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<p>This study focuses on the production of bioactive secondary metabolites (SMs) with antifungal potential against <i>Macrophomina phaseolina</i> using microbial solid-state fermentation (SSF) technology. Two <i>Trichoderma</i> strains (MJ1 and MJ3) were propagated to bioprocessing with agro-industrial substrates (broken white rice). Solid: liquid partition (1:3) was used for extraction with solvent ethyl acetate (EtOAc), and a rotary vacuum evaporator facilitated solvent evaporation and compound collection. The crude extract was dissolved in DMSO. While optimal conditions for enhanced metabolite yield and antifungal activity were identified using both the one-factor-at-a-time (OFAT) and the Box–Behnken design (BBD) of response surface methodology (RSM). OFAT experimentations were applied using critical physical growth parameters (GPs) such as incubation days (3rd, 5th, 7th, 10th, 13th, and 17th day), temperature (22&#xa0;°C, 25&#xa0;°C, 28&#xa0;°C, 30&#xa0;°C, and 32&#xa0;°C), initial moisture content % (50%, 70%, and 80%), and pH (5, 6, 7, 8, 9, and 10). Based on the observations of OFAT, a BBD matrix consisting of 27 experimental trials was generated as combinations of selected GPs by Design–Expert software. For MJ1, comparing the results obtained from OFAT and RSM with the highest value obtained in the zone of inhibition (mm) (ZOI) and yield (mg/mL), the percentage improvement calculated as ZOI is 60% in the RSM optimization compared to OFAT and yield is 27% in RSM compared to OFAT. For MJ3 inhibition zone improved by 20% in the RSM optimization compared to OFAT and yield improved by 34.81% in RSM compared to OFAT. GC–MS analysis revealed 22 compounds in MJ1 and 19 compounds in MJ3. The dominant compounds included 2,3-butanediol (5.5117%), phenylethyl alcohol (2.3171%), 6-pentyl-2H-pyran-2-one (1.0935%), trans-alpha-alpha-bergamotene (3.1030%), n-hexadecanoic acid (22.1247%), 9,12-octadecadienoic acid (Z,Z)- (56.0125%), and eicosanoic acid (0.7213%). Majority of the SMs exhibit reported bioactivity against <i>M. phaseolina</i>, whereas phenylethyl alcohol and 6-pentyl-2H-pyran-2-one have not been reported for the same. This study demonstrates that the bioprocessing using broken white rice as substrate with <i>Trichoderma</i> strains, coupled with statistical modeling, determines that the RSM approach is significantly more effective in enhancing both antifungal activity and yield and thus promotes antifungal SM production. Furthermore, it suggests the feasibility of industrial scale-up using cost-effective substrates. The present study indicates that MJ1 and MJ3 give better antifungal activity under optimized GPs and thus produce antifungal specific SMs, which were analyzed by the GC–MS technique and concludes their antagonistic nature towards pathogen <i>M. phaseolina</i>.</p>

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Response surface methodology: a comparative optimization of antifungal metabolite production by Trichoderma viride and Trichoderma harzianum using solid-state fermentation

  • 期刊论文

  • 发表日期：

    2025年2月5日

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  • Jasleen Kaur,
  • Dweipayan Goswami,
  • Meenu Saraf

摘要

This study focuses on the production of bioactive secondary metabolites (SMs) with antifungal potential against Macrophomina phaseolina using microbial solid-state fermentation (SSF) technology. Two Trichoderma strains (MJ1 and MJ3) were propagated to bioprocessing with agro-industrial substrates (broken white rice). Solid: liquid partition (1:3) was used for extraction with solvent ethyl acetate (EtOAc), and a rotary vacuum evaporator facilitated solvent evaporation and compound collection. The crude extract was dissolved in DMSO. While optimal conditions for enhanced metabolite yield and antifungal activity were identified using both the one-factor-at-a-time (OFAT) and the Box–Behnken design (BBD) of response surface methodology (RSM). OFAT experimentations were applied using critical physical growth parameters (GPs) such as incubation days (3rd, 5th, 7th, 10th, 13th, and 17th day), temperature (22 °C, 25 °C, 28 °C, 30 °C, and 32 °C), initial moisture content % (50%, 70%, and 80%), and pH (5, 6, 7, 8, 9, and 10). Based on the observations of OFAT, a BBD matrix consisting of 27 experimental trials was generated as combinations of selected GPs by Design–Expert software. For MJ1, comparing the results obtained from OFAT and RSM with the highest value obtained in the zone of inhibition (mm) (ZOI) and yield (mg/mL), the percentage improvement calculated as ZOI is 60% in the RSM optimization compared to OFAT and yield is 27% in RSM compared to OFAT. For MJ3 inhibition zone improved by 20% in the RSM optimization compared to OFAT and yield improved by 34.81% in RSM compared to OFAT. GC–MS analysis revealed 22 compounds in MJ1 and 19 compounds in MJ3. The dominant compounds included 2,3-butanediol (5.5117%), phenylethyl alcohol (2.3171%), 6-pentyl-2H-pyran-2-one (1.0935%), trans-alpha-alpha-bergamotene (3.1030%), n-hexadecanoic acid (22.1247%), 9,12-octadecadienoic acid (Z,Z)- (56.0125%), and eicosanoic acid (0.7213%). Majority of the SMs exhibit reported bioactivity against M. phaseolina, whereas phenylethyl alcohol and 6-pentyl-2H-pyran-2-one have not been reported for the same. This study demonstrates that the bioprocessing using broken white rice as substrate with Trichoderma strains, coupled with statistical modeling, determines that the RSM approach is significantly more effective in enhancing both antifungal activity and yield and thus promotes antifungal SM production. Furthermore, it suggests the feasibility of industrial scale-up using cost-effective substrates. The present study indicates that MJ1 and MJ3 give better antifungal activity under optimized GPs and thus produce antifungal specific SMs, which were analyzed by the GC–MS technique and concludes their antagonistic nature towards pathogen M. phaseolina.