Design and Evaluation of a Inonotus obliquus-AgNP-Maltodextrin Delivery System: Antioxidant, Antimicrobial, Acetylcholinesterase Inhibitory and Cytotoxic Potential

Abstract

Inonotus obliquus, a medicinal mushroom valued for its bioactive compounds, has not been previously characterized from Romanian sources. This study presents the first comprehensive chemical and biological screening of I. obliquus, introducing novel polymer-based encapsulation systems to enhance the stability and bioavailability of its bioactive constituents. Two distinct delivery systems were designed to enhance the functionality of I. obliquus extracts: (i) microencapsulation in maltodextrin (MIO) and (ii) a sequential approach involving preparation of silver nanoparticle-loaded I. obliquus (IO-AgNPs), followed by microencapsulation to yield the hybrid MIO-AgNP system. Comprehensive metabolite profiling using GC-MS and ESI-QTOF-MS revealed 142 bioactive constituents, including terpenoids, flavonoids, phenolic acids, amino acids, coumarins, styrylpyrones, fatty acids, and phytosterols. Structural integrity and successful encapsulation were confirmed by XRD, FTIR, and SEM analyses. Both IO-AgNPs and MIO-AgNPs demonstrated potent antioxidant activity, significant acetylcholinesterase inhibition, and robust antimicrobial effects against Staphylococcus aureus, Bacillus cereus, Pseudomonas aeruginosa, and Escherichia coli. Cytotoxicity assays revealed pronounced activity against MCF-7, HCT116, and HeLa cell lines, with MIO-AgNPs exhibiting superior efficacy. The synergistic integration of maltodextrin and AgNPs enhanced compound stability and bioactivity. As the first report on Romanian I. obliquus, this study highlights its therapeutic potential and establishes polymer-based nanoencapsulation as an effective strategy for optimizing its applications in combating microbial resistance and cancer.

Keywords: Inonotus obliquus; anti-acetylcholinesterase activity; antimicrobial screening; antioxidant potential; in vitro cytotoxicity; micro-spray encapsulation; silver nanoparticles.

Figure 1

Total ion chromatogram of I. obliquus sample.

Figure 2

The mass spectrum of I. obliquus sample.

Figure 3

The VOC sensory profile of the constituents identified in the I. obliquus sample. VOC: Volatile organic compound.

Figure 4

FTIR spectra of I. obliquus sample (black line), IO–AgNPs (green line), MIO (red line), and MIO–AgNPs (blue line) systems. FTIR: Fourier-transform infrared; IO–AgNPs: I. obliquus–silver nanoparticles; MIO: Maltodextrin—I. obliquus; MIO–AgNPs: Maltodextrin—I. obliquus–silver nanoparticles.

Figure 5

XRD patterns of the I. obliquus sample (black line) and the IO–AgNP system (red line). XRD: X-ray diffraction.

Figure 6

SEM micrograph of I. obliquus sample (a), IO–AgNPs (b), MIO (c), and MIO–AgNPs (d) systems. SEM: Scanning electron microscopy.

Figure 7

EDX analysis of the I. obliquus sample (a) and IO–AgNP system (b). EDX: Energy-dispersive X-ray.

Figure 8

DLS pattern of I. obliquus sample (red curve) and IO–AgNP system (green curve). DLS: Dynamic light scattering.

Figure 9

PSD curves from 10 consecutive measurements conducted over a two-minute period for MIO (a) and MIO–AgNPs (b) systems. PSD: Particle size distribution.

Figure 10

Comparative TG (a), DTG (b), and HF (c) thermoanalytical curves of I. obliquus sample (black line), IO–AgNPs (green line), MIO (red line), and MIO–AgNPs (blue line) systems. DTG: Differential thermogravimetry; HF: Heat flow; TG: Thermogravimetry.

Figure 11

Results of TPC (a), FRAP (b), and DPPH (c) assays for the I. obliquus extract, IO–AgNPs, MIO, and MIO–AgNP systems. Data are presented as mean ± SD (n = 3). Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test to compare samples (* p < 0.05). SD, standard deviation.

Figure 11

Results of TPC (a), FRAP (b), and DPPH (c) assays for the I. obliquus extract, IO–AgNPs, MIO, and MIO–AgNP systems. Data are presented as mean ± SD (n = 3). Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test to compare samples (* p < 0.05). SD, standard deviation.

Figure 12

Results of AChE inhibitory assay for of I. obliquus sample, IO–AgNPs, MIO, MIO–AgNP systems and AgNPs. Data are presented as mean ± SD (n = 3). Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test (* p < 0.05, ** p < 0.01 vs. control). AChE: Acetylcholinesterase.

Figure 13

Viability of MCF-7, HCT116, and HeLa cell lines, assessed at 24, 48, and 72 h after co-incubation with varying concentrations (75–200 μg/mL) of I. obliquus sample (a), IO–AgNPs (b), MIO (c), and MIO–AgNPs (d) systems. Negative control wells included untreated cells, while positive control wells included cells treated with a known cytotoxic agent; MTT solution and DMSO were used in the assay. Data are presented as mean ± SD (n = 3). Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test (* p < 0.05, ** p < 0.01 vs. control). DMSO: Dimethyl sulfoxide; MTT: 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide.

Figure 13

Viability of MCF-7, HCT116, and HeLa cell lines, assessed at 24, 48, and 72 h after co-incubation with varying concentrations (75–200 μg/mL) of I. obliquus sample (a), IO–AgNPs (b), MIO (c), and MIO–AgNPs (d) systems. Negative control wells included untreated cells, while positive control wells included cells treated with a known cytotoxic agent; MTT solution and DMSO were used in the assay. Data are presented as mean ± SD (n = 3). Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test (* p < 0.05, ** p < 0.01 vs. control). DMSO: Dimethyl sulfoxide; MTT: 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide.

Figure 14

IC₅₀ values of in vitro cytotoxicity of I. obliquus extract (IO), citrate-coated AgNPs, IO–AgNPs, MIO, and MIO–AgNP systems against MCF-7, HCT116, and HeLa cancer cell lines, assessed by MTT assay. Untreated cells served as the control. Data are presented as mean ± SD (n = 3). Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test (* p < 0.05, ** p < 0.01 vs. control).