Preliminary study on the function of natural persimmon frost fungi

Preliminary study on the function of natural persimmon frost fungi | 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 Article Preliminary study on the function of natural persimmon frost fungi Xiaoyi Gao, Rongfan Xu, Yiran Yan, Xiaodong Sun This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6276251/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract China is the largest persimmon producer, with the largest cultivation area and yield in the world. These rich persimmons, in addition to direct consumption, can also be made into dried persimmon. Dried persimmon is a traditional Chinese processed food which preserves the nutrition of persimmons. Persimmon frost, icing sugar on the surface of dried persimmon, is formed by the diffusion of sugar inside the dried persimmon to the surface, which has the effect of moistening lungs and relieving cough. During the long and open process of persimmon frost formation, there are a lot of airborne fungi falling freely on the surface of persimmon frost, but there are a few reports on persimmon frost fungi so far. In this study, eight fungal strains, including three strains of Aureobasidium pullulans , Filobasidium magnum , Epicoccum nigrum , Aspergillus versicolor , Metschnikowia pulcherrima and Papiliotrema flavescens , were isolated from dried persimmon samples freshly prepared by natural drying method. Sucrose was used as carbon source for single strain fermentation. The fermentation broths were passed through the dialysis bag (retaining a relative molecular weight of 3.5KD), and the polysaccharides were detected inside the bags using the concentrated sulfuric acid-phenol method. The results showed that the polysaccharides were detected out in all the fermentation broths, and the concentrations of the three Aureobasidium pullulans strains were significantly higher than those of the other fungi. The filtrate outside the dialysis bag was detected for ten monosaccharides and sugar acids by mass spectrometry detection, including glucose, galacturonic acid, rhamnose, fucose, mannose, galactose, glucuronic acid, ribose, arabinose and xylose. There are great differences in the compositions and concentrations of the monosaccharides and sugar acids in the fermentation broths of the 8 strains, and glucose is the most important monosaccharide. Galacturonic acid, rhamnose and fucose were not detected in the fermentation broth of the 8 strains, and the concentrations of mannose, galactose, glucuronic acid, ribose, arabinose and xylose were very different. It can be concluded that these fungi participate in the transformation of sugars and the formation of polysaccharides, which may influence and change the composition and functions of persimmon frost. This research provides a new field for screening fungi that produce extracellular polysaccharides and developing new functional polysaccharides. Biological sciences/Microbiology Biological sciences/Molecular biology Natural persimmon frost Filamentous fungi Yeast Monosaccharide Polysaccharide Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Introduction As the world’s largest producer of persimmons, China ranks the first worldwide in terms of the cultivation area and yield of persimmons 1 Apart from direct consumption, persimmons can also be made into dried persimmons. There are currently two techniques of processing dried persimmons in China, i.e. natural drying and artificial drying. As the traditional processing technology, the natural drying method has a long history and is still being used in many places nowadays. Figure 1 shows the main process of the natural drying method. It involves a long production cycle about 30–50 days 2 During this process, the sugars in the persimmon gradually release, and dry white crystals gradually form on the surface of the persimmon as the water evaporates, which being referred to as the persimmon frost. Having been regarded as the essence of dried persimmons ever since ancient times, persimmon frost plays an effective role in treating oral ulcer and acute pharyngitis. However, there are currently few researches on persimmon frost in the world, and the very few researches available are limited to China. These researches mainly focus on the ingredients, formation and effects of persimmon frosMa et al. t preliminarily extracted persimmon frost with organic solvents such as petroleum ether and 95% ethanol, and the sugars detected in the persimmon frost included glucose, fructose, sucrose, xylose and a trace of mannose 3 . By measuring the content of protein, fat, total acid and total sugar in persimmon frost, Zhang (2002) found that total sugar accounted for 95% in persimmon frost, among which polysaccharide took up for 7%, sucrose accounted for 6%, glucose 37%, and fructose 45% 4 Zhang and Huang (2009) found that reducing sugars accounted for 58.21% of the total weight of persimmon frost, and these reducing sugars were glucose, fructose, a trace of rhamnose and arabinose, without galactose and xylose 5 . During the long-term formation of persimmon frost, many fungi in the air will inevitably accumulate on the surface. By isolating the fungi in persimmon frost, Wang (2010) obtained various species of fungi such as Penicillium citrinum , P. chrysogenum , P. oxalicum , Aspergillus flavus and A. niger. Bai (2017) separated Penicillium chrysogenum and Aspergillus niger from persimmon frost, and relevant analysis suggested that they were the contaminating strains that caused the dried persimmon to discolor and rot 6 , 7 . Eight strains, including three strains of Aureobasidium pullulans , Filobasidium magnum , Epicoccum nigrum , Aspergillus versicolor , Metschnikowia pulcherrima and Papiliotrema flavescens , were isolated from dried persimmon samples freshly prepared by natural drying method. Unlike previous studies which only obtained Penicillium and Aspergillus species from persimmon frost, these strains of fungus were isolated from persimmon frost for the first time. The surface of persimmon frost is rich in sugar. Are these persimmon frost fungi involved in the conversion of sugars in persimmon frost? Is it possible to synthesize polysaccharides using the sugars on the surface of persimmon frost? At present, there is no relevant report. In order to clarify the function of persimmon frost fungi, the change of sugars in the fermentation broth before and after fermentation were studied by means of single strain fermentation and mass spectrometry detection. The results of this study provide a powerful reference for evaluating the ability of sugar conversion and polysaccharide synthesis of persimmon frost fungi. Materials and Methods Activation of strains The strains were inoculated into PDA plate for activation. Primary and secondary fermentation The strains were inoculated into 100 mL of the primary fermentation broth to be cultured in the dark for 96h, with the temperature of 18℃ and the rotating speed being 140r/min. The primary broth, with an inoculated dose of 2%, was transferred into a conical flask equipped with 100 mL of re-fermentation medium to be cultured for 7d with the temperature of 18°C and the rotating speed being 140r/min. All strains were repeated three times. The primary fermentation medium (seed medium) is composed of 50.0 g of sucrose, 4.0 g of K 2 HPO 4 , 0.8g of (NH 4 ) 2 SO 4 , 0.2g of MgSO 4 , 2.0g of NaCl and 1.5 g of yeast powder (pH = 7.0). The medium was sterilized at 121°C for 20min 8 . The re-fermentation medium consists of 50.0g of sucrose, 6.0g of K 2 HPO 4 , 0.6g of (NH 4 ) 2 SO 4 , 0.5g of MgSO 4 , 4.0g of NaCl and 0.9g of yeast powder (initial pH = 6.5). The medium was sterilized at 121°C for 20min. Detection of polysaccharides The fermentation broth was centrifuged at 12000r/min for 5 min, the fungi mycelium and sediments were discarded, the fermentation broth was filtered, and the supernatant was collected. The supernatant passed through the dialysis bag (retaining a relative molecular weight of 3.5KD), and distilled water was added until the liquid in the dialysis bag reached uniformly 100 mL. The liquid outside the dialysis bag was also collected (the dialysis bag was placed in a beaker and the final volume of water in the beaker was maintained at 300 mL). The concentration of polysaccharides in the dialysis bag was detected using the concentrated sulfuric acid-phenol method 9 , 10 1 mL of fermentation broth, 1 mL of distilled water, 1 mL of 6% phenol solution and 5 mL of concentrated sulfuric acid were added to a 25 mL colorimetric tube which was then covered and shaken up. After that, the colorimetric tube experienced a boiling water bath for 15 min, then be cooled. The OD value was measured at 490 nm, with the above operation repeated on each sample for three times. Distilled water replaced the sample solution to join the reaction system as the blank control. In this study, as a standard stock solution, the 1mg/mL glucose solution was diluted to a standard solution with a concentration of 0, 0.02, 0.04, 0.08, 0.1, and 0.15 mg/mL, respectively. With the reaction operated per the above steps, the absorbance was measured at 490 nm. A standard curve was plotted with glucose concentration as the abscissa and absorbance as the ordinate. By introducing the absorbance of the sample into the standard curve, the concentration of polysaccharides in the sample was calculated. Detection of monosaccharides and sugar acids Metabolites extraction 1)The liquid outside the dialysis bag were collected and 50µL liquid samples or standard solution were taken for derivative reaction. Derivatization: add 100 µL methanol solution of 1-phenyl-3-methyl-5-pyrazolone (10 mg / mL) and 30 µL of 0.1 mol / L NaOH solution to the sample (or equivalent volume of standard solution), and then react at 70 ℃ for 30 min. After cooling to room temperature, 30 µL of 0.1 mol / L HCl solution was added. After concentration and drying, 600 µL of water was redissolved, and 1 mL of chloroform was used to extract three times to remove the excessive derivatization reagent. 2)After the reaction, the mixture was dried and dissolved with 200 L methanol. The supernatant was taken for determination. Preparation of standard solution Prepare 10 mmol/L standard stock solution. Take the corresponding amount of standard stock solution into a 10 mL volumetric flask and prepare the mixed standard solution. Dilute the standard solution in turn to obtain a series of calibration solutions. See Table 1 for details of each standard substance. Table 1 Specific information about the standard substances Compound name Molecular Weight Manufacturer Purity D-Mannose 180.16 TCI, China ≥ 98.0% D-Galactose 180.16 Yuanye, China ≥ 98.0% D-Glucose 180.16 DR.E, Germany ≥ 99.7% D-Glucuronic Acid 194.14 TRC, Canada ≥ 98.0% Alpha-D-Galacturonic Acid Hydrate 212.16 FLUKA, Switzerland ≥ 97.0% L-Rhamnose monohydrate 182.17 SIGMA, America ≥ 99.0% L-Fucose 164.16 TCI, China ≥ 97.0% D-Ribose 150.13 Zhenzhun, China ≥ 98.0% DL-Arabinose 150.13 VETEC, China ≥ 99% D (+)-Xylose 150.13 TCI, China ≥ 98.0% 5-Methyl-2-phenyl-1,2-dihydropyrazol-3-one 174.20 TCI, China ≥ 98.0% Machine Detection In this study, a Waters Acquity UPLC ultra high performance liquid chromatograph was used to perform chromatographic separation of the target compounds with a Waters Acquity UPLC BEH C18 liquid chromatographic column (100 × 2.1mm, 1.7µm, Waters). The mobile phase A was 0.1% formic acid aqueous solution, and the mobile phase B was 0.1% formic acid acetonitrile solution. The column temperature was 45 ℃, the sample tray was set at 4℃, and the injection volume was 2 µL. The Xevo G2-XS QTOF high resolution mass spectrometer was used for full scan mass spectrometry analysis. Ion source parameters are as follows: Capillary voltage = 2000 V, Sampling cone = 40 V, Source temperature = 115℃, Desolvation temperature = 500°C, Desolvation gas = 900 L/h, Scan Range = 400 ~ 600, Scan time = 0.3 s. For each target compound, the derived parent ion was used for quantitative analysis. See Table 2 for details. Table 2 Mobile phase gradient and MS parameters Mobile phase gradient Time A (H 2 O) B (ACN) Flow 0.0 min 95% 5% 0.40 mL/min 8.0 min 75% 25% 0.40 mL/min 9.0 min 0% 100% 0.40 mL/min 11.0 min 0% 100% 0.40 mL/min 11.5 min 95% 5% 0.40 mL/min 13.0 min 95% 5% 0.40 mL/min MS parameters Compound Name RT (min) m/z Polarity D-Mannose 5.39 511.2193 Positive D-Galactose 7.31 511.2193 Positive D-Glucose 7.54 511.2193 Positive D-Glucuronic Acid 8.86 525.1985 Positive Alpha-D-Galacturonic Acid Hydrate 9.15 525.1985 Positive L-Rhamnose monohydrate 6.74 495.2244 Positive L-Fucose 9.00 495.2244 Positive D-Ribose 6.35 481.2087 Positive DL-Arabinose 7.91 481.2087 Positive D (+) - Xylose 8.40 481.2087 Positive Standard curves The standard curve of polysaccharides The standard curves of polysaccharide drawn based on the concentration and absorbance of standard glucose solution were shown in Fig. 2 , where x represents the glucose concentration and y stands for the absorbance. By substituting the absorbance of the sample into the equation, the concentrations of polysaccharide in the fermentation broth were calculated. The standard curves of monosaccharides and sugar acids The UPLC-HRMS analysis of the standard solution of various sugars was carried out using the method described above, and the standard curve was obtained. Where y represents the peak area of the target compound and x represents the concentration of the target compound. When the least square method was used for regression analysis and the weight was set as 1/x, the calibration solution recovery rate (accuracy) and correlation coefficient (R 2 ) were the best. During the detection, it was found that the glucose contents in the samples were too high. Finally, the internal standard method was used for quantitative analysis, and the internal standard was 13C 6 -glucose (ALDRICH, purity ≥ 99%). In the curve of glucose, y represents the ratio of the peak area of the target compound to the peak area of the corresponding internal standard, and x represents the concentration of the target compound. See Table 3 for details. Table 3 The standard curves of monosaccharides and sugar acids Compound Name Linear Range (µg/mL) Calibration curve R 2 D-Glucose 6.053-121.068 y = 1.31322x-2.77829 0.9997 D-Mannose 0.535-107.015 y = 39437.9x-6812.69 0.9981 D-Galactose 0.077–38.734 y = 58417.9x-2240.98 0.9982 D-Glucuronic Acid 0.670-66.978 y = 3892.19x-890.331 0.9902 Alpha-D-Galacturonic Acid Hydrate 0.115–22.983 y = 7105.2x + 29.7572 0.9992 L-Rhamnose monohydrate 0.089–44.495 y = 49763.3x-1806.84 0.986 L-Fucose 0.050-10.000 y = 101929x-2861.82 0.9991 D-Ribose 0.116–58.025 y = 54799.5x-3715.83 0.998 DL-Arabinose 0.132–26.393 y = 87997x-4172.61 0.9958 D (+)-Xylose 0.067–13.497 y = 89690.4x-4105.06 0.9985 Results The polysaccharide concentration in fermentation broth of each strain In this study, polysaccharide contents were detected in the fermentation broth of 8 strains. The polysaccharide concentrations of the three Aureobasidium pullulans were significantly higher than those of the other fungi. Epicoccum nigrum and Filobasidium magnum had lower polysaccharide concentrations. See Table 4 for details. Table 4 The contents of polysaccharides produced by the strains Strain number Species A Polysaccharide (mg/mL) SD3-1-1 Epicoccum nigrum 0.414 ± 0.017 0.075 ± 0.003 FZ2-3-5 Aspergillus versicolor 0.343 ± 0.016 *2 0.124 ± 0.006 SD3-1-2 Aureobasidium pullulans 0.366 ± 0.038 *4 0.263 ± 0.028 G3-3-1 Metschnikowia pulcherrima 0.419 ± 0.042 *2 0.151 ± 0.015 SX1-2-3 Papiliotrema flavescens 0.619 ± 0.042 0.112 ± 0.008 SD3-2-1 Filobasidium magnum 0.394 ± 0.008 0.071 ± 0.001 SD2-3-1 Aureobasidium pullulans 0.439 ± 0.008 *4 0.317 ± 0.006 SD1-2-4 Aureobasidium pullulans 0.473 ± 0.009 *3 0.256 ± 0.005 Note: A is the absorbance value of polysaccharide. The number behind the asterisk is the sample’s dilution multiple. The values in the table are average ± standard deviation (n = 3). The concentrations of monosaccharides and sugar acids 1)Liquid chromatograms of standard solutions The chromatogram of standard solutions is shown in Fig. 3 , from which it can be seen that all target compounds present symmetrical chromatographic peaks, realizing the chromatographic separation of each target compound. The peak-side notes are compounds, retention time, peak area, and concentration in order 2)The concentrations of sugars in the fermentation broth The quantitative results of monosaccharides and sugar acids in the samples are shown in Table 5 . It can be seen from the table that there are great differences in the composition of ten monosaccharides and sugar acids in the fermentation broth of the 8 strains. Glucose is the most important monosaccharide component in the fermentation broth of each strain. Among them, Filobasidium magnum SD3-2-1 had the highest glucose concentration, reaching 36316.67 ± 394.774 mg / L. The concentrations of Epicoccum nigrum SD3-1-1, Aspergillus versicolor FZ2-3-5, Papiliotrema flavescens SX1-2-3 and Aureobasidium pullulans SD2-3-1 were also relatively high. The glucose concentration of Metschnikowia pulcherrima G3-3-1 was the lowest, only 28.87 ± 7.777 mg / L. Galacturonic acid, rhamnose and fucose were not detected in the fermentation broth of the 8 strains, or their response intensity was lower than the minimum concentration of the standard curve, so they could not be quantified. The concentrations of mannose, galactose, glucuronic acid, ribose, arabinose and xylose in the fermentation broth of the eight strains were very different. Galactose and glucuronic acid were only detected in the fermentation broth of two strains SD3-1-2 and SD1-2-4 of Aureobasidium pullulans . Mannose, ribose, arabinose and xylose were found in the fermentation broth of eight strains. The contents of mannose, ribose and arabinose in the fermentation broth of different strains were quite different, while xylose was a small amount in the fermentation broth of each strain. See Figs. 4 and 5 for details. Table 5 The types and concentrations of monosaccharides in fermentation broth of each strain Sample Name Metabolite Concentration (mg/L) Glucose Mannose Galactose Glucuronic acid Galacturonic acid Rhamnose Fucose Ribose Arabinose Xylose SD3-1-2 9271.73 ± 771.053 (×40) 12.99 ± 4.222 26.34 ± 6.909 1.19 ± 0.029 N.D. N.D. <LLOQ 45.11 ± 5.345 2.20 ± 0.447 0.33 ± 0.029 SD1-2-4 3676.67 ± 324.385 (×100) 9.57 ± 0.847 10.48 ± 3.877 1.54 ± 0.156 N.D. <LLOQ <LLOQ 28.67 ± 5.761 1.19 ± 0.202 0.16 ± 0.005 SD2-3-1 19451.67 ± 763.182 (×500) 3.48 ± 1.032 N.D. <LLOQ N.D. N.D. <LLOQ 24.40 ± 3.849 0.92 ± 0.088 0.11 ± 0.010 SD3-2-1 36316.67 ± 394.774 (×500) 21.30 ± 2.557 N.D. N.D. N.D. N.D. <LLOQ 0.39 ± 0.035 0.71 ± 0.064 0.08 ± 0.001 SD3-1-1 23548.33 ± 157.056 (×500) 18.72 ± 0.721 N.D. N.D. N.D. N.D. <LLOQ 1.69 ± 0.117 0.40 ± 0.080 0.13 ± 0.014 FZ2-3-5 25816.67 ± 1710.967 (×500) 19.52 ± 0.913 N.D. N.D. N.D. N.D. <LLOQ 5.33 ± 0.575 0.30 ± 0.031 0.13 ± 0.005 G3-3-1 28.87 ± 7.777 0.28 ± 0.041 N.D. N.D. N.D. N.D. <LLOQ 1.56 ± 0.291 20.99 ± 0.957 0.07 ± 0.001 SX1-2-3 27485.00 ± 509.200 (×500) 3.69 ± 1.040 N.D. <LLOQ N.D. N.D. <LLOQ 1.15 ± 0.350 0.58 ± 0.260 0.38 ± 0.090 The target metabolite concentration in the sample is the final concentration measured by the instrument multiplied by the dilution ratio of the sample in the extraction solvent, in mg / L. Among them, glucose is diluted data and other compounds are undiluted data. "N.D." indicates that the target compound in the sample is not detected. “<LLOQ” means that the target compounds in the sample have chromatographic peaks, but their response strength is lower than the minimum concentration of the standard curve, and cannot be quantified. The values listed in the glucose column are multiples of the sample dilution. Note: The values in the table are average ± standard deviation (n = 3) Discussion Two filamentous fungi, Aspergillus versicolor and Epicoccum nigrum with polysaccharide activity were detected in the study. As common fungal groups in the air, Aspergillus are abundant in naturally fermented foods and they participate in the fermentation process of the foods. In this genus, Aspergillus oryzae and A. niger are important strains of fermentation, which play an important role in the fermentation of many foods 11 , 12 Aspergillus versicolor is also an important functional strain, and has been widely reported. As an endophytic fungus, it can produce a variety of antibacterial substances, and some strains isolated from the deep sea can metabolize to produce polysaccharides 13 – 16 Epicocum nigrum can produce yellow pigment, as well as polyketide, carotenoids and flavonoids, and these bioactive compounds are widely used in the production of functional food, medicine and cosmetics 17 However, there are few reports on the production of polysaccharides by Epicocum nigrum . Six polysaccharide-producing yeast strains were obtained in the study. It is known to all that Aureobasidium pullulans can produce extracellular polysaccharide Pullulan during fermentation, and Pullulan has been extensively used in the fields of food, pharmacy and biomedicine 18 Whether the three strains of Aureobasidium pullulans produce Pullulan or other types of polysaccharides still requires further intensive research. Metschnikowia commonly found on the surface of grapes or in naturally fermented wines and fruit juices, can produce alcohol and volatile organic compounds during metabolism. Besides, studies have shown that Metschnikowia can produce extracellular polysaccharides 19 , 20 As dimorphic basidiomycetes, Papiliotrema and Filobasidium can produce yeast-like spores in their life histories. There are few studies on the application of the two kinds of fungi and on their production of polysaccharides. These 8 strains were isolated from persimmon frost for the first time, and the polysaccharide activity was detected in all of them. This provides a new direction for screening polysaccharide producing strains and developing new polysaccharides. Sugar transformation and formation of polysaccharides by fungi Several kinds of monosaccharides and sugar acids were detected in the fermentation broth outside the dialysis bag. It was found that glucose was the main carbohydrate substance in the filtrate of each strain, and various sugars were also detected. Sucrose is the only carbohydrate in the original fermentation medium. Sucrose can be hydrolyzed into glucose under the action of the sucrase produced by the strains. Many studies have isolated and obtained the sucrase from fungi and yeast 21 – 26 Under the action of glucokinase, the produced glucose turns into glucose-6-phosphate, which is converted into the precursor nucleotide sugar of various sugars by the action of other synthetases, mutases, isomerases and other enzyme systems produced by the strains. It is the activation form of various monosaccharides and sugar acids, and the starting material of polysaccharide synthesis. Then, under the action of highly specific glycosyltransferases (GTS), the extension, reversion and polymerization of polysaccharide repeat units are carried out, and the polysaccharide output is also carried out 27 , 28 The research of polysaccharide synthesis mainly focuses on bacteria and a few medicinal fungi. Although the structure of polysaccharides varies greatly, the synthetic pathway is almost the same. In this study, polysaccharides were detected from the dialysate bags of each strain. It can be inferred that these eight strains have the function of utilizing, transforming sugars and synthesizing polysaccharides. Potential functions of fungi in persimmon frost There are few reports on the study of persimmon frost polysaccharides. Li (2010) used the ultrasonic-assisted hot water extraction method to extract crude persimmon polysaccharide on the surface of dried persimmons, and detected the antioxidative competence of persimmon polysaccharides 29 Is it rational to speculate that there are abundant fungal groups on the surface of persimmon frost and some species can produce polysaccharides using the sugar on the surface of persimmon frost? As a matter of fact, the formation of persimmon frost polysaccharide is the process of metabolism and transformation of carbohydrates with the participation of fungi. Different fungi produce various polysaccharide mixtures with different structures and functions on the surface of dried persimmon. The function and efficacy of persimmon frost are probably a result of these polysaccharides working with other beneficial ingredients in the persimmon frost. In addition, as the main carbon source, sucrose in the original fermentation broth was hydrolyzed by the strains, and many kinds of other sugars was detected in the fermentation broth, which indicating that sugar transformation occurred in the fermentation broths to form other sugars. It is speculated that the sugars on the surface of persimmon frost consists of two parts, namely, the sugars inside the persimmon that being evaporated to the surface of dried persimmon, and the sugar converted from other carbohydrates by the fungal metabolism on the surface of persimmon frost. The fungi on the surface of persimmon frost take an active part in both the formation of polysaccharides and the mutual conversion of carbohydrates, influencing and altering the composition and function of carbohydrates in the persimmon frost. Due to the traditional methods of separation, fermentation and detection, there will be some growth and metabolic limitations of these fungi, as well as errors caused by human factors. However, the results can roughly reveal the ability of multiple fungi on persimmon frost utilizing sugars in persimmon frost and generating polysaccharides. Therefore, not all fungi in the persimmon frost are contaminating species, and many of them are functional strains that can produce polysaccharides. Due to the unique macromolecular structure and obvious biological activity of polysaccharides, we should focus on these new polysaccharides in future researches. Conclusion Carbohydrates not only provide the human body with necessary energy, but are also equipped with a variety of biological activities. Researches on carbohydrates have attracted extensive attention in recent years, of which the researches on polysaccharides are the most popular. In the current stage, to screen polysaccharide-producing strains and develop new functional polysaccharides, attempts have been made to separate microorganisms that can metabolize polysaccharides from different habitats such as oceans, soils, plants and extreme environments. Most of these microorganisms are bacteria, but there are also a small amount of fungi. Dried persimmon is a food with Chinese characteristics. As the essence on the surface of dried persimmon, persimmon frost contains rich carbohydrates that provide a favorable carbon source for the growth and metabolism of fungi. In this study, eight strains were isolated from dried persimmon samples. All of them are capable of transforming carbohydrates and producing polysaccharides. With persimmon frost as the substrate, these fungi could change the composition and function of carbohydrates in persimmon frost. This study took the initiative to explore the fungi in persimmon frost, the results of which provide a new orientation for screening fungi that produce extracellular polysaccharides and developing new functional polysaccharides. Declarations Declaration of Competing Interest The authors have declared no conflict of interest. Acknowledgements This research was supported by the funds of Fundamental Research Funds for the Central Universities of Dalian Minzu University. Data availability statement format guidelines The datasets generated during and/or analysed during the current study are available from the corresponding author on reasonable request. Author contributions statements Xiaoyi Gao,Rongfan Xu wrote the main manuscript text, Yiran Yan was responsible for data integration.All authors reviewed the manuscript. References Li, X. M., Qin, Z. Q. & Tu, J. F. Evolvement and development trend of the tree canopy on persimmon ( Diospyros kaki Linn. f.) in China. J. 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Characterization and comparison of yeasts from different sources for some probiotic properties and exopolysaccharide production. Food Sci. Technol. 39 (Suppl. 2), 646–653 (2019). Rossouw, D. & Bauer, F. F. Exploring the phenotypic space of non- Saccharomyces wine yeast biodiversity. Food Microbiol. 55 , 32–46 (2016). Linde, D. et al. Molecular and biochemical characterization of β-fructofuranosidase from Xanthophyllomyces dendrorhous . Appl. Environ. Microbiol. 75 , 1065–1073 (2009). Goosen, C. et al. Molecular and biochemical characterization of a novel intracellular invertase from Aspergillus niger with transfructosylating activity. Eukaryot. Cell. 6 , 674–681 (2007). Hernalsteens, S. & Maugeri, F. Purification and characterization of a fructosyltransferase from Rhodotorula sp. Appl. Microbiol. Biotechnol. 79 , 589–596 (2008). Ghazi, I., Fernández-Arrojo, L., Garcia-Arellano, H., Plou, F. J. & Ballesteros, A. Purification and kinetic characterization of a fructosyltransferase from Aspergillus aculeatus . J. Biotechnol. 128 , 204–211 (2007). Belcarz, A., Ginalska, G., Lobarzewski, J. & Penel, C. The novel non-glycosylated invertase from Candida utilis (the properties and the conditions of and purification). Biochim. Biophys. Acta . 1594 , 40–53 (2002). Álvaro-Benito, M. et al. Characterization of a β-fructofuranosidase from Schwanniomyces occidentalis with transfructosylating activity yielding the prebiotic 6-kestose. J. Biotechnol. 132 , 75–81 (2007). Zeng, H. W., Zheng, H. H., Chen, H., Liao, X. R. & Cai, Y. J. Progress in research on biosynthesis and metabolic engineering of microbial polysaccharides. J. Shaanxi Univ. Technol. (Natural Sci. Edition) . 31 , 49–58 (2015). Jochen, S. Recent insights in microbial exopolysaccharide biosynthesis and engineering strategies. Curr. Opin. Biotechnol. 53 , 130–136 (2018). Li, C. M. The preliminary study on functional characteristic of dried pers (2010). Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-6276251","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":445926367,"identity":"c3580b90-3c01-404a-831a-7723f9e816d9","order_by":0,"name":"Xiaoyi Gao","email":"","orcid":"","institution":"Dalian Minzu University","correspondingAuthor":false,"prefix":"","firstName":"Xiaoyi","middleName":"","lastName":"Gao","suffix":""},{"id":445926368,"identity":"6d9d9a59-16f2-4a7f-8142-946d1052c5d5","order_by":1,"name":"Rongfan Xu","email":"","orcid":"","institution":"Shenyang University","correspondingAuthor":false,"prefix":"","firstName":"Rongfan","middleName":"","lastName":"Xu","suffix":""},{"id":445926369,"identity":"0e8273ae-2de8-4ced-ba78-cea245eff189","order_by":2,"name":"Yiran Yan","email":"","orcid":"","institution":"Dalian Minzu University","correspondingAuthor":false,"prefix":"","firstName":"Yiran","middleName":"","lastName":"Yan","suffix":""},{"id":445926373,"identity":"fa863485-a383-4039-8061-1a500748c189","order_by":3,"name":"Xiaodong Sun","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAqklEQVRIiWNgGAWjYHACwwc8BWCGAdFajA14DEjUYiZBmhb5GcnbKt4Y3ElsYG/eJsFQc4ewFoMzx8puzjF4ltjAc6xMguHYMyK0sPeY3eYxOJzYIJFjJsHYcJgIhzXzmBWDtci/IVILw/EeM2aILTxEagH6pVhyjsFh4zaetGKLhGPEOGxG8sYPbyoOy/azH95440MNMQ6DATYQkUCChlEwCkbBKBgFeAAAgTM2ms3s+ZIAAAAASUVORK5CYII=","orcid":"","institution":"Dalian Minzu University","correspondingAuthor":true,"prefix":"","firstName":"Xiaodong","middleName":"","lastName":"Sun","suffix":""}],"badges":[],"createdAt":"2025-03-21 09:38:27","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-6276251/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-6276251/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":81170724,"identity":"30def46f-fe7c-4538-a241-dbb819577a7a","added_by":"auto","created_at":"2025-04-23 05:10:55","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":257688,"visible":true,"origin":"","legend":"\u003cp\u003eThe making process of dried persimmon\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-6276251/v1/753dce8655054d4f5ae8781a.png"},{"id":81169761,"identity":"a6e0b7b9-38f0-43f7-8a07-de6dbd116f11","added_by":"auto","created_at":"2025-04-23 05:02:55","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":29999,"visible":true,"origin":"","legend":"\u003cp\u003eStandard curve of glucose\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-6276251/v1/00bd956b7d59a6c1fb81f720.png"},{"id":81169759,"identity":"5ac75426-1138-43ab-857d-f5171f4ac94b","added_by":"auto","created_at":"2025-04-23 05:02:55","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":79064,"visible":true,"origin":"","legend":"\u003cp\u003eliquid chromatograms of various sugar standard solutions\u003c/p\u003e","description":"","filename":"3.png","url":"https://assets-eu.researchsquare.com/files/rs-6276251/v1/cd5c75e29ec88a9c5c1a572f.png"},{"id":81169764,"identity":"b5ca48d3-d00b-4526-bbc7-55172cf0f23f","added_by":"auto","created_at":"2025-04-23 05:02:55","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":40846,"visible":true,"origin":"","legend":"\u003cp\u003eThe partial sugar composition in fermentation broth of different strains\u003c/p\u003e","description":"","filename":"4.png","url":"https://assets-eu.researchsquare.com/files/rs-6276251/v1/8f33e39eace0be47b713a3eb.png"},{"id":81169767,"identity":"7a3f0e87-36fd-4734-b845-d1152a7b8985","added_by":"auto","created_at":"2025-04-23 05:02:55","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":33787,"visible":true,"origin":"","legend":"\u003cp\u003eGlucose concentration in fermentation broth of different strains (concentration of G3-3-1 is 28.87 ± 7.777 mg / L)\u003c/p\u003e","description":"","filename":"5.png","url":"https://assets-eu.researchsquare.com/files/rs-6276251/v1/5fa8e13d96c99ed5df27514a.png"},{"id":81932005,"identity":"2af630fd-0265-4ce5-ab28-ec563ec27fc2","added_by":"auto","created_at":"2025-05-05 05:32:32","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1460317,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6276251/v1/73938828-2305-4ef1-92de-c649b500e320.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Preliminary study on the function of natural persimmon frost fungi","fulltext":[{"header":"Introduction","content":"\u003cp\u003eAs the world\u0026rsquo;s largest producer of persimmons, China ranks the first worldwide in terms of the cultivation area and yield of persimmons\u003csup\u003e\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003e Apart from direct consumption, persimmons can also be made into dried persimmons. There are currently two techniques of processing dried persimmons in China, i.e. natural drying and artificial drying. As the traditional processing technology, the natural drying method has a long history and is still being used in many places nowadays. Figure\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e shows the main process of the natural drying method. It involves a long production cycle about 30\u0026ndash;50 days\u003csup\u003e\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u003c/sup\u003e During this process, the sugars in the persimmon gradually release, and dry white crystals gradually form on the surface of the persimmon as the water evaporates, which being referred to as the persimmon frost. Having been regarded as the essence of dried persimmons ever since ancient times, persimmon frost plays an effective role in treating oral ulcer and acute pharyngitis. However, there are currently few researches on persimmon frost in the world, and the very few researches available are limited to China.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThese researches mainly focus on the ingredients, formation and effects of persimmon frosMa et al. t preliminarily extracted persimmon frost with organic solvents such as petroleum ether and 95% ethanol, and the sugars detected in the persimmon frost included glucose, fructose, sucrose, xylose and a trace of mannose\u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u003c/sup\u003e. By measuring the content of protein, fat, total acid and total sugar in persimmon frost, Zhang (2002) found that total sugar accounted for 95% in persimmon frost, among which polysaccharide took up for 7%, sucrose accounted for 6%, glucose 37%, and fructose 45%\u003csup\u003e4\u003c/sup\u003e Zhang and Huang (2009) found that reducing sugars accounted for 58.21% of the total weight of persimmon frost, and these reducing sugars were glucose, fructose, a trace of rhamnose and arabinose, without galactose and xylose\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eDuring the long-term formation of persimmon frost, many fungi in the air will inevitably accumulate on the surface. By isolating the fungi in persimmon frost, Wang (2010) obtained various species of fungi such as \u003cem\u003ePenicillium citrinum\u003c/em\u003e, \u003cem\u003eP. chrysogenum\u003c/em\u003e, \u003cem\u003eP. oxalicum\u003c/em\u003e, \u003cem\u003eAspergillus flavus\u003c/em\u003e and \u003cem\u003eA. niger.\u003c/em\u003e Bai (2017) separated \u003cem\u003ePenicillium chrysogenum\u003c/em\u003e and \u003cem\u003eAspergillus niger\u003c/em\u003e from persimmon frost, and relevant analysis suggested that they were the contaminating strains that caused the dried persimmon to discolor and rot\u003csup\u003e\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e,\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eEight strains, including three strains of \u003cem\u003eAureobasidium pullulans\u003c/em\u003e, \u003cem\u003eFilobasidium magnum\u003c/em\u003e, \u003cem\u003eEpicoccum nigrum\u003c/em\u003e, \u003cem\u003eAspergillus versicolor\u003c/em\u003e, \u003cem\u003eMetschnikowia pulcherrima\u003c/em\u003e and \u003cem\u003ePapiliotrema flavescens\u003c/em\u003e, were isolated from dried persimmon samples freshly prepared by natural drying method. Unlike previous studies which only obtained \u003cem\u003ePenicillium\u003c/em\u003e and \u003cem\u003eAspergillus\u003c/em\u003e species from persimmon frost, these strains of fungus were isolated from persimmon frost for the first time. The surface of persimmon frost is rich in sugar. Are these persimmon frost fungi involved in the conversion of sugars in persimmon frost? Is it possible to synthesize polysaccharides using the sugars on the surface of persimmon frost? At present, there is no relevant report. In order to clarify the function of persimmon frost fungi, the change of sugars in the fermentation broth before and after fermentation were studied by means of single strain fermentation and mass spectrometry detection. The results of this study provide a powerful reference for evaluating the ability of sugar conversion and polysaccharide synthesis of persimmon frost fungi.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eActivation of strains\u003c/h2\u003e \u003cp\u003eThe strains were inoculated into PDA plate for activation.\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003ePrimary and secondary fermentation\u003c/h3\u003e\n\u003cp\u003eThe strains were inoculated into 100 mL of the primary fermentation broth to be cultured in the dark for 96h, with the temperature of 18℃ and the rotating speed being 140r/min. The primary broth, with an inoculated dose of 2%, was transferred into a conical flask equipped with 100 mL of re-fermentation medium to be cultured for 7d with the temperature of 18\u0026deg;C and the rotating speed being 140r/min. All strains were repeated three times.\u003c/p\u003e \u003cp\u003eThe primary fermentation medium (seed medium) is composed of 50.0 g of sucrose, 4.0 g of K\u003csub\u003e2\u003c/sub\u003eHPO\u003csub\u003e4\u003c/sub\u003e, 0.8g of (NH\u003csub\u003e4\u003c/sub\u003e)\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, 0.2g of MgSO\u003csub\u003e4\u003c/sub\u003e, 2.0g of NaCl and 1.5 g of yeast powder (pH\u0026thinsp;=\u0026thinsp;7.0). The medium was sterilized at 121\u0026deg;C for 20min\u003csup\u003e\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eThe re-fermentation medium consists of 50.0g of sucrose, 6.0g of K\u003csub\u003e2\u003c/sub\u003eHPO\u003csub\u003e4\u003c/sub\u003e, 0.6g of (NH\u003csub\u003e4\u003c/sub\u003e)\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, 0.5g of MgSO\u003csub\u003e4\u003c/sub\u003e, 4.0g of NaCl and 0.9g of yeast powder (initial pH\u0026thinsp;=\u0026thinsp;6.5). The medium was sterilized at 121\u0026deg;C for 20min.\u003c/p\u003e\n\u003ch3\u003eDetection of polysaccharides\u003c/h3\u003e\n\u003cp\u003eThe fermentation broth was centrifuged at 12000r/min for 5 min, the fungi mycelium and sediments were discarded, the fermentation broth was filtered, and the supernatant was collected. The supernatant passed through the dialysis bag (retaining a relative molecular weight of 3.5KD), and distilled water was added until the liquid in the dialysis bag reached uniformly 100 mL. The liquid outside the dialysis bag was also collected (the dialysis bag was placed in a beaker and the final volume of water in the beaker was maintained at 300 mL).\u003c/p\u003e \u003cp\u003eThe concentration of polysaccharides in the dialysis bag was detected using the concentrated sulfuric acid-phenol method\u003csup\u003e\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e,\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u003c/sup\u003e 1 mL of fermentation broth, 1 mL of distilled water, 1 mL of 6% phenol solution and 5 mL of concentrated sulfuric acid were added to a 25 mL colorimetric tube which was then covered and shaken up. After that, the colorimetric tube experienced a boiling water bath for 15 min, then be cooled. The OD value was measured at 490 nm, with the above operation repeated on each sample for three times. Distilled water replaced the sample solution to join the reaction system as the blank control. In this study, as a standard stock solution, the 1mg/mL glucose solution was diluted to a standard solution with a concentration of 0, 0.02, 0.04, 0.08, 0.1, and 0.15 mg/mL, respectively. With the reaction operated per the above steps, the absorbance was measured at 490 nm. A standard curve was plotted with glucose concentration as the abscissa and absorbance as the ordinate. By introducing the absorbance of the sample into the standard curve, the concentration of polysaccharides in the sample was calculated.\u003c/p\u003e\n\u003ch3\u003eDetection of monosaccharides and sugar acids\u003c/h3\u003e\n\u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003eMetabolites extraction\u003c/h2\u003e \u003cp\u003e1)The liquid outside the dialysis bag were collected and 50\u0026micro;L liquid samples or standard solution were taken for derivative reaction.\u003c/p\u003e \u003cp\u003eDerivatization: add 100 \u0026micro;L methanol solution of 1-phenyl-3-methyl-5-pyrazolone (10 mg / mL) and 30 \u0026micro;L of 0.1 mol / L NaOH solution to the sample (or equivalent volume of standard solution), and then react at 70 ℃ for 30 min. After cooling to room temperature, 30 \u0026micro;L of 0.1 mol / L HCl solution was added. After concentration and drying, 600 \u0026micro;L of water was redissolved, and 1 mL of chloroform was used to extract three times to remove the excessive derivatization reagent.\u003c/p\u003e \u003cp\u003e2)After the reaction, the mixture was dried and dissolved with 200 L methanol. The supernatant was taken for determination.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003ePreparation of standard solution\u003c/h2\u003e \u003cp\u003ePrepare 10 mmol/L standard stock solution. Take the corresponding amount of standard stock solution into a 10 mL volumetric flask and prepare the mixed standard solution. Dilute the standard solution in turn to obtain a series of calibration solutions. See Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e for details of each standard substance.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eSpecific information about the standard substances\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCompound name\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eMolecular Weight\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eManufacturer\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePurity\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD-Mannose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e180.16\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eTCI, China\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026ge;\u0026thinsp;98.0%\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD-Galactose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e180.16\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eYuanye, China\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026ge;\u0026thinsp;98.0%\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD-Glucose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e180.16\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eDR.E, Germany\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026ge;\u0026thinsp;99.7%\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD-Glucuronic Acid\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e194.14\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eTRC, Canada\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026ge;\u0026thinsp;98.0%\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eAlpha-D-Galacturonic Acid Hydrate\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e212.16\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eFLUKA, Switzerland\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026ge;\u0026thinsp;97.0%\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eL-Rhamnose monohydrate\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e182.17\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSIGMA, America\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026ge;\u0026thinsp;99.0%\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eL-Fucose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e164.16\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eTCI, China\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026ge;\u0026thinsp;97.0%\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD-Ribose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e150.13\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eZhenzhun, China\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026ge;\u0026thinsp;98.0%\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eDL-Arabinose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e150.13\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eVETEC, China\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026ge;\u0026thinsp;99%\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD (+)-Xylose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e150.13\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eTCI, China\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026ge;\u0026thinsp;98.0%\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e5-Methyl-2-phenyl-1,2-dihydropyrazol-3-one\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e174.20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eTCI, China\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u0026ge;\u0026thinsp;98.0%\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eMachine Detection\u003c/h3\u003e\n\u003cp\u003eIn this study, a Waters Acquity UPLC ultra high performance liquid chromatograph was used to perform chromatographic separation of the target compounds with a Waters Acquity UPLC BEH C18 liquid chromatographic column (100 \u0026times; 2.1mm, 1.7\u0026micro;m, Waters). The mobile phase A was 0.1% formic acid aqueous solution, and the mobile phase B was 0.1% formic acid acetonitrile solution. The column temperature was 45 ℃, the sample tray was set at 4℃, and the injection volume was 2 \u0026micro;L. The Xevo G2-XS QTOF high resolution mass spectrometer was used for full scan mass spectrometry analysis. Ion source parameters are as follows: Capillary voltage\u0026thinsp;=\u0026thinsp;2000 V, Sampling cone\u0026thinsp;=\u0026thinsp;40 V, Source temperature\u0026thinsp;=\u0026thinsp;115℃, Desolvation temperature\u0026thinsp;=\u0026thinsp;500\u0026deg;C, Desolvation gas\u0026thinsp;=\u0026thinsp;900 L/h, Scan Range\u0026thinsp;=\u0026thinsp;400\u0026thinsp;~\u0026thinsp;600, Scan time\u0026thinsp;=\u0026thinsp;0.3 s. For each target compound, the derived parent ion was used for quantitative analysis. See Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e for details.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eMobile phase gradient and MS parameters\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colspan=\"4\" nameend=\"c4\" namest=\"c1\"\u003e \u003cp\u003eMobile phase gradient\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTime\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eA (H\u003csub\u003e2\u003c/sub\u003eO)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eB (ACN)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eFlow\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e0.0 min\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e95%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e5%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.40 mL/min\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e8.0 min\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e75%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e25%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.40 mL/min\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e9.0 min\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e100%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.40 mL/min\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e11.0 min\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e100%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.40 mL/min\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e11.5 min\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e95%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e5%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.40 mL/min\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e13.0 min\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e95%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e5%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.40 mL/min\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"4\" nameend=\"c4\" namest=\"c1\"\u003e \u003cp\u003e\u003cb\u003eMS parameters\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCompound Name\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eRT (min)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003em/z\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePolarity\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD-Mannose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e5.39\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e511.2193\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePositive\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD-Galactose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e7.31\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e511.2193\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePositive\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD-Glucose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e7.54\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e511.2193\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePositive\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD-Glucuronic Acid\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e8.86\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e525.1985\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePositive\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eAlpha-D-Galacturonic Acid Hydrate\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e9.15\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e525.1985\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePositive\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eL-Rhamnose monohydrate\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e6.74\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e495.2244\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePositive\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eL-Fucose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e9.00\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e495.2244\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePositive\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD-Ribose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e6.35\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e481.2087\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePositive\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eDL-Arabinose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e7.91\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e481.2087\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePositive\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD (+) - Xylose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e8.40\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e481.2087\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePositive\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e\n\u003ch3\u003eStandard curves\u003c/h3\u003e\n\u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003eThe standard curve of polysaccharides\u003c/h2\u003e \u003cp\u003eThe standard curves of polysaccharide drawn based on the concentration and absorbance of standard glucose solution were shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e, where x represents the glucose concentration and y stands for the absorbance. By substituting the absorbance of the sample into the equation, the concentrations of polysaccharide in the fermentation broth were calculated.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003eThe standard curves of monosaccharides and sugar acids\u003c/h2\u003e \u003cp\u003eThe UPLC-HRMS analysis of the standard solution of various sugars was carried out using the method described above, and the standard curve was obtained. Where y represents the peak area of the target compound and x represents the concentration of the target compound. When the least square method was used for regression analysis and the weight was set as 1/x, the calibration solution recovery rate (accuracy) and correlation coefficient (R\u003csup\u003e\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u003c/sup\u003e) were the best. During the detection, it was found that the glucose contents in the samples were too high. Finally, the internal standard method was used for quantitative analysis, and the internal standard was 13C\u003csub\u003e6\u003c/sub\u003e-glucose (ALDRICH, purity\u0026thinsp;\u0026ge;\u0026thinsp;99%). In the curve of glucose, y represents the ratio of the peak area of the target compound to the peak area of the corresponding internal standard, and x represents the concentration of the target compound. See Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e3\u003c/span\u003e for details.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab3\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eThe standard curves of monosaccharides and sugar acids\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCompound Name\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eLinear Range (\u0026micro;g/mL)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCalibration curve\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eR\u003csup\u003e\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD-Glucose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e6.053-121.068\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ey\u0026thinsp;=\u0026thinsp;1.31322x-2.77829\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9997\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD-Mannose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.535-107.015\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ey\u0026thinsp;=\u0026thinsp;39437.9x-6812.69\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9981\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD-Galactose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.077\u0026ndash;38.734\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ey\u0026thinsp;=\u0026thinsp;58417.9x-2240.98\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9982\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD-Glucuronic Acid\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.670-66.978\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ey\u0026thinsp;=\u0026thinsp;3892.19x-890.331\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9902\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eAlpha-D-Galacturonic Acid Hydrate\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.115\u0026ndash;22.983\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ey\u0026thinsp;=\u0026thinsp;7105.2x\u0026thinsp;+\u0026thinsp;29.7572\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9992\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eL-Rhamnose monohydrate\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.089\u0026ndash;44.495\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ey\u0026thinsp;=\u0026thinsp;49763.3x-1806.84\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.986\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eL-Fucose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.050-10.000\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ey\u0026thinsp;=\u0026thinsp;101929x-2861.82\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9991\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD-Ribose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.116\u0026ndash;58.025\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ey\u0026thinsp;=\u0026thinsp;54799.5x-3715.83\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.998\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eDL-Arabinose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.132\u0026ndash;26.393\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ey\u0026thinsp;=\u0026thinsp;87997x-4172.61\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9958\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD (+)-Xylose\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.067\u0026ndash;13.497\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ey\u0026thinsp;=\u0026thinsp;89690.4x-4105.06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.9985\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e"},{"header":"Results","content":"\u003cdiv id=\"Sec14\" class=\"Section2\"\u003e \u003ch2\u003eThe polysaccharide concentration in fermentation broth of each strain\u003c/h2\u003e \u003cp\u003eIn this study, polysaccharide contents were detected in the fermentation broth of 8 strains. The polysaccharide concentrations of the three \u003cem\u003eAureobasidium pullulans\u003c/em\u003e were significantly higher than those of the other fungi. \u003cem\u003eEpicoccum nigrum\u003c/em\u003e and \u003cem\u003eFilobasidium magnum\u003c/em\u003e had lower polysaccharide concentrations. See Table\u0026nbsp;\u003cspan refid=\"Tab4\" class=\"InternalRef\"\u003e4\u003c/span\u003e for details.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab4\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 4\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eThe contents of polysaccharides produced by the strains\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eStrain number\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSpecies\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eA\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePolysaccharide (mg/mL)\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSD3-1-1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eEpicoccum nigrum\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.414\u0026thinsp;\u0026plusmn;\u0026thinsp;0.017\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e0.075\u0026thinsp;\u0026plusmn;\u0026thinsp;0.003\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eFZ2-3-5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eAspergillus versicolor\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.343\u0026thinsp;\u0026plusmn;\u0026thinsp;0.016 *2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e0.124\u0026thinsp;\u0026plusmn;\u0026thinsp;0.006\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSD3-1-2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eAureobasidium pullulans\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.366\u0026thinsp;\u0026plusmn;\u0026thinsp;0.038 *4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e0.263\u0026thinsp;\u0026plusmn;\u0026thinsp;0.028\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eG3-3-1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eMetschnikowia pulcherrima\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.419\u0026thinsp;\u0026plusmn;\u0026thinsp;0.042 *2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e0.151\u0026thinsp;\u0026plusmn;\u0026thinsp;0.015\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSX1-2-3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003ePapiliotrema flavescens\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.619\u0026thinsp;\u0026plusmn;\u0026thinsp;0.042\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e0.112\u0026thinsp;\u0026plusmn;\u0026thinsp;0.008\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSD3-2-1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eFilobasidium magnum\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.394\u0026thinsp;\u0026plusmn;\u0026thinsp;0.008\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e0.071\u0026thinsp;\u0026plusmn;\u0026thinsp;0.001\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSD2-3-1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eAureobasidium pullulans\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.439\u0026thinsp;\u0026plusmn;\u0026thinsp;0.008 *4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e0.317\u0026thinsp;\u0026plusmn;\u0026thinsp;0.006\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSD1-2-4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cem\u003eAureobasidium pullulans\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.473\u0026thinsp;\u0026plusmn;\u0026thinsp;0.009 *3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e0.256\u0026thinsp;\u0026plusmn;\u0026thinsp;0.005\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"4\"\u003eNote: A is the absorbance value of polysaccharide. The number behind the asterisk is the sample\u0026rsquo;s dilution multiple. The values in the table are average\u0026thinsp;\u0026plusmn;\u0026thinsp;standard deviation (n\u0026thinsp;=\u0026thinsp;3).\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec15\" class=\"Section2\"\u003e \u003ch2\u003eThe concentrations of monosaccharides and sugar acids\u003c/h2\u003e \u003cp\u003e1)Liquid chromatograms of standard solutions\u003c/p\u003e \u003cp\u003eThe chromatogram of standard solutions is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e, from which it can be seen that all target compounds present symmetrical chromatographic peaks, realizing the chromatographic separation of each target compound.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThe peak-side notes are compounds, retention time, peak area, and concentration in order\u003c/p\u003e \u003cp\u003e2)The concentrations of sugars in the fermentation broth\u003c/p\u003e \u003cp\u003eThe quantitative results of monosaccharides and sugar acids in the samples are shown in Table\u0026nbsp;\u003cspan refid=\"Tab5\" class=\"InternalRef\"\u003e5\u003c/span\u003e. It can be seen from the table that there are great differences in the composition of ten monosaccharides and sugar acids in the fermentation broth of the 8 strains. Glucose is the most important monosaccharide component in the fermentation broth of each strain. Among them, \u003cem\u003eFilobasidium magnum\u003c/em\u003e SD3-2-1 had the highest glucose concentration, reaching 36316.67\u0026thinsp;\u0026plusmn;\u0026thinsp;394.774 mg / L. The concentrations of \u003cem\u003eEpicoccum nigrum\u003c/em\u003e SD3-1-1, \u003cem\u003eAspergillus versicolor\u003c/em\u003e FZ2-3-5, \u003cem\u003ePapiliotrema flavescens\u003c/em\u003e SX1-2-3 and \u003cem\u003eAureobasidium pullulans\u003c/em\u003e SD2-3-1 were also relatively high. The glucose concentration of \u003cem\u003eMetschnikowia pulcherrima\u003c/em\u003e G3-3-1 was the lowest, only 28.87\u0026thinsp;\u0026plusmn;\u0026thinsp;7.777 mg / L. Galacturonic acid, rhamnose and fucose were not detected in the fermentation broth of the 8 strains, or their response intensity was lower than the minimum concentration of the standard curve, so they could not be quantified. The concentrations of mannose, galactose, glucuronic acid, ribose, arabinose and xylose in the fermentation broth of the eight strains were very different. Galactose and glucuronic acid were only detected in the fermentation broth of two strains SD3-1-2 and SD1-2-4 of \u003cem\u003eAureobasidium pullulans\u003c/em\u003e. Mannose, ribose, arabinose and xylose were found in the fermentation broth of eight strains. The contents of mannose, ribose and arabinose in the fermentation broth of different strains were quite different, while xylose was a small amount in the fermentation broth of each strain. See Figs.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e and \u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e for details.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab5\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 5\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eThe types and concentrations of monosaccharides in fermentation broth of each strain\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"12\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c10\" colnum=\"10\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c11\" colnum=\"11\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c12\" colnum=\"12\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eSample Name\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"10\" nameend=\"c11\" namest=\"c2\"\u003e \u003cp\u003eMetabolite Concentration (mg/L)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"1\" nameend=\"c12\" namest=\"c12\"\u003e\u0026nbsp;\u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eGlucose\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eMannose\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eGalactose\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eGlucuronic acid\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eGalacturonic acid\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eRhamnose\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003eFucose\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c9\"\u003e \u003cp\u003eRibose\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c10\"\u003e \u003cp\u003eArabinose\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c11\"\u003e \u003cp\u003eXylose\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"1\" nameend=\"c12\" namest=\"c12\"\u003e\u0026nbsp;\u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSD3-1-2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e9271.73\u0026thinsp;\u0026plusmn;\u0026thinsp;771.053 (\u0026times;40)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e12.99\u0026thinsp;\u0026plusmn;\u0026thinsp;4.222\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e26.34\u0026thinsp;\u0026plusmn;\u0026thinsp;6.909\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e1.19\u0026thinsp;\u0026plusmn;\u0026thinsp;0.029\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;LLOQ\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e45.11\u0026thinsp;\u0026plusmn;\u0026thinsp;5.345\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e2.20\u0026thinsp;\u0026plusmn;\u0026thinsp;0.447\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e0.33\u0026thinsp;\u0026plusmn;\u0026thinsp;0.029\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"1\" nameend=\"c12\" namest=\"c12\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSD1-2-4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e3676.67\u0026thinsp;\u0026plusmn;\u0026thinsp;324.385 (\u0026times;100)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e9.57\u0026thinsp;\u0026plusmn;\u0026thinsp;0.847\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e10.48\u0026thinsp;\u0026plusmn;\u0026thinsp;3.877\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e1.54\u0026thinsp;\u0026plusmn;\u0026thinsp;0.156\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u0026lt;LLOQ\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;LLOQ\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e28.67\u0026thinsp;\u0026plusmn;\u0026thinsp;5.761\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e1.19\u0026thinsp;\u0026plusmn;\u0026thinsp;0.202\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e0.16\u0026thinsp;\u0026plusmn;\u0026thinsp;0.005\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"1\" nameend=\"c12\" namest=\"c12\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSD2-3-1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e19451.67\u0026thinsp;\u0026plusmn;\u0026thinsp;763.182 (\u0026times;500)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3.48\u0026thinsp;\u0026plusmn;\u0026thinsp;1.032\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt;LLOQ\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;LLOQ\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e24.40\u0026thinsp;\u0026plusmn;\u0026thinsp;3.849\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e0.92\u0026thinsp;\u0026plusmn;\u0026thinsp;0.088\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e0.11\u0026thinsp;\u0026plusmn;\u0026thinsp;0.010\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"1\" nameend=\"c12\" namest=\"c12\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSD3-2-1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e36316.67\u0026thinsp;\u0026plusmn;\u0026thinsp;394.774 (\u0026times;500)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e21.30\u0026thinsp;\u0026plusmn;\u0026thinsp;2.557\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;LLOQ\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e0.39\u0026thinsp;\u0026plusmn;\u0026thinsp;0.035\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e0.71\u0026thinsp;\u0026plusmn;\u0026thinsp;0.064\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e0.08\u0026thinsp;\u0026plusmn;\u0026thinsp;0.001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"1\" nameend=\"c12\" namest=\"c12\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSD3-1-1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e23548.33\u0026thinsp;\u0026plusmn;\u0026thinsp;157.056 (\u0026times;500)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e18.72\u0026thinsp;\u0026plusmn;\u0026thinsp;0.721\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;LLOQ\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e1.69\u0026thinsp;\u0026plusmn;\u0026thinsp;0.117\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e0.40\u0026thinsp;\u0026plusmn;\u0026thinsp;0.080\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e0.13\u0026thinsp;\u0026plusmn;\u0026thinsp;0.014\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"1\" nameend=\"c12\" namest=\"c12\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eFZ2-3-5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e25816.67\u0026thinsp;\u0026plusmn;\u0026thinsp;1710.967 (\u0026times;500)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e19.52\u0026thinsp;\u0026plusmn;\u0026thinsp;0.913\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;LLOQ\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e5.33\u0026thinsp;\u0026plusmn;\u0026thinsp;0.575\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e0.30\u0026thinsp;\u0026plusmn;\u0026thinsp;0.031\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e0.13\u0026thinsp;\u0026plusmn;\u0026thinsp;0.005\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"1\" nameend=\"c12\" namest=\"c12\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eG3-3-1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e28.87\u0026thinsp;\u0026plusmn;\u0026thinsp;7.777\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.28\u0026thinsp;\u0026plusmn;\u0026thinsp;0.041\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;LLOQ\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e1.56\u0026thinsp;\u0026plusmn;\u0026thinsp;0.291\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e20.99\u0026thinsp;\u0026plusmn;\u0026thinsp;0.957\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e0.07\u0026thinsp;\u0026plusmn;\u0026thinsp;0.001\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"1\" nameend=\"c12\" namest=\"c12\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSX1-2-3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e27485.00\u0026thinsp;\u0026plusmn;\u0026thinsp;509.200 (\u0026times;500)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3.69\u0026thinsp;\u0026plusmn;\u0026thinsp;1.040\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u0026lt;LLOQ\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eN.D.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e\u0026lt;LLOQ\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e1.15\u0026thinsp;\u0026plusmn;\u0026thinsp;0.350\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c10\"\u003e \u003cp\u003e0.58\u0026thinsp;\u0026plusmn;\u0026thinsp;0.260\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c11\"\u003e \u003cp\u003e0.38\u0026thinsp;\u0026plusmn;\u0026thinsp;0.090\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"1\" nameend=\"c12\" namest=\"c12\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"11\" nameend=\"c11\" namest=\"c1\"\u003e \u003cp\u003eThe target metabolite concentration in the sample is the final concentration measured by the instrument multiplied by the dilution ratio of the sample in the extraction solvent, in mg / L. Among them, glucose is diluted data and other compounds are undiluted data. \"N.D.\" indicates that the target compound in the sample is not detected. \u0026ldquo;\u0026lt;LLOQ\u0026rdquo; means that the target compounds in the sample have chromatographic peaks, but their response strength is lower than the minimum concentration of the standard curve, and cannot be quantified. The values listed in the glucose column are multiples of the sample dilution.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colspan=\"1\" nameend=\"c12\" namest=\"c12\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eNote: The values in the table are average\u0026thinsp;\u0026plusmn;\u0026thinsp;standard deviation (n\u0026thinsp;=\u0026thinsp;3)\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e"},{"header":"Discussion","content":"\u003cp\u003eTwo filamentous fungi, \u003cem\u003eAspergillus versicolor\u003c/em\u003e and \u003cem\u003eEpicoccum nigrum\u003c/em\u003e with polysaccharide activity were detected in the study. As common fungal groups in the air, \u003cem\u003eAspergillus\u003c/em\u003e are abundant in naturally fermented foods and they participate in the fermentation process of the foods. In this genus, \u003cem\u003eAspergillus oryzae\u003c/em\u003e and \u003cem\u003eA. niger\u003c/em\u003e are important strains of fermentation, which play an important role in the fermentation of many foods\u003csup\u003e\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e,\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e\u003c/sup\u003e \u003cem\u003eAspergillus versicolor\u003c/em\u003e is also an important functional strain, and has been widely reported. As an endophytic fungus, it can produce a variety of antibacterial substances, and some strains isolated from the deep sea can metabolize to produce polysaccharides \u003csup\u003e\u003cspan additionalcitationids=\"CR14 CR15\" citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e\u003c/sup\u003e \u003cem\u003eEpicocum nigrum\u003c/em\u003e can produce yellow pigment, as well as polyketide, carotenoids and flavonoids, and these bioactive compounds are widely used in the production of functional food, medicine and cosmetics\u003csup\u003e\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u003c/sup\u003e However, there are few reports on the production of polysaccharides by \u003cem\u003eEpicocum nigrum\u003c/em\u003e.\u003c/p\u003e \u003cp\u003eSix polysaccharide-producing yeast strains were obtained in the study. It is known to all that \u003cem\u003eAureobasidium pullulans\u003c/em\u003e can produce extracellular polysaccharide Pullulan during fermentation, and Pullulan has been extensively used in the fields of food, pharmacy and biomedicine\u003csup\u003e\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e\u003c/sup\u003e Whether the three strains of \u003cem\u003eAureobasidium pullulans\u003c/em\u003e produce Pullulan or other types of polysaccharides still requires further intensive research. \u003cem\u003eMetschnikowia\u003c/em\u003e commonly found on the surface of grapes or in naturally fermented wines and fruit juices, can produce alcohol and volatile organic compounds during metabolism. Besides, studies have shown that \u003cem\u003eMetschnikowia\u003c/em\u003e can produce extracellular polysaccharides\u003csup\u003e\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e,\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e\u003c/sup\u003e As dimorphic basidiomycetes, \u003cem\u003ePapiliotrema\u003c/em\u003e and \u003cem\u003eFilobasidium\u003c/em\u003e can produce yeast-like spores in their life histories. There are few studies on the application of the two kinds of fungi and on their production of polysaccharides.\u003c/p\u003e \u003cp\u003eThese 8 strains were isolated from persimmon frost for the first time, and the polysaccharide activity was detected in all of them. This provides a new direction for screening polysaccharide producing strains and developing new polysaccharides.\u003c/p\u003e \u003cdiv id=\"Sec17\" class=\"Section2\"\u003e \u003ch2\u003eSugar transformation and formation of polysaccharides by fungi\u003c/h2\u003e \u003cp\u003eSeveral kinds of monosaccharides and sugar acids were detected in the fermentation broth outside the dialysis bag. It was found that glucose was the main carbohydrate substance in the filtrate of each strain, and various sugars were also detected. Sucrose is the only carbohydrate in the original fermentation medium. Sucrose can be hydrolyzed into glucose under the action of the sucrase produced by the strains. Many studies have isolated and obtained the sucrase from fungi and yeast\u003csup\u003e\u003cspan additionalcitationids=\"CR22 CR23 CR24 CR25\" citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e\u003c/sup\u003e Under the action of glucokinase, the produced glucose turns into glucose-6-phosphate, which is converted into the precursor nucleotide sugar of various sugars by the action of other synthetases, mutases, isomerases and other enzyme systems produced by the strains. It is the activation form of various monosaccharides and sugar acids, and the starting material of polysaccharide synthesis. Then, under the action of highly specific glycosyltransferases (GTS), the extension, reversion and polymerization of polysaccharide repeat units are carried out, and the polysaccharide output is also carried out\u003csup\u003e\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e,\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e\u003c/sup\u003e The research of polysaccharide synthesis mainly focuses on bacteria and a few medicinal fungi. Although the structure of polysaccharides varies greatly, the synthetic pathway is almost the same. In this study, polysaccharides were detected from the dialysate bags of each strain. It can be inferred that these eight strains have the function of utilizing, transforming sugars and synthesizing polysaccharides.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec18\" class=\"Section2\"\u003e \u003ch2\u003ePotential functions of fungi in persimmon frost\u003c/h2\u003e \u003cp\u003eThere are few reports on the study of persimmon frost polysaccharides. Li (2010) used the ultrasonic-assisted hot water extraction method to extract crude persimmon polysaccharide on the surface of dried persimmons, and detected the antioxidative competence of persimmon polysaccharides\u003csup\u003e\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e\u003c/sup\u003e Is it rational to speculate that there are abundant fungal groups on the surface of persimmon frost and some species can produce polysaccharides using the sugar on the surface of persimmon frost? As a matter of fact, the formation of persimmon frost polysaccharide is the process of metabolism and transformation of carbohydrates with the participation of fungi. Different fungi produce various polysaccharide mixtures with different structures and functions on the surface of dried persimmon. The function and efficacy of persimmon frost are probably a result of these polysaccharides working with other beneficial ingredients in the persimmon frost. In addition, as the main carbon source, sucrose in the original fermentation broth was hydrolyzed by the strains, and many kinds of other sugars was detected in the fermentation broth, which indicating that sugar transformation occurred in the fermentation broths to form other sugars. It is speculated that the sugars on the surface of persimmon frost consists of two parts, namely, the sugars inside the persimmon that being evaporated to the surface of dried persimmon, and the sugar converted from other carbohydrates by the fungal metabolism on the surface of persimmon frost. The fungi on the surface of persimmon frost take an active part in both the formation of polysaccharides and the mutual conversion of carbohydrates, influencing and altering the composition and function of carbohydrates in the persimmon frost.\u003c/p\u003e \u003cp\u003eDue to the traditional methods of separation, fermentation and detection, there will be some growth and metabolic limitations of these fungi, as well as errors caused by human factors. However, the results can roughly reveal the ability of multiple fungi on persimmon frost utilizing sugars in persimmon frost and generating polysaccharides. Therefore, not all fungi in the persimmon frost are contaminating species, and many of them are functional strains that can produce polysaccharides. Due to the unique macromolecular structure and obvious biological activity of polysaccharides, we should focus on these new polysaccharides in future researches.\u003c/p\u003e \u003c/div\u003e"},{"header":"Conclusion","content":"\u003cp\u003eCarbohydrates not only provide the human body with necessary energy, but are also equipped with a variety of biological activities. Researches on carbohydrates have attracted extensive attention in recent years, of which the researches on polysaccharides are the most popular. In the current stage, to screen polysaccharide-producing strains and develop new functional polysaccharides, attempts have been made to separate microorganisms that can metabolize polysaccharides from different habitats such as oceans, soils, plants and extreme environments. Most of these microorganisms are bacteria, but there are also a small amount of fungi. Dried persimmon is a food with Chinese characteristics. As the essence on the surface of dried persimmon, persimmon frost contains rich carbohydrates that provide a favorable carbon source for the growth and metabolism of fungi. In this study, eight strains were isolated from dried persimmon samples. All of them are capable of transforming carbohydrates and producing polysaccharides. With persimmon frost as the substrate, these fungi could change the composition and function of carbohydrates in persimmon frost. This study took the initiative to explore the fungi in persimmon frost, the results of which provide a new orientation for screening fungi that produce extracellular polysaccharides and developing new functional polysaccharides.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eDeclaration of Competing Interest\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors have declared no conflict of interest.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis research was supported by the funds of Fundamental Research Funds for the Central Universities of Dalian Minzu University.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData availability statement format guidelines\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe datasets generated during and/or analysed during the current study are available from the corresponding author on reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor contributions statements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eXiaoyi Gao,Rongfan Xu wrote the main manuscript text, Yiran Yan was responsible for data integration.All authors reviewed the manuscript.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eLi, X. 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Progress in research on biosynthesis and metabolic engineering of microbial polysaccharides. \u003cem\u003eJ. Shaanxi Univ. Technol. (Natural Sci. Edition)\u003c/em\u003e. \u003cb\u003e31\u003c/b\u003e, 49\u0026ndash;58 (2015).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJochen, S. Recent insights in microbial exopolysaccharide biosynthesis and engineering strategies. \u003cem\u003eCurr. Opin. Biotechnol.\u003c/em\u003e \u003cb\u003e53\u003c/b\u003e, 130\u0026ndash;136 (2018).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLi, C. M. The preliminary study on functional characteristic of dried pers (2010).\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Natural persimmon frost, Filamentous fungi, Yeast, Monosaccharide, Polysaccharide","lastPublishedDoi":"10.21203/rs.3.rs-6276251/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6276251/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eChina is the largest persimmon producer, with the largest cultivation area and yield in the world. These rich persimmons, in addition to direct consumption, can also be made into dried persimmon. Dried persimmon is a traditional Chinese processed food which preserves the nutrition of persimmons. Persimmon frost, icing sugar on the surface of dried persimmon, is formed by the diffusion of sugar inside the dried persimmon to the surface, which has the effect of moistening lungs and relieving cough. During the long and open process of persimmon frost formation, there are a lot of airborne fungi falling freely on the surface of persimmon frost, but there are a few reports on persimmon frost fungi so far. In this study, eight fungal strains, including three strains of \u003cem\u003eAureobasidium pullulans\u003c/em\u003e, \u003cem\u003eFilobasidium magnum\u003c/em\u003e, \u003cem\u003eEpicoccum nigrum\u003c/em\u003e, \u003cem\u003eAspergillus versicolor\u003c/em\u003e, \u003cem\u003eMetschnikowia pulcherrima\u003c/em\u003e and \u003cem\u003ePapiliotrema flavescens\u003c/em\u003e, were isolated from dried persimmon samples freshly prepared by natural drying method.\u003c/p\u003e \u003cp\u003eSucrose was used as carbon source for single strain fermentation. The fermentation broths were passed through the dialysis bag (retaining a relative molecular weight of 3.5KD), and the polysaccharides were detected inside the bags using the concentrated sulfuric acid-phenol method. The results showed that the polysaccharides were detected out in all the fermentation broths, and the concentrations of the three \u003cem\u003eAureobasidium pullulans\u003c/em\u003e strains were significantly higher than those of the other fungi. The filtrate outside the dialysis bag was detected for ten monosaccharides and sugar acids by mass spectrometry detection, including glucose, galacturonic acid, rhamnose, fucose, mannose, galactose, glucuronic acid, ribose, arabinose and xylose. There are great differences in the compositions and concentrations of the monosaccharides and sugar acids in the fermentation broths of the 8 strains, and glucose is the most important monosaccharide. Galacturonic acid, rhamnose and fucose were not detected in the fermentation broth of the 8 strains, and the concentrations of mannose, galactose, glucuronic acid, ribose, arabinose and xylose were very different.\u003c/p\u003e \u003cp\u003eIt can be concluded that these fungi participate in the transformation of sugars and the formation of polysaccharides, which may influence and change the composition and functions of persimmon frost. This research provides a new field for screening fungi that produce extracellular polysaccharides and developing new functional polysaccharides.\u003c/p\u003e","manuscriptTitle":"Preliminary study on the function of natural persimmon frost fungi","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-04-23 05:02:50","doi":"10.21203/rs.3.rs-6276251/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"a6422f56-d920-4bc2-b39a-93d3f0f42189","owner":[],"postedDate":"April 23rd, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":47467252,"name":"Biological sciences/Microbiology"},{"id":47467253,"name":"Biological sciences/Molecular biology"}],"tags":[],"updatedAt":"2025-05-05T05:08:24+00:00","versionOfRecord":[],"versionCreatedAt":"2025-04-23 05:02:50","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-6276251","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6276251","identity":"rs-6276251","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}