ). On the other hand, protein percentage exhibited an increase in curds of L0B1-treated plants and L2B1-treated plants. The role of protein in the plant cover expanded ambit through their structural, enzymatic, metabolic, regulatic and storage functions (Rasheed et al., 2020 ). Foliar application with biostimulants that are rich with protein such as BYE (Demirgul et al., 2022 ) or amino acid such as LRY (Wang et al., 2015 ) or both could increase the curds content of protein and promote yield quality from the healthy components. Brassica curds are good source for phenolic compounds particularly flavonoids (F. A. Ahmed & R. F. Ali, 2013). Phenols including flavonoids are natural compounds that are synthesized by the phenylpropanoid pathway and widely distributed in plants to promote and maintain its growth and development. Besides, they have several medical benefits such as anticancer, antiinflammation and antioxidant properties (Kuljarusnont et al., 2024 ). In our study, the total of phenols and flavonoids in curds have not shown significantly affected due to the foliar application with LRY or BYE. However, an insignificant increase was observed in phenols and flavonoids content in curds of L0B2-treated plants and L1B2-treated plants respectively. This increase may be related to the role of LRY and BYE in boosting and stimulating secondary metabolites production including total phenols and flavonoids, conforming with obtained outcomes by Younes in Allium cepa (Younes et al., 2021 ) and Dawood in Glycine max (Dawood et al., 2013 ). In the antioxidant context, previous literature indicated the active constituents in Brassicaceae vegetables and their related antioxidant properties (Mulțescu et al., 2021 ). The antioxidants activity in Brassica curds is correlated to various compounds such as phenols, flavonoids, sulfurs compounds, glucosinolates and others. This activity may be influenced extremely by several factors including cultivar, maturity, growing and harvesting conditions and storge circumstances (Podsędek, 2007 ). Following the effect of LRE and BYE on antioxidant activity of curds methanol extract, L0B2-treated plant and L1B0-treated plants recorded a significant activity. Likewise, curds extract of plants sprayed with combination of both LRY and BYE concentrations recorded also antioxidant activity against DPPH. The augmentation of DPPH scavenging efficiency due to LRE implementing was observed previously in the extract of Stevia rebaudiana bertoni under normal conditions (Khashan & AL-Taweel, 2024 ) and Phaseolus vulgaris L. under salinity conditions (Rady et al., 2019 ), both found LRE promoted antioxidant activity in plants due to increase the non-enzymatic antioxidant (e.g. proline, phenols, tocopherols, carotenoids, etc). In the current study, the capacity of curds extract to scavenging DPPH radicals could be increased regarding the increase of Dimethyl trisulfide, d-Proline, 9,12,15-Octadecatrienoic acid and gamma.-Sitosterol (Table 1). Likewise, BYE application boosted antioxidants properties in Allium sativum L. as a tolerance strategy against drought stress through increasing of antioxidant enzymes and antioxidants compounds such as proline and ascorbic acid (Abdelaal et al., 2021 ). Our result indicated that BYE enhanced the scavenging of free radicals in curds extract of treated plants which could be imputed to induced role of BY in promoting compounds with antioxidants features including Dimethyl trisulfide, d-Proline, 9,12,15-Octadecatrienoic acid, Methyl 9-cis,11-trans-octadecadienoate, Glycerol monopalmitate and gamma.-Sitosterol. These compounds increased also in curds extract of sprayed plants with combination of LRE and BYE according to GC-MS analysis (Table 1) which consequently enhanced its antioxidants efficiency. Microelements (N, P, K, S) increased compatibly with biostimulants foliar application. Brassica curds have been reported to contain a high macro elements content (Ekholm et al., 2007 ). However, cultivars, growing and environmental conditions, and harvesting and post harvesting circumstances influence curds content of macro and micro elements (Kalisz et al., 2019 ; Kisko et al., 2021 ). In a present study potassium recorded the highest content in Brassica curds and the content increased remarkably with LRE and BRE at L0B2 and L1B2-treated plants. Likewise, other macronutrients including N, P, and S increased in curds of treated plants: for N at L0B1 and L2B1, for P at L1B1, L2B0, and L2B1, and for S at L1B0 and L2B1. The increase of macro elements in curds may be referred to a rich content of elements in LRE (Gormez & Sengul, 2014 ) and BRE (Tao et al., 2023 ), these elements either move through or interact with the apoplast space which subsequently effect on their translocation to different plant parts including the curds (Fernández & Brown, 2013 ). Additionally, this increase may be linked to enhanced root capacity for nutrients uptake from the soil post biostimulants application (Halpern et al., 2015 ). It has been reported that LRE and BYE increased minerals content in mango fruit and potato tuber when biostimulants applied on the plants (Orján et al., 2023 ; Yousif et al., 2024 ). Macro minerals almost participate in all human body functions and act significantly in metabolic processes. For example, phosphorus serve important role in the bone, teeth, phosphoproteins and nucleic acids formation, fat and starch metabolism and heartbeat and kidney function normalization (Elser, 2012 ). GC-MS is mostly favoured technique to identify the quality and quantity of compounds in complex mixture due to its sensitivity and selectivity. Numerous agents influence GC-MS analysis results, some are related to plants (plants species, growing conditions, extraction methods) and others are related to GC-MS instrumentation conditions (temperature program, injection technique, detector sensitivity). Under our experimental conditions, GC-MS result of curds methanolic extract indicated high alteration in the peak area of most important detected compounds. Several compounds increased due to LRE or BYE or both applications such as Dimethyl trisulfide (L1B0, L1B2), d-Proline (L0B1, L0B2, L1B0, L1B1, L2B0, L2B1), 1H-Indole-3-acetonitrile (L1B1, L1B2), Hexadecanoic acid (all treatments), Methyl 9-cis,11-trans-octadecadienoate (all treatments), Glycerol monopalmitate (all treatment except L2B2) and gamma.-Sitosterol (L0B1, L0B2, L1B0, L2B0, L2B1, L2B2). On the contrary, several compounds decreased due to individual or combination of foliar applications such as Dimethyl trisulfide (L0B1, L0B2, L1B0, L1B1, L2B0, L2B1, L2B2), Phenol, 4-ethenyl-2,6-dimethoxy- (all treatments), 4-Ethoxy-2-(methylamino) tropone (L0B1, L0B2, L1B2, L2B0, L2B1), 1H-Indole-3-acetonitrile (L0B1, L0B2, L1B0, L2B0, L2B1, L2B2), and Glycerol monopalmitate (L2B2). Interestingly, some compounds disappeared due to LRE or BYE or both applications such as 4-Ethoxy-2-(methylamino) tropone (L1B0, L1B1, L2B2), Furan-2-carboxaldehyde, 5-(1-piperidyl)- (L0B0, L0B1, L0B2, L1B2, L2B0, L2B1), d-Proline (L1B2, L2B2), and gamma.-Sitosterol (L1B1, L1B2). The alteration of active compounds in Brassica curds extract with LRE foliar spray may be correlated to rich content of LRE from sugars, vitamins, mineral nutrients, amino acids, selenium, and phytohormones which effect on secondary metabolites biosynthesis pathways such as sulfurs-containing compounds pathway (Nakai & Maruyama-Nakashita, 2020 ), shikimate/phenylpropanoid pathway (Tak et al., 2023 ), hormone biosynthesis pathways (Bajguz & Piotrowska-Niczyporuk, 2023 ), and sterol biosynthesis pathway (Du et al., 2022 ). Likewise, BYE application caused a variation in peak areas of secondary compounds in Brassica curds extract which may be referred to the BYE content of effective components including amino acids, vitamins, proteins and hormones (Medani & Taha, 2015 ). Some of these components serve as precursors for secondary metabolites biosynthesis pathways such as amino acids (Trovato et al., 2021 ), while some act as enzymes cofactors, signaling molecules, and genes expression regulators such as vitamins and hormones (Parra et al., 2018 ; Rui et al., 2020 ). These active compounds in Brassica curds have crucial role in maintaining human health when incorporated into the daily dietary due to their therapy functions as antioxidants, antiinflammation, anticancer, antiviral, anti-obesities, antihypertensive, and antigenotoxicity (Zhang et al., 2025 ). For example, 1H-Indole-3-acetonitrile displayed a broad-spectrum antiviral potential against influenza A, HSV-1, VSV and SARS-CoV-2 (Hui et al., 2023 ; Zhao et al., 2021 ). Thus, the increase of their amount after LRE and BYE application may enhance the nutritional, health-promoting and therapeutic value of brassica curds. 5- Conclusion In summary, we found that the combination of biostimulants including LRE and BYE could be affected prominently on metabolites content of Brassica curds. When two biostimulants are applied as combination, their act may complement each other or act synergistically to enhance the overall effect, resulting in improving plants yield quantity and quality in the region that has desert soil properties. Our results indicated an increase in primary and secondary metabolites in Brassica curds after foliar application of LRE and BYE concentrations. Although the induvial treatments had an obvious impact on experimental parameters, the combination of study agents also acted synergistically to boost these parameters. In crops cultivation under desert conditions, obtained findings could be considered a promising result within the sustainable required direction. The remarkable function of LRE and BYE in improving the quality components of Brassica curds including phenols, flavonoids, macronutrients, and active compounds provides compelling evidence of the alternative role that biostimulants may play in eco-friendly agriculture. This improvement not only increase the nutritional value of Brassica curds but also enrich them with active compounds that contribute to the promoting of healthy foods sources. Declarations Author contribution Z.A designed research studies and performed field work, W.N and F.H performed lab experiments and data analysis. W.N and F.A wrote/edited the manuscript. 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10:58:16","extension":"xml","order_by":17,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":127844,"visible":true,"origin":"","legend":"","description":"","filename":"26cb706bd6d64a54a451d74f7da3c29b1structuring.xml","url":"https://assets-eu.researchsquare.com/files/rs-7614364/v1/2e3f6b8cb9c28bb4ae950d3a.xml"},{"id":92587704,"identity":"8b98d821-28e1-4fcd-bc1a-6f93d81e71c0","added_by":"auto","created_at":"2025-10-01 10:58:16","extension":"html","order_by":18,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":134172,"visible":true,"origin":"","legend":"","description":"","filename":"earlyproof.html","url":"https://assets-eu.researchsquare.com/files/rs-7614364/v1/799b6cfd90e67e777c889917.html"},{"id":92587949,"identity":"6ec379ae-1a49-44a8-882e-20c49f855379","added_by":"auto","created_at":"2025-10-01 11:06:15","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":20536,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eThe effect of foliar application with Liquorice root extract and Bread yeast extract on carbohydrates and protein content in Brassica curds.\u003c/strong\u003e (A) Representative the effect of Liquorice root extract and Bread yeast extract on the carbohydrates content of Brassica curds. L0B0 control or untreated plants. L0B1treated plants with Bread yeast extract at 10 g/L. L0B2 treated plants with Bread yeast extract at 20 g/L. L1B0 treated plants with 10 g/L of Liquorice root extract. L1B1treated plants with 10 g/L of Liquorice root extract and 10 g/L of Bread yeast extract. L1B2 treated plants with 10 g/L of Liquorice root extract and 20 g/L of Bread yeast extract. L2B0 treated plant with Liquorice root extract. L2B1 treated plants with 20 g/L of Liquorice root extract and 10 g/L of Bread yeast extract. L2B2 treated plants with 20 g/L of Liquorice root extract and Bread yeast extract. (B) Representative the effect of Liquorice root extract and Bread yeast extract on the protein content of Brassica curds. A multiple ANOVA was performed using ordinary one-way ANOVA multiple comparisons to compare the averages of treatments with L0B0. Significance was designated as follows: *p \u0026lt; 0.05, ***p \u0026lt; 0.0001.\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-7614364/v1/281c9d4577417e0950c879fe.png"},{"id":92587685,"identity":"7d0a36dd-b26b-4f7b-b080-8a0044a67fa1","added_by":"auto","created_at":"2025-10-01 10:58:15","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":32584,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eThe effect of foliar application with Liquorice root extract and Bread yeast extract on phenols and flavonoids content and DPPH scavenging in Brassica curds.\u003c/strong\u003e (A) Representative the effect of Liquorice root extract and Bread yeast extract on the phenols content of Brassica curds. L0B0 control or untreated plants. L0B1treated plants with Bread yeast extract at 10 g/L. L0B2 treated plants with Bread yeast extract at 20 g/L. L1B0 treated plants with 10 g/L Liquorice root extract. L1B1treated plants with 10 g/L of Liquorice root extract and 10 g/L of Bread yeast extract. L1B2 treated plants with 10 g/L of Liquorice root extract and 20 g/L of Bread yeast extract. L2B0 treated plant with Liquorice root extract. L2B1 treated plants with 20 g/L of Liquorice root extract and 10 g/L of Bread yeast extract. L2B2 treated plants with 20 g/L of Liquorice root extract and Bread yeast extract. (B) Representative the effect of Liquorice root extract and Bread yeast extract on the flavonoids content of Brassica curds. (C) Representative the effect of Liquorice root extract and Bread yeast extract on the DPPH scavenging of Brassica curds extract. A multiple ANOVA was performed using ordinary one-way ANOVA multiple comparisons to compare the averages of treatments with L0B0. Significance was designated as follows: *p \u0026lt; 0.05, **P \u0026lt; 0.01, ****p \u0026lt; 0.0001.\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-7614364/v1/9aeadfdfd39cc2afb2b01540.png"},{"id":92587687,"identity":"63593863-a6fa-450d-b7c7-e61957fcae81","added_by":"auto","created_at":"2025-10-01 10:58:15","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":39473,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eThe effect of foliar application with Liquorice root extract and Bread yeast extract on macro elements content in Brassica curds. \u003c/strong\u003e(A) Representative the effect of Liquorice root extract and Bread yeast extract on the nitrogen content of Brassica curds. L0B0 control or untreated plants. L0B1treated plants with Bread yeast extract at 10 g/L. L0B2 treated plants with Bread yeast extract at 20 g/L. L1B0 treated plants with 10 g/L Liquorice root extract. L1B1treated plants with 10 g/L of Liquorice root extract and 10 g/L of Bread yeast extract. L1B2 treated plants with 10 g/L of Liquorice root extract and 20 g/L of Bread yeast extract. L2B0 treated plant with Liquorice root extract. L2B1 treated plants with 20 g/L of Liquorice root extract and 10 g/L of Bread yeast extract. L2B2 treated plants with 20 g/L of Liquorice root extract and Bread yeast extract. (B) Representative the effect of Liquorice root extract and Bread yeast extract on the phospho content of Brassica curds. (C) Representative the effect of Liquorice root extract and Bread yeast extract on the potassium content of Brassica curds. (D) Representative the effect of Liquorice root extract and Bread yeast extract on the sulfur content of Brassica curds A multiple ANOVA was performed using ordinary one-way ANOVA multiple comparisons to compare the averages of treatments with L0B0. Significance was designated as follows: *p \u0026lt; 0.05, ****p \u0026lt; 0.0001.\u003c/p\u003e","description":"","filename":"3.png","url":"https://assets-eu.researchsquare.com/files/rs-7614364/v1/9ec6a60416c86c8254e65fda.png"},{"id":104252025,"identity":"f5fdd7d8-0802-41b1-8710-425c8a64fc20","added_by":"auto","created_at":"2026-03-09 16:16:45","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":815872,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7614364/v1/1a92b69e-bced-4054-9cf6-6a98d03537a3.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Biostimulants combination enhance the metabolites content of Brassica oleracea var. botrytis curds cultivated in the desert region of Basrah governorate","fulltext":[{"header":"1- Introduction","content":"\u003cp\u003eTo achieve sustainability in the agricultural sector, it is necessary to eliminate synthetic fertilizers and apply instead biostimulants (BIOs) to enhance both the quantity and quality of crops. Biostimulants are substances or/and microorganisms that optimize plant growth through enhance nutrition efficiency, soil fertility, abiotic stress tolerance, and crop quality without direct effect on the nutrients content (Du Jardin, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). BIOs are commercially available in numerous forms and could be designated as product contain mixture of both substances and microorganism. Patrick du Jardin divided biostimulants into seven categories: humic and fulvic acids, protein hydrolysates (PHs), oligopeptides and polypeptides, seaweed and botanical extracts, biopolymers, and beneficial fungi and bacteria (Du Jardin, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). BIOs derived from plants have received a notable attention in the recent years and researchers highly investigated their potential effect on plants growth under normal and stress conditions (Han et al., \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2024\u003c/span\u003e; keya Tudu et al., \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Among plants that used their extracts as BIOs, Liquorice (\u003cem\u003eGlycyrrhiza glabra\u003c/em\u003e) root extract (LRE) is used particularly due to the rich content of osmoprotectants, phytohormones and antioxidants (E. Desoky et al., \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). LRE has been reported as a promising natural biostimulants due to its role in enhancement the plant ability to uptake nutrients (Salih et al., \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e2021\u003c/span\u003e), promote a good growth and yield (Elrys \u0026amp; Merwad, \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2017\u003c/span\u003e), and improve abiotic stress tolerance (E.-S. M. Desoky, A. S. Elrys, et al., 2019; E.-S. M. Desoky, A. I. ElSayed, et al., 2019). Numerous active compounds exist in LRE extract such as antioxidants, phenols, flavonoids, triterpenoid and saponins and help to stimulate its favourable role when applied on crops (Mamedov \u0026amp; Egamberdieva, \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). Likewise, recent utilization of bread yeast \u003cem\u003eSaccharomyces cerevisiae\u003c/em\u003e extract (BYE) as biostimulants have garnered attention for their potential to enhance plant growth and contribute to sustainable agriculture (Vargas et al., \u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). BYE derived from yeast cells is rich in effective components such as low-molecular-weight organic substances, amino acids, nucleotides, peptides, nitrogen, phosphorus, and trace elements with absence of chemical hormones and toxic ingredients (Vieira et al., \u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). The foliar application of BYE on plants can stimulate plant metabolism and enhance the vitality, leading to boost nutrient uptake, increase biomass, improv biochemical traits and consequently optimize the high value productivity in cabbage and tomato crops (Dima et al., \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Dima et al., \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2020\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eWhite cauliflower Brassica \u003cem\u003eoleracea\u003c/em\u003e var. \u003cem\u003ebotrytis\u003c/em\u003e is annual plant belongs to the Brassica family, is an edible and highly desirable crop among Iraqi consumers. The edible parts of the plants are immature inflorescences, heads, or curds. Curds are rich with a privileged content of nutrients and active constituents such as fibres, soluble sugars, minerals, carotenoids, vitamins, glucosinolates, polyphenols, and flavonoids (Fouad A Ahmed \u0026amp; Rehab FM Ali, 2013). White cauliflower is one of crops that are voracious for fertilizers special the nitrogen. Achieving high and good productivity of crop could be required massive amounts of fertilizers that are costly, polluted, and not ensuring the sustainable requirements. Thus, exploring suitable alternatives such as biofertilizers and biostimulants is necessary to reduce the agriculture cost, optimize the quality and quantity of yield, minimize global carbon footprint and protect our environment (D\u0026iacute;az-P\u0026eacute;rez et al., \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2024\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eCurrent study is aimed to investigate the effect of LRE and BYE combination as biostimulants on nutrient value and active component profile and their antioxidant activity in Brassica \u003cem\u003eoleracea\u003c/em\u003e var. \u003cem\u003ebotrytis\u003c/em\u003e curds that grown in desert area in Basrah.\u003c/p\u003e"},{"header":"2- Materials and methods","content":"\u003cp\u003e2.1. Cultivation and treatments\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe verity cauliflower hybrid F1 Alnahar seeds from Monarch seeds, Noord-Scharwoude, Holland, cultivated in trays with total peat moss substrate. After 40 days of cultivation, seedlings transferred to permanent field, treatment with biostimulants started after 30 days of transferring. Biostimulants treatments were performed weakly with LRE and BYE at three concentrations (0, 10, 20) g/L, the period between each treatment was 2 weeks. \u0026nbsp;\u003c/p\u003e\n\u003cp\u003e2.2. Biochemical traits of curds\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe cruds of treated plants were dried at room temperature and used to estimate the content of carbohydrates, protein, phenols, flavonoids, and elements as well as their active constituent’s profile\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e2.2.1. Carbohydrates content\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eCarbohydrates content was determined by Phenol-Sulphuric Acid method according to (DuBois et al., 1956). From dried samples 0.5 g homogenized with 70 ml of distilled water at 70 °C for 1 hr. Then, 5 ml of filtrated extract placed in a volumetric flask and made up to 30 ml by distilled water. From diluted extract, 1 ml placed in a new tube and homogenized with 1m of 5% phenol and 5 ml of pure sulfuric acid by vortex. After brining the mixture to the room temperature, the absorption recorded by spectrophotometer at 490 nm and carbohydrates concentration was valuated from glucose standard curve and expressed as mg of glucose equivalents per gram of sample dry weight (mg GU/g dw).\u003c/p\u003e\n\u003cp\u003e2.2.2. Protein content\u0026nbsp;\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eProtein content was determined in dried plant samples following Kjeldahl method. From dried samples 0.20 g was digested with 5 ml of pure sulfuric acid at 350 °C for 30 minutes. After the samples cooling, 3 ml of the mixture of 96 ml sulfuric acid and 6 ml perchloric acid was added to the sample solution and placed on digestion plate at 350°C until the solution being clear. The volume of sample solution was made up to 50 ml by distilled water. From diluted sample 10 ml was mixed with 10 ml of 6 N of sodium hydroxide in the tube and placed in Kjeldahl analyzer instrument. The distillate was collected in a conical flask containing 2 % of Boric acid and 2% of bromocresol green and methyl red indicator solution (0.099 g + 0.066 g in 100 ml Ethanol). The collection was titrated with 2 % HCl until the solution has a slightly violet color, the percentage of nitrogen was calculated through the equation:\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e%\u003cimg width=\"314\" height=\"22\" src=\"data:image/png;base64,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\" alt=\"image\"\u003e\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;%N= Percentage of nitrogen\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eV= Volume of titration acid (HCl)\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eN= Normality of HCl\u003c/p\u003e\n\u003cp\u003eMW= Molecular weight of nitrogen\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eTV= Total volume of digestion solution\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eSW= Weight of plant sample\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eVS= Volume of digestion solution\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe percentage of protein was calculated following the question:\u003c/p\u003e\n\u003cp\u003e\u003cimg width=\"157\" height=\"22\" src=\"data:image/png;base64,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\" alt=\"image\"\u003e\u0026nbsp;(conversion factor)\u003c/p\u003e\n\u003cp\u003e2.2.3. Total phenols content\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eCruds content of phenols estimated by Folin–Ciocalteu method according to (Porter, 2012). With 5 ml of 80% methanol, 0.25 g of dry sample was homogenized and kept 48 hrs in dark, 24 hrs at room temperature and 24 hrs at 4 °C. To 0.5 ml of the methanolic extract, 0.5 ml of Folin–Ciocalteu reagent was added and followed with 8 ml of distilled water. After 5min of incubation, 1ml of 20% sodium carbonate (Na2CO3) was added. After 30 mins incubation at ambient temperature in dark, an intense blue color developed, and its absorbance was measured at 760 nm. Gallic acid was used as a standard by preparing stock solution at concentration 1 mg/ml and used for the calibration. The results were expressed as mg of gallic acid equivalents per gram of the dry weight sample (mg GAE/g dw)\u003c/p\u003e\n\u003cp\u003e2.2.4. Total flavonoids content\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eTotal flavonoids content in cruds of control and treated plants was evaluated following (Samidha Kamtekar, 2014). To 0.5 ml of extract that placed in 10 ml tubes, 0.5 ml of 80% methanol added and followed by the adding of 10 ml of distilled water and 0.3 ml of 5% sodium nitrite (NaNO3). After incubation for 5 mins, 0.3 ml of 10% aluminum chloride (AlCl3) and 2 ml of 1M sodium hydroxide (NaOH) were added. The volume was made up to 10 ml with distilled water and vortexed, the absorbance was recorded spectrophotometry at 510 nm after orange-yellowish color development. Quercetin was used as a standard by preparing stock solution at concentration 1 mg/ml and used for the calibration. The results were expressed as mg of quercetin equivalent per gram of the dry weight sample (mg QE/g dw) . \u0026nbsp;\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e2.2.4. Elements composition\u0026nbsp;\u003c/p\u003e\n\u003cp\u003ePotassium, phosphors, and sulfur were determined by the dry ash method (Akinyele \u0026amp; Shokunbi, 2015). In porcelain crucibles, 1 g of cruds samples were placed and incinerated at an initial temperature of 200\u003csup\u003e◦\u003c/sup\u003eC for 1h on a hot plate until the smoke disappears. Then, samples kept in Nabertherm muffle furnace at 556 °C for 5 hrs for total incineration. Next, ashes were dissolved by a mixture of 4 ml of nitric acid (65%) and 10 ml of distilled water on a hot plate. The solution was filtered and made up to 25ml with distilled water and used for elements (P, K , S) analysis by atomic absorption/emission spectroscopy. The calibration curves were generated using the standards of each element, and the results were expressed as mg of element per gram of the dry weight sample.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e2.2.5. GC-MS analysis\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e2.2.5.1. Samples preparation\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eFrom dried cruds, 0.250 g homogenized with 5 ml of 80% methanol, vortexed to achieve a good mixing and incubated in dark at room temperature overnight. Next day, supernatant collected carefully by micropipette and sent for injection in GCMC instrument\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e2.2.5.2 GCMC conditions\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eGC-MS analysis was carried out by using an Agilent Technologies, 7890B GC system coupled to an Agilent Technologies 5977A MSD with EI Signal detector, using HP-5ms 5% phenyl, 95% methyl siloxane (30m*250um*0.25). The oven temperature was set at 40 °C hold for 5 mins then raised to 8 °C/min to 300 °C for 20 mins, Helium carrier gas flow rate was 1 ml/min and purge flow of 3 ml/min. The injection mode was pulsed Splitless with injection temperature 290 °C and the injection sample volume was 0.5 µl. The mass spectrometer used Ion Source Temperature 230 °C, with scan speed 1562(N2), and the mass range 44-750 m/z. Compound identification was implemented using mass spectral data and retention indices, in comparison with authentic standards and spectral libraries, including the NIST 2014, 2020 database.\u003c/p\u003e"},{"header":"3- Results","content":"\u003cp\u003eFigure 1 illustrates the effect of biostimulants spray on curds content of carbohydrates and proteins, the spray with LBE and BYE at different concentration caused reduce the carbohydrates content of curds compared to the control plants, the range of values between 19.32 mg GU/g dw and 22.86 mg GU/g dw. On the other hand, protein proportion increased remarkably in curds of 10 g/L of BYE-treated plants (15.89 %) and plants that sprayed with 20 g/L of LRE and 10 g/L of BYE combination (14.87 %) (Fig.1A\u0026amp;B).\u003c/p\u003e\n\u003cp\u003eTotal phenolic content and total flavonoids content in curds exhibited non-significant increase due to foliar application of both extracts on Brassica plants. The increase of phenols content was in curds of 20 g/L of BYE-treated plants (17.03 mg GAE/g dw), while flavonoids content increased in curds of combination of 10 g/L of LRE and 20 g/L of BYE-treated plant (16.54 mg QE/g dw) (Fig. 2A\u0026amp;B). Antioxidants activity of curds extracts against 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals reinforced in curds methanolic extract of LRE-sprayed plants and BYE-sprayed plants and their combination-sprayed plants recorded 75.00 %, 77.59 %, and 74.09 % respectively (Fig. 2C).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eElements composition including nitrogen, phosphors, calcium and sulfur displayed the effect of foliar application of biostimulants on macro elements content in Brassica curds. Nitrogen portion increased in curds of 10 g/L BYE-treated plants and 20 g/L LRE and 10 g/L BYE-treated plants (2.54, 2.38) % (Fig. 3A). Phosphor content was impacted with individual and combination applications of LRE and BYE, P content promoted in curds of 10 g/L of LRE and BYE-treated plants (5.67 mg/g), 20 g/L LRE-treated plants (6.00 mg/g) and 20 g/L of LRE and BYE-treated plants (6.10 m/g) of dry weight (Fig. 3B). Furthermore, potassium content increased subsequently the foliar spray with LRE and BYE. Curds of 20 g/L BRE-treated plants (28.83 mg/g), 10 g/L of LRE-treated plants (23.31 mg/g) and the combination 10 g/L of LRE and 20 g/L of BRE-treated plants (28.52 mg/g) of dry weight (Fig. 3C). Sulfur content in Brassica curds was boosted in curds of plants that treated with 10 g/L of LRE and the combination of 20 g/L of LRE and 10 g/L of BRE that both recorded 3. 60 mg/g of dry weight (Fig. 3D).\u003cbr\u003eGC-MS profile of methanolic Brassica curds extract of treated plants represents alteration in the active constituents compared to control plants (Tab. 1). The alteration may involve increase or decrease or disappearance of detected compounds area percentage according to applied treatments. For example, dimethyl trisulfide (compound is responsible for off-flavour in cooked Brassica vegetable) increased in curds of 10 g/L-treated plants and plants that sprayed with combination of 10 g/L and 20 g/L of LRE and BYE respectively. While compounds like phenol decreased in curds of sprayed plants with both biostimulants comparing to untreated plants. Saturated fatty acid Hexadecanoic acid and polyunsaturated fatty acid 9,12,15-Octadecatrienoic acid increased substantially in plants curds due to foliar application with biostimulants particularly with combination applications. Methyl ester compound such as Methyl 9-cis,11-trans-octadecadienoate was inexistent in untreated plants and gradually increasing with individual or combinated applications according to GC-MS profile. However, several compounds such as 4-Ethoxy-2-(methylamino) tropon, Furan-2-carboxaldehyde, 5-(1-piperidyl)- and gamma.-Sitosterol vanished in methanolic extract of Brassica curds due to influence of individual or combination applied treatments.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eTable (1): GC-MS analysis profile of Brassica methanolic extract\u003c/p\u003e\n\u003cdiv align=\"center\"\u003e\n \u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"798\" class=\"fr-table-selection-hover\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 42px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eR.T.\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 180px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eCompound name\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 120px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eChemical class\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eL0B0\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eL0B1\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eL0B2\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eL1B0\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eL1B1\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eL1B2\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eL2B0\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eL2B1\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eL2B2\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 42px;\"\u003e\n \u003cp\u003e9.26\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 180px;\"\u003e\n \u003cp\u003eDimethyl trisulfide\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 120px;\"\u003e\n \u003cp\u003eOrganosulfur\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e3.68\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e3.58\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e3.13\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e4.06\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e2.87\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e4.98\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e2.81\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e2.79\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e2.89\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 42px;\"\u003e\n \u003cp\u003e18.43\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 180px;\"\u003e\n \u003cp\u003ePhenol, 4-ethenyl-2,6-dimethoxy-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 120px;\"\u003e\n \u003cp\u003ePhenolic compounds\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e1.47\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e0.72\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e0.75\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e1.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e0.83\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e0.60\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e0.52\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e0.58\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e0.64\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 42px;\"\u003e\n \u003cp\u003e19.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 180px;\"\u003e\n \u003cp\u003e4-Ethoxy-2-(methylamino) tropone\u003c/p\u003e\n \u003cp\u003eFuran-2-carboxaldehyde, 5-(1-piperidyl)-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 120px;\"\u003e\n \u003cp\u003eTroponoid\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003cp\u003eVolatile compound\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e8.48\u003c/p\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e5.79\u003c/p\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e6.16\u003c/p\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e7.23\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e6.37\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e4.10\u003c/p\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e5.29\u003c/p\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e5.84\u003c/p\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e6.54\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 42px;\"\u003e\n \u003cp\u003e20.72\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 180px;\"\u003e\n \u003cp\u003ed-Proline\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 120px;\"\u003e\n \u003cp\u003eAmino acid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e0.97\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e1.46\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e1.55\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e2.19\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e1.86\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e3.31\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e2.69\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 42px;\"\u003e\n \u003cp\u003e21.26\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 180px;\"\u003e\n \u003cp\u003e1H-Indole-3-acetonitrile\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 120px;\"\u003e\n \u003cp\u003eGlycosides\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e1.65\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e1.53\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e1.55\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e1.53\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e1.68\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e1.83\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e1.15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e1.10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e1.03\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 42px;\"\u003e\n \u003cp\u003e22.19\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 180px;\"\u003e\n \u003cp\u003eHexadecanoic acid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 120px;\"\u003e\n \u003cp\u003eSat. Fatty acid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e1.23\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e3.26\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e2.67\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e2.27\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e4.21\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e5.07\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e4.59\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e7.44\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e4.43\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 42px;\"\u003e\n \u003cp\u003e23.82\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 180px;\"\u003e\n \u003cp\u003eMethyl 9-cis,11-trans-octadecadienoate\u003c/p\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 120px;\"\u003e\n \u003cp\u003e\u0026nbsp;Methyl ester\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e2.59\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e2.04\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e1.56\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e2.032\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e4.65\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e3.18\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e4.73\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e2.89\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 42px;\"\u003e\n \u003cp\u003e23.89\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 180px;\"\u003e\n \u003cp\u003e9,12,15-Octadecatrienoic acid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 120px;\"\u003e\n \u003cp\u003eUnsat. Fatty acid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e1.24\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e10.12\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e9.35\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e7.42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e10.83\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e21.10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e13.11\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e15.30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e9.99\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 42px;\"\u003e\n \u003cp\u003e27.33\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 180px;\"\u003e\n \u003cp\u003eGlycerol monopalmitate\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 120px;\"\u003e\n \u003cp\u003eFatty acid\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e1.33\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e1.18\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e1.51\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e1.57\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e2.18\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e1.38\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e2.18\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e1.73\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e0.86\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 42px;\"\u003e\n \u003cp\u003e33.29\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 180px;\"\u003e\n \u003cp\u003e.gamma.-Sitosterol\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 120px;\"\u003e\n \u003cp\u003eSterol\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e0.45\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e0.51\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e0.76\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e2.11\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 48px;\"\u003e\n \u003cp\u003e1.93\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e2.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 54px;\"\u003e\n \u003cp\u003e1.53\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003c/table\u003e\n\u003c/div\u003e"},{"header":"4- Discussion","content":"\u003cp\u003eThe utilization of botanical extract as biofertilizer or biostimulants to enhance plant growth or plant tolerance to stress conditions has gained considerable attention in recent years. Obtained results showed that carbohydrates content in curds decreased when foliar spray applied on Brassica plants which could be attributed to exploiting carbohydrates to increase the growth parameters of sprayed plants including plant high, leaves number, leaf area and biomass (data not shown). Furthermore, total carbohydrates were utilized also by treated plants to increase the productivity traits such as yield, diameter, biomass, and weight of Brassica cruds (data not shown). Under similar conditions, instead of carbohydrates accumulation in induvial organs, plant direct total carbohydrates to promote its growth and yield (Van den Ende, \u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). On the other hand, protein percentage exhibited an increase in curds of L0B1-treated plants and L2B1-treated plants. The role of protein in the plant cover expanded ambit through their structural, enzymatic, metabolic, regulatic and storage functions (Rasheed et al., \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Foliar application with biostimulants that are rich with protein such as BYE (Demirgul et al., \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2022\u003c/span\u003e) or amino acid such as LRY (Wang et al., \u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e2015\u003c/span\u003e) or both could increase the curds content of protein and promote yield quality from the healthy components.\u003c/p\u003e\u003cp\u003eBrassica curds are good source for phenolic compounds particularly flavonoids (F. A. Ahmed \u0026amp; R. F. Ali, 2013). Phenols including flavonoids are natural compounds that are synthesized by the phenylpropanoid pathway and widely distributed in plants to promote and maintain its growth and development. Besides, they have several medical benefits such as anticancer, antiinflammation and antioxidant properties (Kuljarusnont et al., \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). In our study, the total of phenols and flavonoids in curds have not shown significantly affected due to the foliar application with LRY or BYE. However, an insignificant increase was observed in phenols and flavonoids content in curds of L0B2-treated plants and L1B2-treated plants respectively. This increase may be related to the role of LRY and BYE in boosting and stimulating secondary metabolites production including total phenols and flavonoids, conforming with obtained outcomes by Younes in \u003cem\u003eAllium cepa\u003c/em\u003e (Younes et al., \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e2021\u003c/span\u003e) and Dawood in \u003cem\u003eGlycine max\u003c/em\u003e (Dawood et al., \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2013\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eIn the antioxidant context, previous literature indicated the active constituents in Brassicaceae vegetables and their related antioxidant properties (Mulțescu et al., \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). The antioxidants activity in Brassica curds is correlated to various compounds such as phenols, flavonoids, sulfurs compounds, glucosinolates and others. This activity may be influenced extremely by several factors including cultivar, maturity, growing and harvesting conditions and storge circumstances (Podsędek, \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e2007\u003c/span\u003e). Following the effect of LRE and BYE on antioxidant activity of curds methanol extract, L0B2-treated plant and L1B0-treated plants recorded a significant activity. Likewise, curds extract of plants sprayed with combination of both LRY and BYE concentrations recorded also antioxidant activity against DPPH. The augmentation of DPPH scavenging efficiency due to LRE implementing was observed previously in the extract of \u003cem\u003eStevia rebaudiana bertoni\u003c/em\u003e under normal conditions (Khashan \u0026amp; AL-Taweel, \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2024\u003c/span\u003e) and \u003cem\u003ePhaseolus vulgaris\u003c/em\u003e L. under salinity conditions (Rady et al., \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2019\u003c/span\u003e), both found LRE promoted antioxidant activity in plants due to increase the non-enzymatic antioxidant (e.g. proline, phenols, tocopherols, carotenoids, etc). In the current study, the capacity of curds extract to scavenging DPPH radicals could be increased regarding the increase of Dimethyl trisulfide, d-Proline, 9,12,15-Octadecatrienoic acid and gamma.-Sitosterol (Table\u0026nbsp;1). Likewise, BYE application boosted antioxidants properties in \u003cem\u003eAllium sativum\u003c/em\u003e L. as a tolerance strategy against drought stress through increasing of antioxidant enzymes and antioxidants compounds such as proline and ascorbic acid (Abdelaal et al., \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Our result indicated that BYE enhanced the scavenging of free radicals in curds extract of treated plants which could be imputed to induced role of BY in promoting compounds with antioxidants features including Dimethyl trisulfide, d-Proline, 9,12,15-Octadecatrienoic acid, Methyl 9-cis,11-trans-octadecadienoate, Glycerol monopalmitate and gamma.-Sitosterol. These compounds increased also in curds extract of sprayed plants with combination of LRE and BYE according to GC-MS analysis (Table\u0026nbsp;1) which consequently enhanced its antioxidants efficiency.\u003c/p\u003e\u003cp\u003eMicroelements (N, P, K, S) increased compatibly with biostimulants foliar application. Brassica curds have been reported to contain a high macro elements content (Ekholm et al., \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2007\u003c/span\u003e). However, cultivars, growing and environmental conditions, and harvesting and post harvesting circumstances influence curds content of macro and micro elements (Kalisz et al., \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Kisko et al., \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). In a present study potassium recorded the highest content in Brassica curds and the content increased remarkably with LRE and BRE at L0B2 and L1B2-treated plants. Likewise, other macronutrients including N, P, and S increased in curds of treated plants: for N at L0B1 and L2B1, for P at L1B1, L2B0, and L2B1, and for S at L1B0 and L2B1. The increase of macro elements in curds may be referred to a rich content of elements in LRE (Gormez \u0026amp; Sengul, \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2014\u003c/span\u003e) and BRE (Tao et al., \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e2023\u003c/span\u003e), these elements either move through or interact with the apoplast space which subsequently effect on their translocation to different plant parts including the curds (Fern\u0026aacute;ndez \u0026amp; Brown, \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2013\u003c/span\u003e). Additionally, this increase may be linked to enhanced root capacity for nutrients uptake from the soil post biostimulants application (Halpern et al., \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). It has been reported that LRE and BYE increased minerals content in mango fruit and potato tuber when biostimulants applied on the plants (Orj\u0026aacute;n et al., \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Yousif et al., \u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). Macro minerals almost participate in all human body functions and act significantly in metabolic processes. For example, phosphorus serve important role in the bone, teeth, phosphoproteins and nucleic acids formation, fat and starch metabolism and heartbeat and kidney function normalization (Elser, \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2012\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eGC-MS is mostly favoured technique to identify the quality and quantity of compounds in complex mixture due to its sensitivity and selectivity. Numerous agents influence GC-MS analysis results, some are related to plants (plants species, growing conditions, extraction methods) and others are related to GC-MS instrumentation conditions (temperature program, injection technique, detector sensitivity). Under our experimental conditions, GC-MS result of curds methanolic extract indicated high alteration in the peak area of most important detected compounds. Several compounds increased due to LRE or BYE or both applications such as Dimethyl trisulfide (L1B0, L1B2), d-Proline (L0B1, L0B2, L1B0, L1B1, L2B0, L2B1), 1H-Indole-3-acetonitrile (L1B1, L1B2), Hexadecanoic acid (all treatments), Methyl 9-cis,11-trans-octadecadienoate (all treatments), Glycerol monopalmitate (all treatment except L2B2) and gamma.-Sitosterol (L0B1, L0B2, L1B0, L2B0, L2B1, L2B2). On the contrary, several compounds decreased due to individual or combination of foliar applications such as Dimethyl trisulfide (L0B1, L0B2, L1B0, L1B1, L2B0, L2B1, L2B2), Phenol, 4-ethenyl-2,6-dimethoxy- (all treatments), 4-Ethoxy-2-(methylamino) tropone (L0B1, L0B2, L1B2, L2B0, L2B1), 1H-Indole-3-acetonitrile (L0B1, L0B2, L1B0, L2B0, L2B1, L2B2), and Glycerol monopalmitate (L2B2). Interestingly, some compounds disappeared due to LRE or BYE or both applications such as 4-Ethoxy-2-(methylamino) tropone (L1B0, L1B1, L2B2), Furan-2-carboxaldehyde, 5-(1-piperidyl)- (L0B0, L0B1, L0B2, L1B2, L2B0, L2B1), d-Proline (L1B2, L2B2), and gamma.-Sitosterol (L1B1, L1B2). The alteration of active compounds in Brassica curds extract with LRE foliar spray may be correlated to rich content of LRE from sugars, vitamins, mineral nutrients, amino acids, selenium, and phytohormones which effect on secondary metabolites biosynthesis pathways such as sulfurs-containing compounds pathway (Nakai \u0026amp; Maruyama-Nakashita, \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2020\u003c/span\u003e), shikimate/phenylpropanoid pathway (Tak et al., \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e2023\u003c/span\u003e), hormone biosynthesis pathways (Bajguz \u0026amp; Piotrowska-Niczyporuk, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2023\u003c/span\u003e), and sterol biosynthesis pathway (Du et al., \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Likewise, BYE application caused a variation in peak areas of secondary compounds in Brassica curds extract which may be referred to the BYE content of effective components including amino acids, vitamins, proteins and hormones (Medani \u0026amp; Taha, \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). Some of these components serve as precursors for secondary metabolites biosynthesis pathways such as amino acids (Trovato et al., \u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e2021\u003c/span\u003e), while some act as enzymes cofactors, signaling molecules, and genes expression regulators such as vitamins and hormones (Parra et al., \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Rui et al., \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). These active compounds in Brassica curds have crucial role in maintaining human health when incorporated into the daily dietary due to their therapy functions as antioxidants, antiinflammation, anticancer, antiviral, anti-obesities, antihypertensive, and antigenotoxicity (Zhang et al., \u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e2025\u003c/span\u003e). For example, 1H-Indole-3-acetonitrile displayed a broad-spectrum antiviral potential against influenza A, HSV-1, VSV and SARS-CoV-2 (Hui et al., \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Zhao et al., \u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Thus, the increase of their amount after LRE and BYE application may enhance the nutritional, health-promoting and therapeutic value of brassica curds.\u003c/p\u003e"},{"header":"5- Conclusion","content":"\u003cp\u003eIn summary, we found that the combination of biostimulants including LRE and BYE could be affected prominently on metabolites content of Brassica curds. When two biostimulants are applied as combination, their act may complement each other or act synergistically to enhance the overall effect, resulting in improving plants yield quantity and quality in the region that has desert soil properties. Our results indicated an increase in primary and secondary metabolites in Brassica curds after foliar application of LRE and BYE concentrations. Although the induvial treatments had an obvious impact on experimental parameters, the combination of study agents also acted synergistically to boost these parameters. In crops cultivation under desert conditions, obtained findings could be considered a promising result within the sustainable required direction. The remarkable function of LRE and BYE in improving the quality components of Brassica curds including phenols, flavonoids, macronutrients, and active compounds provides compelling evidence of the alternative role that biostimulants may play in eco-friendly agriculture. This improvement not only increase the nutritional value of Brassica curds but also enrich them with active compounds that contribute to the promoting of healthy foods sources.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAuthor contribution\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eZ.A designed research studies and performed field work, W.N and F.H performed lab experiments and data analysis. W.N and F.A wrote/edited the manuscript. All authors have read and approved the final version of the manuscript\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDeclarations\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eEthical approval and consent to participate\u003c/p\u003e\n\u003cp\u003eNot applicable\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eConsent for publication\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e\n\u003cp\u003eFunding\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe research was not supported by any funding\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eclinical trial\u003c/p\u003e\n\u003cp\u003eNot applicable\u0026nbsp;\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eAbdelaal, K., Attia, K. A., Niedbała, G., Wojciechowski, T., Hafez, Y., Alamery, S., Alateeq, T. K., \u0026amp; Arafa, S. A. (2021). 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Brassica vegetables-an undervalued nutritional goldmine. \u003cem\u003eHortic Res\u003c/em\u003e,\u003cem\u003e 12\u003c/em\u003e(2), uhae302. https://doi.org/10.1093/hr/uhae302\u003c/li\u003e\n\u003cli\u003eZhao, X., Zhao, L., Zhao, Y., Huang, K., Gong, W., Yang, Y., Zhao, L., Xia, X., Li, Z., \u0026amp; Sheng, F. (2021). 3-Indoleacetonitrile is highly effective in treating influenza a virus infection in vitro and in vivo. \u003cem\u003eViruses\u003c/em\u003e,\u003cem\u003e 13\u003c/em\u003e(8), 1433.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"bmc-plant-biology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"pbio","sideBox":"Learn more about [BMC Plant Biology](http://bmcplantbiol.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/pbio/default.aspx","title":"BMC Plant Biology","twitterHandle":"BMC_series","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Biostimulants, Liquorice root extract, Bread yeast extract, Cauliflower curds, Metabolites. Desert region ","lastPublishedDoi":"10.21203/rs.3.rs-7614364/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7614364/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eAmong Brassica vegetables, cauliflower (\u003cem\u003eBrassica oleracea\u003c/em\u003e var. \u003cem\u003ebotrytis\u003c/em\u003e) is widely recognized as a consumer favorite due to its rich and healthy content of nutrients and bioactive compounds. The enhancement of productivity and quality of preferred crops has gained highly importance in recent decades, driven by increasing customers demand for affordable and healthy food. Away from chemical materials and their risky side effects, biostimulants and biofertilizers could be a suitable alternative for traditional fertilizers under desert conditions. In a current study the extract of Liquorice root LRE and bread yeast BYE applied both with 0, 10, and 20 g/L on Brassica plant to explore their influence in curds metabolites. Obtained results showed that while carbohydrates reduced in curds of treated plants, protein increased with both stimulants application. The non-significant increasing in phenols and flavonoids content was observed in curds of LRE and BYE-treated plants with an elevation in antioxidant activity in their methanolic extracts. Both LRE and BYE applications promoted the curds content of macronutrients and caused a notable alteration in active compounds profile. Noteworthy, the values of acquired outcomes increased or decreased according to applied concentrations of individual or combination of both biostimulants. Based on our finding, the foliar application with LRE and BYE enhanced the primary and secondary metabolites of Brassica curds. However, the combination of 10 g/L of LRE and 20 g/L of BYE is recommended due to its act in promoting of flavonoids content, antioxidant capacity, potassium content and active compounds profile including organosulfur, troponoid, methyl 9-cis,11-trans-octadecadienoate, 1H-Indole-3-acetonitrile and 9,12,15-Octadecatrienoic acid.\u003c/p\u003e","manuscriptTitle":"Biostimulants combination enhance the metabolites content of Brassica oleracea var. botrytis curds cultivated in the desert region of Basrah governorate","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-10-01 10:58:11","doi":"10.21203/rs.3.rs-7614364/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2025-10-15T17:19:50+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-15T17:01:47+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-15T05:21:53+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"313484877094632807148147309029102417277","date":"2025-10-13T06:41:53+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"152644901131244696442902658598785703159","date":"2025-10-10T16:41:59+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"202647097519640137527402710455072187077","date":"2025-10-10T02:50:14+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"24762148199311460054775016986841810403","date":"2025-10-08T19:01:39+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-09-20T10:38:43+00:00","index":"","fulltext":""},{"type":"editorInvited","content":"","date":"2025-09-18T10:29:59+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-09-16T15:01:33+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-09-16T15:00:36+00:00","index":"","fulltext":""},{"type":"submitted","content":"BMC Plant Biology","date":"2025-09-14T19:03:47+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"bmc-plant-biology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"pbio","sideBox":"Learn more about [BMC Plant Biology](http://bmcplantbiol.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/pbio/default.aspx","title":"BMC Plant Biology","twitterHandle":"BMC_series","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"95653ce0-e9b8-418b-9957-f38ea3e4dbe7","owner":[],"postedDate":"October 1st, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2026-03-09T16:14:14+00:00","versionOfRecord":{"articleIdentity":"rs-7614364","link":"https://doi.org/10.1186/s12870-026-08237-y","journal":{"identity":"bmc-plant-biology","isVorOnly":false,"title":"BMC Plant Biology"},"publishedOn":"2026-03-04 15:59:53","publishedOnDateReadable":"March 4th, 2026"},"versionCreatedAt":"2025-10-01 10:58:11","video":"","vorDoi":"10.1186/s12870-026-08237-y","vorDoiUrl":"https://doi.org/10.1186/s12870-026-08237-y","workflowStages":[]},"version":"v1","identity":"rs-7614364","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7614364","identity":"rs-7614364","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}