1. Introduction
Cordyceps, which are widely recognized as medicinal mushrooms to improve human health and wellbeing, have been emerging as a miracle cure for many common ailments in Eastern and Chinese medicine [1,2,3]. Among them, the fungus Cordyceps militaris (CM), which grows as a parasite in Lepidopteran pupa and is yellow or orange in its fruiting body, is one of the members of this genus of Cordyceps. Based on historical and research evidence, the mycelium and fruiting body of CM are widely used in the pharmaceutical and health care industries as medical materials, dietary supplements, and food additives. The potential components found in the mycelia and fruiting bodies are cordycepin (Cor), polysaccharides, proteins, and phenolic compounds, which can be extremely effective to enhance oxygen mobilization, improve ATP synthesis, stabilize blood sugar metabolism, and actualize DNA repair [4,5,6,7,8]. Moreover, the health effects of compounds found in this species can positively assist the treatment of cancer and diabetes and enhance the human immune system [4]. Although bioactive compounds extracted from CM are preferably used in healthy foods and dietary supplements, Cor and Adenosine (Ado) (Figure 1) are among the most active compounds that can contribute to tumor cell apoptosis, a decrease in tumor cell proliferation, and several therapeutic potentials (antineoplastic, antioxidant, anti-inflammatory activities) [9].
Due to the precious properties it can bring to human health and the high demand of the medicine industry, the marketing requirements for this species are continuously increasing. However, hosts for the growth of CM are specialized and rare in nature; the natural development of this fungus happens slowly only in specific places and ends with a small fruiting body [10]. Several solid media such as rice, green beans, corn, sunflower seeds, and insects are reported as possible alternative protein sources to cultivate CM at both laboratory and industrial levels [11]. Chemical components in CM cultivated via these artificial methods, such as Cor, Ado, proteins, carotenoids, and trace minerals, are in line with those of the fungus grown naturally [12]. Bioactive compounds are mostly found in the fruiting body of this fungus. Thus, the fruiting body of CM is considered the most valuable part, which is usually collected and utilized as a food product for human consumption. The remaining part of CM, so-called “solid media” or “solid-based residues” (SBRs), is usually discarded as waste [13,14]. The unused SBRs of CM cause problems for the environment [15]. However, SBRs may contain bioactive substances [13]. A considerable amount of Cor was extracted, separated, and purified successfully from this waste medium of CM [13].
The idea of “green solvents” has been raised since the 1990s from the concept of green chemistry [16,17]. The environmental and health hazards of conventional solvents, such as benzene, carbon disulfide, carbon tetrachloride, diethyl ether, hexane, nitromethane, and ethylene glycol dimethyl ether, has required the use of less hazardous solvents [16,18]. Thus, several solvent selection guides have emerged from many organizations including GSK [19,20,21], ACS [22], Pfizer [23], and Sanofi [24]. Ethanol and water are among the most recommended extractants [19,20,21,22,23,24]. Although ethanol shares the same attributes as other alcohols (methanol, isopropanol), e.g., solvent strength, dielectric constant, critical point, and hydrogen-donation ability, it has become the favorable selection because it is safe [25], easily available in high purity, low in price, and completely biodegradable [26]. Ethanol is well known as an efficient solvent to extract polyphenols [27], flavonoids, and other polar compounds [28]. Meanwhile, water also plays an important role in green chemistry and is categorized as “green” because it is non-flammable, affordable, readily available, and, more importantly, non-toxic. Moreover, water is also a high polar solvent, having advantages regarding reactivity and selectivity, and thus it is favorable for organometallic catalysis [29]. A mixture of EtOH and water was found to be environmentally preferable [17].
In this study, two solvents, ethanol (EtOH) and water, and their combinations were used to evaluate optimum yields of Cor and Ado and relevant biological activities in relation to allelopathy, antioxidants, and xanthine oxidase (XOD). Allelopathy was determined according to the inhibitory levels of SBRs against the growth of R. sativus, L. sativa, and the two harmful weeds E. crus-galli and B. pilosa.
2. Results
2.1. Adenosine and Cordycepin Content in Cordyceps Militaris SBRs
A reversed-phase column was employed to quantify the content of the two bioactive compounds in the SBRs of CM. A good separation of Ado and Cor was conducted by the HPLC system integrated with a UV-Vis detector at a wavelength of 260 nm. Two sharp peaks of Ado and Cor were detected at the retention times of 9.110 min and 12.617 min in the standard chromatogram (Figure 2), respectively. Table 1 shows that Ado and Cor were detected and quantified in most of the extracts, except for W8–W10 and Wt where the contents of Ado were lower than LOD (0.274 µg/mL). With modifications of the solvent ratio of 10% (v/v), the levels of Ado and Cor varied unproportionately. The optimum yields of both Ado and Cor were recorded in fraction W4 (E:W = 6:4) and W6 (E:W = 4:6) when the content of these two compounds reached levels of 0.381 ± 0.002 mg/g and 0.561 ± 0.011 mg/g (W4), followed by 0.255 ± 0.001 mg/g and 0.511 ± 0.014 mg/g (W6), respectively. The highest content of Cor (16.515 ± 0.283 mg/g) was determined by hot water (Wt); however, Ado was not detected in this fraction.
2.2. Antioxidant Activities and In Vitro Inhibition of Xanthine Oxidase (XOD)
The free radical inhibitory activity of 13 extracts is presented in Table 2. It is obvious that the antioxidant activity of the SBR extracts was not proportional to the alteration of the solvent system ratio and varied unpredictably. The highest inhibitory activity of the extract was identified in two fractions, W4 and W6, where the IC50 of all three assays (DPPH, ABTS, and XOD) ranged from 118.5 ± 2.9 to 194.2 ± 4.9 µg/mL and from 118.2 ± 5.4 to 198.0 ± 7.7 µg/mL, respectively. These fractions confirmed a certain inhibition effect on oxidative agents due to their comparable values to the controls, including BHT (18.0 ± 0.3 and 40.0 ± 0.6 µg/mL) and allopurinol (20.8 ± 0.7 µg/mL). W10 was found to exert the lowest antioxidant activity, of which the IC50 ranged from 1103.5 ± 6.4 to 2007.0 ± 6.8 µg/mL. However, in the DPPH and ABTS assays, the inhibitory capability against oxidants of Wt was much higher than W10, revealing IC50 values of 413.5 ± 7.7 and 413.4 ± 6.7 µg/mL, respectively.
2.3. Screening Analysis of GC-MS
By GC-MS, major compounds in the SBRs relating to the different extracts were identified, and these are shown in Table 3. They included 14 compounds: 1H-pyrazole,1,3,5-trimethyl-; 1-tetradecanol; acetic acid; aqua cera; bis(2-ethylhexyl) phthalate; glycerin; hexadecanoic acid; methyl oleate; myristic acid; oxacyclododecan-2-one; phloroglucinol; ricinoleic acid; stearic acid; and trehalose. These chemicals belonged to disaccharides, fatty acid, and phenolic compounds. The presence of the chemicals varied among the extracting protocols, of which trehalose accounted for greater peak areas as compared to other putative constituents. Ricinoleic acid was found in W0–W6 and W7, whilst stearic acid was detected in W0, W2, W3, W4, and W5.
2.4. Growth-Inhibitory Activity on Plants of Cordyceps Militaris SBRs Extract
The inhibitory effect of the SBR extracts on the development of two indicator plants and two noxious weeds, expressed in IC50 values, is illustrated in Table 4. As can be seen, water played an important role in the solvent system to prove the weed-resistance activity of the SBR extracts. The MeOH extract fraction (W0) yielded the highest IC50 value for all four weed species, ranging from 438.0 ± 13.0 to 529.8 ± 15.4 μg/mL. In contrast, the inhibitory concentration of the solvent systems from W1 to W10 ranged below 178.3 ± 8.7 μg/mL, which was at least three times lower than W0. Among the 14 extracts, Wt exhibited the highest inhibitory activity against the growth of the two indicator plants and the two noxious weeds (IC50 value ranged from 54.9 ± 3.8 to 64.7 ± 1.0 μg/mL), followed by the extract W4 (IC50 value ranged from 64.7 ± 1.4 to 78.1 ± 3.9 μg/mL).
3. Discussion
Bioactive compounds, including Cor, Ado, polysaccharides, ergosterol, mannitol, and phenolic compounds, are reported to play important roles in pharmacological and medicinal activities of C. militaris [30]. Ado is regarded as the principal active compound in CM fruiting body cultured in China [31,32]. Meanwhile, Cor has shown capability in cancer cell inhibition [32]. In this study, these two compounds were both observed in the SBRs, although their doses varied among the applied solvents of EtOH-water and hot water. Pintathong et al. [15] employed barley, white rice, riceberry rice, and wheat to culture CM and obtained contents of Cor and Ado of between 0.78 and 1.09 mg/g dried weight [15]. Elsewhere, the concentration of Ado in CM waste medium was 0.47 mg/g DW [33]. The contents of Cor and Ado in the corpus of CM were 3.6 and 0.6 mg/g of CM, respectively [34]. Compared to the mentioned papers, the contents of Ado and Cor in this research (0.002–0.381 mg/g and 0.018–0.67 mg/g, respectively) were lower. The medium used to culture CM, which has a major effect on the yield of Ado and Cor from the fruiting body, may regulate the contents of Ado and Cor of this study [35,36].
Reactive oxygen species (ROS) is the main factor for oxidative stress to cause various diseases, such as hepatopathies, atherosclerosis, and aging [37,38]. The mechanism of antioxidants, which comprises their direct reaction with free radicals including hydrogen peroxide (H2O2), superoxide anion (O2•−) radicals, and hydroxyl (OH−) radicals, etc., to prevent the formation of peroxide, is the factor that mitigates the impact of those radicals on cells [33]. CM might be a good source to scavenge the activities of certain types of ROS, including xanthine oxidase (XOD), DPPH•, and ABTS• radicals [39,40,41]. In this study, hot water was the best medium to optimize the yields of the major constituents of CM, and it also exhibited the strongest antioxidant capacity. In addition, the combination of EtOH and water at the ratio of 4:6 (W6) was the ideal fraction to scavenge the activities of XOD, DPPH•, and ABTS•. The concentrations of SBRs to inhibit the three ROS were 185.2 ± 6.0 µg/mL, 118.2 ± 5.4 µg/mL, and 198.0 ± 7.7 µg/mL (IC50), respectively (Table 2). Wang et al. (2015) indicated that, at a concentration of 15 mg/mL, CM waste medium extract can inhibit 50% of TBARS formation, while the highest values of scavenging activity against DPPH• (within the range of 40.12 ± 0.85 to 66.62 ± 2.10 mg TEAC/g extract) were found in hot water extracts of SBRs [15,33].
Bioactive compounds such as polyphenols, flavonoids, polysaccharides, sterols, and peptides are associated with antioxidant capacity in CM extracts [42]. Polysaccharides are the most potential compounds in mushrooms to be responsible for their antioxidant properties [43,44]. Polysaccharides separated from CM decreased the level of malondialdehyde, which is considered an indicator for oxidant damage in cells, in mice blood [45,46]. The GC-MS analysis of this study revealed the presence of 14 compounds, of which trehalose, stearic acid, and methyl oleate have been described as increasing antioxidant activities [47]. The antioxidant activity, xanthine oxidase, and inhibitory activity of W4 and W6 may be largely attributed to these compounds. Trehalose can modify the structural conformation of the enzyme, therefore enhancing the activity of urate oxidase and improving its catalytic efficiency [48,49,50,51].
Allelochemicals from fungi have been described as having potential for weed control in agriculture [52,53,54]. The toxic effect of fungi on plant growth is driven by the variety and concentration of allelochemicals [55]. This study revealed that Wt and W4 exerted the maximum inhibitory effects on the germination, shoot height, and root length of two indicator plants and two noxious weeds, as compared to other fractions (Table 2). This suggests that Wt and W4 might contain certain allelochemical content relevant to inhibitory activity. Cor isolated from CM strongly inhibited the growth of radish seedlings [39]. Particularly, at a concentration of 0.04 mg/mL, Cor inhibited the germination and growth of R. sativus 3.8–5.9 and 3.3–3.7 times more than benzoic acid and glyphosate, respectively [56]. Meanwhile, the analysis of HPLC also confirmed the noticeable presence of Cor in Wt and W4. The immersion of EtOH provided an amount of Cor, but the highest yield of Cor was found in Wt (hot water treatment), followed by W4 and W6 (treatments of EtOH 60% and EtOH 40%, respectively) (Table 1). The amounts of Cor varied among the extracts and are proportional to the observed inhibitory activities against the tested plants (Table 1 and Table 3). Cor has been described as a potential compound to alter glyphosate, which has recently been debated and removed from the market in some countries [56].
4. Materials and Methods
4.1. Materials
SBRs of C. militaris were obtained from Center for Research and Development of Mushroom, Agricultural Genetics Institute (AGI), Vietnam in January 2021. The drying process of SBRs was conducted at 40 ± 1 °C for 3 days until SBR weight was constant. The dried SBRs were then ground into powders. An alteration of 10% of the solvent system (EtOH:water; E:W) was applied to each extraction procedure to identify the most active fraction. A 50 g sample was mixed with a solvent (500 mL) at a solid–liquid ratio of 1:10 w/v, then the obtained product was processed with an orbital shaker, conditioning at 125 rpm for 6 h. Exceptionally, hot water extraction was conducted with an incubator shaker, conditioned at 95 ± 1 °C with a rate of 125 rpm for 4 h. All experiments were replicated thrice. Then, the filtrate of the mixture was collected via a Whatman filter (no.1) in a vacuum filtration system. Solvent vaporization was conducted using a rotary evaporator to obtain the concentrated crude extract, which was then kept in MeOH and stored at −20 °C for further analysis.
4.2. Reagents
1,1-dipheynyl 2-picrylhydrazyl (DPPH), 2,2′-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), dimethyl sulfoxide (DMSO), monopotassium phosphate, dipotassium phosphate, sodium hydroxide, hydrochloric acid, xanthine, microbial xanthine oxidase, and allopurinol were purchased from Sigma-Aldrich Pte Ltd., Singapore. Analytical grade reagents were utilized for all experimentation.
4.3. Adenosine and Cordycepin Content in Cordyceps militaris SBRs
Quantification of Cor and Ado was performed by HPLC system, adapting the method of Li et al. [57]. In brief, 5 mL of SBR extract (in MeOH) was filtered through a hydrophilic filter (0.2 μm) before being injected into the HPLC system (Thermo UltiMate 3000, column Hypersil Gold 250 × 4.6 mm–5 µm). Two solvent systems, (A)—H2O, ammonium acetate 10 mM, acetic acid 0.1% and (B)—MeOH 90%, ammonium acetate 10 mM, acetic acid 0.1%, were employed for the gradient program. This included 3 phases: The first 20 min involved a gradient concentration of B (5% to 95%), which inclined to 100% in the next 5 min and decreased to 5% in the last 5 min. The HPLC system was conditioned with a wavelength of 260 nm, flow rate of 1.6 mL/min, column temperature of 40 °C, and sample temperature of 15 °C.
4.4. Antioxidant and Xanthine Oxidase Inhibition (XOD) Activities
DPPH (2,2-diphenyl-1-picrylhydrazyl) radical scavenging and ABTS (2,2′-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid)) radical cation decolorization assays were used to determine the antioxidant activity of all fractions [58,59]. Slight modifications from the reported spectrophotometric technique were used to measure the inhibitory effect on xanthine oxidase (XO) of all fractions [60].
4.5. GC-MS Screening Analysis
A GC-MS system (GC Trace 1310, Thermo Fisher Scientific, Waltham, USA) equipped with TriPlus RSH liquid autosampler and coupled with electron impact ionization-tandem mass spectrometry (TSQ 9000, Thermo Fisher Scientific, Waltham, USA) was employed to determine the chemical constituents of fractions from W0 to Wg [61]. A Thermo Scientific TG-5MS column (30 m × 0.25 mm ID× 0.25 μm in thickness) was utilized for chemical separation. Splitless mode was selected for the liquid autosampler, of which the injection volume and temperature were set at 1 µL and 280 °C, respectively. The carrier gas was helium (99.999% purity), and the flow rate was maintained at 1.0 mL min−1 throughout separation time. The temperature program of the GC oven was as follows: A hold time of 3 min and an initial temperature of 80 °C were set for the first stage. Then, the temperature was increased to 200 °C with a rate of 15 °C min−1 before continuously rising to 300 °C with a lower rate of 8 °C min−1. Finally, a temperature of 300 °C was held for 5 min to end the program. Constant temperatures of 280 °C and 150 °C were used for the electron impact ionization source and quadrupole, respectively. Volatile compound detection was performed by positive EI-MS/MS under MRM mode and at an energy ionization level of 70 eV.
4.6. Growth-Inhibitory Activity Bioassays
Echinochloa crus-galli, Bidens pilosa L., Raphanus sativus L., and Lactuca sativa L. seeds were selected as experimental subjects to evaluate the growth-constraint potential of the SBR extracts. The seeds were germinated in a growth chamber, which was conditioned with a photoperiodic setting of 12/12 h (day/night) and corresponding temperatures of 25/23 °C. The preparation of the SRB-derived test solution was conducted by diluting the extract in Tween water (0.2%) to attain different concentrations (10, 100, 500, 1000, and 2000 μg/mL). An aliquot of the SRB solution (100 μL) was dropped through filter papers laid on top of a 12-well plate (22 mm diameter × 18 mm height of each well). The selected seeds were put in corresponding wells filled with 100 μL of distilled water. Changes in length of the root and shoot were monitored and recorded daily for 7 days. Six replications were applied in this bioassay. The half-maximal concentration (IC50) of the SBRs inhibiting shoot and root growth was also determined following the previous method [62].
4.7. Statistical Analysis
Results are presented as mean ± standard deviation (SD). Duncan’s test (α = 5%) was employed to present significant differences in the results. Moreover, significant differences of the solvent system were determined by one-way analysis of variance (ANOVA) using the statistical software Minitab®19.2020.1 (Philadelphia, PA, USA).
5. Conclusions
This study optimized the quantification of cordycepin (Cor) and adenosine (Ado) in solid-based residues of C. militaris using different combinations of EtOH-water and hot water and examined the relevant potential of antioxidant activities, antihyperuricemia, and plant growth-inhibitory activities. The combination of 60% EtOH-40% water (W4) obtained the highest amounts of Cor and Ado and the strongest levels of the examined biological activities. Hot water (Wt) displayed the maximum inhibition on the growth of lettuce, radish, and the noxious weeds E. crus-galli and B. pilosa. From this study, SBRs of C. militaris appear to be a potential source to exploit its potential in antioxidant and xanthine oxidase activity as well as in weed control.
Conceptualization and methodology, T.N.M.; formal analysis, L.V.A. and T.N.M.; investigation, N.Q.T. and B.Q.M.; resources, T.N.M. and L.V.A.; writing—original draft preparation, L.V.A. and T.N.M.; writing—review and editing, T.D.X.; supervision, T.D.X. and N.Q.T. All authors have read and agreed to the published version of the manuscript.
Not applicable.
Not applicable.
Not applicable.
Center for Research and Technology Transfer, Vietnam is appreciated for partly supporting this research. We thank Do Hoang Giang and Nguyen Tien Dat for helpful advice on this research.
The authors declare no conflict of interest.
Footnotes
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Enlarge this image.Figure 2. Chromatogram of Cor, Ado (a) and their ultraviolet-visible spectrophotometry (b,c).
Adenosine and cordycepin content in Cordyceps militaris SBRs extracted by different solvent systems.
| Extracts | Code | Adenosine |
Cordycepin |
|---|---|---|---|
| EtOH | W0 | 0.002 ± 0.0003 d | 0.018 ± 0.0004 d |
| E:W = 9:1 | W1 | 0.029 ± 0.001 d | 0.25 ± 0.008 c |
| E:W = 8:2 | W2 | 0.096 ± 0.001 c | 0.298 ± 0.012 c |
| E:W = 7:3 | W3 | 0.092 ± 0.001 c | 0.278 ± 0.007 c |
| E:W = 6:4 | W4 | 0.381 ± 0.002 a | 0.561 ± 0.011 b |
| E:W = 5:5 | W5 | 0.089 ± 0.0004 c | 0.251 ± 0.008 c |
| E:W = 4:6 | W6 | 0.255 ± 0.001 b | 0.511 ± 0.014 bc |
| E:W = 3:7 | W7 | 0.074 ± 0.0004 c | 0.316 ± 0.014 c |
| E:W = 2:8 | W8 | nd | 0.26 ± 0.008 c |
| E:W = 1:9 | W9 | nd | 0.212 ± 0.001 c |
| Water | W10 | nd | 0.423 ± 0.007 c |
| Hot water | Wt | nd | 0.67 ± 0.012 a |
| Glue | Wg | nd | nd |
| HPLC | Standards | ||
| Retention time (min) | - | 9.117 ± 0.108 | 12.617 ± 0.122 |
| LOD (µg/mL) | - | 0.274 | 0.366 |
| LOQ (µg/mL) | - | 0.831 | 1.110 |
Values represent means ± SD (standard deviation). Alphabetic letters indicate statistical symbols generated by one-way ANOVA using Duncan’s test (α = 5%). Values with similar letters in each column are not significantly different (p < 0.05) (n = 3). LOD: limit of detection; LOQ: limit of quantification; nd: not detectable; ‘-’: not available.
Antioxidant activity of Cordyceps militaris SBRs extracted by different solvent systems.
| Extract | Code | IC50 (µg/mL) | IC50 (µg/mL) | |
|---|---|---|---|---|
| DPPH | ABTS | XOD | ||
| EtOH | W0 | 1199.9 ± 6.0 b | 1227.8 ± 8.2 b | 782.2 ± 4.6 b |
| E:W = 9:1 | W1 | 1182.2 ± 7.8 b | 1246.0 ± 7.5 b | 731.6 ± 7.2 b |
| E:W = 8:2 | W2 | 411.5 ± 7.4 c | 457.4 ± 4.9 c | 451.7 ± 10.0 c |
| E:W = 7:3 | W3 | 424.1 ± 7.3 c | 459.9 ± 5.8 c | 448.5 ± 9.9 c |
| E:W = 6:4 | W4 | 118.5 ± 2.9 d | 194.2 ± 4.9 d | 186.0 ± 1.9 d |
| E:W = 5:5 | W5 | 225.2 ± 5.3 cd | 234.1 ± 3.1 cd | 450.8 ± 3.7 cd |
| E:W = 4:6 | W6 | 118.2 ± 5.4 d | 198.0 ± 7.7 d | 185.2 ± 6.0 d |
| E:W = 3:7 | W7 | 851.2 ± 7.8 bc | 846.6 ± 7.2 bc | 466.8 ± 10.0 bc |
| E:W = 2:8 | W8 | 1234.8 ± 8.7 b | 1230.3 ± 6.7 b | 778.2 ± 13.6 b |
| E:W = 1:9 | W9 | 1232.5 ± 9.5 b | 1246.2 ± 8.7 b | 750.5 ± 5.0 b |
| Water | W10 | 1964.3 ± 8.0 a | 2007.0 ± 6.8 a | 1103.5 ± 6.4 a |
| Hot water | Wt | 413.5 ± 7.7 c | 413.4 ± 6.7 c | 1190.7 ± 7.7 c |
| Glue | Wg | nd | nd | nd |
| BHT | - | 18.0 ± 0.3 e | 40.0 ± 0.6 e | - |
| Allopurinol | - | - | - | 20.8 ± 0.7 e |
Values represent means ± SD (standard deviation). Alphabetic letters indicate statistical symbols generated by one-way ANOVA using Duncan’s test (α = 5%). Values with similar letters in each column are not significantly different (p < 0.05) (n = 3). BHT: Butylated hydroxytoluene; nd: not detectable; ‘-’: not available.
Principal compounds identified in Cordyceps militaris SBRs by GC-MS screening analysis.
| Extract | Codes | Major Constituents | Retention Times (min) | Peak Area (%) |
|---|---|---|---|---|
| EtOH | W0 | Ste, Tre, |
20.30, 21.45, 22.94, 23.70, 26.56, 27.40 | 24.79, 7.33, 36.12, 8.69, 8.8, 4.22 |
| E:W = 9:1 | W1 | Gly, Tre, Met, Hex, Ric | 20.80, 22.65, 22.98, 23.22, 23.72, | 4.94, 5.76, 28.9, 34.52, 7.79, 0 |
| E:W = 8:2 | W2 | Aqu, Tre, Gly, Hex, Ric, Ste, Met | 20.31, 20.79, 21.45, 23.28, 23.73, 25.60, 29.04 | 11.58, 1.81, 1.27, 18.49, 7.88, 7.98, 45.95 |
| E:W = 7:3 | W3 | Aqu, Tre, Gly, Ric, Hex, Ste, Met | 20.32, 20.81, 21.48, 22.89, 23.29, 23.74, 29.15 | 14.07, 6.76, 1.25, 16.44, 13.14, 4.57, 38.56 |
| E:W = 6:4 | W4 | Tre, Gly, Ric, Hex, Ste, Met | 20.82, 21.49, 22.82, 23.27, 25.65, 29.16 | 19.22, 10.23, 16.68, 17.26, 5.4, 18.93 |
| E:W = 5:5 | W5 | Ace, Tre, Gly, |
11.93, 20.85, 21.53, 22.95, 23.25, 25.62, 29.12 | 2.83, 3.86, 3.25, 12.85, 23.61, 5.18, 35.78 |
| E:W = 4:6 | W6 | Ace, Tre, Gly, Ric, Met | 11.91, 20.83, 21.50, 22.56, 28.35 | 3.67, 7.75, 9.24, 23.34, 24.19 |
| E:W = 3:7 | W7 | Ace, Tre, Gly, Ric, Hex, 1-T | 11.91, 20.83, 21.51, 22.19, 22.86, 23.85 | 4.57, 14.37, 9.76, 32.22, 7.06, 6.28 |
| E:W = 2:8 | W8 | Ace, Tre, Gly, 1-T | 11.88, 20.80, 21.50, 23.83 | 6.09, 15.61, 10.21, 9.41 |
| E:W = 1:9 | W9 | Ace, 1H-, Tre, Gly, Hex, Phl | 11.92, 13.44, 20.83, 21.52, 22.74, 23.88 | 8.18, 3.85, 18.19, 25.46, 5.13, 32.83 |
| Water | W10 | Ace, Tre, Gly, 1-T | 11.95, 20.86, 21.56, 23.90 | 3.28, 8.06, 12.26, 7.69 |
| Hot water | Wt | Ace, Tre, Gly | 11.89, 20.79, 21.48 | 15.43, 8.49, 18.34 |
| Glue | Wg | Tre | 20.77 | 30.16 |
1H-Pyrazole,1,3,5-trimethyl-: 1H-; 1-Tetradecanol: 1-T; acetic acid: Ace; aqua cera: Aqu; bis(2-ethylhexyl) phthalate: Bis; glycerin: Gly; hexadecanoic acid: Hex; methyl oleate: Met; myristic acid: Myr; oxacyclododecan-2-one: Oxa; phloroglucinol: Phl; ricinoleic acid: Ric; stearic acid: Ste; trehalose: Tre.
Inhibitory effects of different Cordyceps militaris SBR extracts on the growth of R. sativus, E. crus-galli, L. sativa, and B. pilosa L.
| Extract |
R. sativus |
E. crus-galli |
L. sativa |
B. pilosa L. |
||||
|---|---|---|---|---|---|---|---|---|
| Shoot | Root | Shoot | Root | Shoot | Root | Shoot | Root | |
| W0 | 512.2 ± 10.8 b | 529.8 ± 15.4 b | 459.6 ± 16.3 b | 438.0 ± 13.0 b | 449.7 ± 13.5 b | 439.7 ± 3.7 b | 522.5 ± 10.5 b | 523.3 ± 12.2 b |
| W1 | 163.0 ± 5.6 c | 172.8 ± 2.6 c | 151.0 ± 3.5 c | 147.5 ± 3.7 c | 150.7 ± 6.7 c | 140.9 ± 7.8 c | 178.3 ± 8.7 c | 172.1 ± 8.8 c |
| W2 | 125.4 ± 2.0 c | 126.4 ± 4.5 c | 114.7 ± 4.3 c | 108.4 ± 3.7 c | 110.4 ± 3.8 c | 108.0 ± 3.5 c | 128.5 ± 2.6 c | 129.9 ± 5.9 c |
| W3 | 122.4 ± 1.5 c | 119.8 ± 5.8 c | 113.3 ± 1.5 c | 108.8 ± 2.5 c | 113.2 ± 1.8 c | 106.7 ± 5.7 c | 127.9 ± 7.5 c | 123.4 ± 4.3 c |
| W4 | 77.5 ± 4.4 d | 78.1 ± 3.9 d | 73.1 ± 3.4 d | 64.7 ± 1.4 d | 71.9 ± 1.5 d | 65.6 ± 0.8 d | 77.3 ± 1.2 d | 76.4 ± 3.2 d |
| W5 | 119.8 ± 3.5 c | 128.9 ± 5.1 c | 115.6 ± 5.3 c | 106.8 ± 2.4 c | 109.8 ± 4.9 c | 107.5 ± 2.3 c | 121.7 ± 5.1 c | 127.9 ± 2.2 c |
| W6 | 79.6 ± 1.0 d | 82.0 ± 1.3 d | 76.0 ± 3.9 d | 70.7 ± 0.8 d | 74.4 ± 0.6 d | 67.7 ± 2.4 d | 86.3 ± 4.1 d | 85.5 ± 3.7 d |
| W7 | 116.8 ± 2.6 c | 121.5 ± 5.6 c | 111.4 ± 1.3 c | 105.5 ± 4.2 c | 107.6 ± 2.6 c | 101.4 ± 6.4 c | 119.6 ± 3.4 c | 122.6 ± 4.1 c |
| W8 | 148.7 ± 6.1 c | 152.0 ± 6.6 c | 136.1 ± 4.9 c | 129.9 ± 5.1 c | 130.5 ± 6.6 c | 126.5 ± 8.8 c | 154.7 ± 1.5 c | 154.7 ± 1.7 c |
| W9 | 164.0 ± 5.0 c | 175.4 ± 9.0 c | 157.2 ± 9.8 c | 145.3 ± 0.7 c | 148.8 ± 6.0 c | 147.2 ± 10.4 c | 176.9 ± 11.7 c | 174.7 ± 1.7 c |
| W10 | 99.1 ± 1.6 cd | 106.2 ± 5.0 cd | 94.6 ± 3.7 cd | 84.5 ± 1.6 cd | 87.4 ± 3.8 cd | 84.6 ± 5.8 cd | 101.6 ± 2.8 cd | 100.4 ± 6.3 cd |
| Wt | 62.0 ± 1.8 d | 63.4 ± 1.0 d | 57.4 ± 0.7 d | 54.5 ± 2.8 d | 56.9 ± 1.9 d | 54.9 ± 3.8 d | 64.7 ± 1.0 d | 63.6 ± 3.5 d |
| Wg | 1025.1 ± 28.6 a | 1074.7 ± 24.2 a | 950.3 ± 27.7 a | 834.6 ± 22.5 a | 915.2 ± 25.3 a | 844.2 ± 12.9 a | 1070.0 ± 28.5 a | 1054.9 ± 21.7 a |
Values represent means ± SD (standard deviation). Alphabetic letters indicate statistical symbols generated by one-way ANOVA using Duncan’s test (α = 5%). Values with similar letters in each column are not significantly different (p < 0.05) (n = 3).
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; Le, Viet Anh 1
; Nguyen, Quang Trung 1
; Bui, Quang Minh 1
; Tran, Dang Xuan 2
1 Center for Research and Technology Transfer, Vietnam Academy of Science and Technology (VAST), Hanoi 122100, Vietnam
2 Transdisciplinary Science and Engineering Program, Graduate School of Advanced Science and Engineering, Hiroshima University, Hiroshima 739-8529, Japan; Center for the Planetary Health and Innovation Science (PHIS), the IDEC Institute, Hiroshima University, Hiroshima 739-8529, Japan