Chemical Composition, Nutritional, and Biological Properties of Extracts Obtained with Different Techniques from Aronia melanocarpa Berries

Abstract

This study investigates the chemical composition, nutritional, and biological properties of extracts obtained from A. melanocarpa berries using different extraction methods and solvents. Hydrodistillation and supercritical fluid extraction with CO₂ allowed us to isolate fruit essential oil (HD_EX) and fixed oil (SFE_EX), respectively. A phenol-enriched extract was obtained using a mild ultrasound-assisted maceration with methanol (UAM_M). The HD_EX most abundant component, using gas chromatography-mass spectrometry (GC/MS), was italicene epoxide (17.2%), followed by hexadecanoic acid (12.4%), khusinol (10.5%), limonene (9.7%), dodecanoic acid (9.7%), and (E)-anethole (6.1%). Linoleic (348.9 mg/g of extract, 70.5%), oleic (88.9 mg/g, 17.9%), and palmitic (40.8 mg/g, 8.2%) acids, followed by α-linolenic and stearic acids, were the main fatty acids in SFE_EX determined using high-performance liquid chromatography coupled with a photodiode array detector and an evaporative light scattering detector (HPLC-DAD/ELSD). HPLC-DAD analyses of SFE_EX identified β-carotene as the main carotenoid (1.7 mg/g), while HPLC with fluorescence detection (FLU) evidenced α-tocopherol (1.2 mg/g) as the most abundant tocopherol isoform in SFE_EX. Liquid chromatography-electrospray ionization-MS (LC-ESI-MS) analysis of UAM_M showed the presence of quercetin-sulfate (15.6%, major component), malvidin 3-O-(6-O-p-coumaroyl) glucoside-4-vinylphenol adduct (pigment B) (9.3%), di-caffeoyl coumaroyl spermidine (7.6%), methyl-epigallocatechin (5.68%), and phloretin (4.1%), while flavonoids (70.5%) and phenolic acids (23.9%) emerged as the most abundant polyphenol classes. UAM_M exerted a complete inhibition of the cholesterol oxidative degradation at 140 °C from 75 μg of extract, showing 50% protection at 30.6 μg (IA₅₀). Furthermore, UAM_M significantly reduced viability (31-48%) in A375 melanoma cells in the range of 500-2000 μg/mL after 96 h of incubation (MTT assay), with a low toxic effect in normal HaCaT keratinocytes. The results of this research extend the knowledge of the nutritional and biological properties of A. melanocarpa berries, providing useful information on specific extracts for potential food, cosmetic, and pharmaceutical applications.

Keywords: Aronia melanocarpa; berries; essential oil; fixed oil; natural extracts; phenolic extract.

Scheme 1

Scheme of preparation of HD_EX, SFE_EX, UAM_M, and UAM_H extracts obtained from dried berries of A. melanocarpa using hydrodistillation in a Clevenger-type apparatus, supercritical CO₂ extraction, and ultrasound-assisted maceration with MeOH and n-hexane, respectively.

Figure 1

GC-MS chromatogram, obtained on an HP-5ms capillary column, of A. melanocarpa fruit essential oil (HD_EX), with the indication of the main volatile compounds as reported in Table 1.

Figure 2

Fatty acid (FA) chromatographic profiles (HPLC analysis) obtained using DAD (200 nm, unsaturated FA) and ELSD (saturated FA) detection of A. melanocarpa fruit oil extracts SFE_EX (a) and UAM_H (b). FA composition (expressed as mg/g of oil extract) determined using HPLC-DAD/ELSD analysis of A. melanocarpa SFE_EX and UAM_H oil extracts after saponification (c). Data are expressed as mean values ± standard deviations (SD) (n = 4); ** = p < 0.01 versus SFE_EX (Student’s unpaired t-test with Welch’s correction).

Figure 3

Quali-quantitative analysis of SFE_EX carotenoids (a) and tocopherols (b) using HPLC-DAD (474 nm) and HPLC-FLU (λ_ex 295 nm, λ_em 330 nm) analysis.

Figure 4

(a) Antioxidant activity (% protection) of different amounts (2.5–100 μg) of A. melanocarpa UAM_M berry extract measured during cholesterol oxidation at 140 °C for 1 h. Three independent experiments were performed, and data are presented as mean and standard deviation (n = 3); *** = p < 0.001, ** = p < 0.01, * = p < 0.05 versus oxidized controls (0% protection). (b) Values of oxysterols 7β-OH and 7-keto (expressed as μg) measured in the controls (Ctrl) and in oxidized samples in the absence (0) or in the presence of different amounts (2.5–100 μg) of UAM_M during cholesterol oxidation; data are presented as mean ± SD (n = 3). *** = p < 0.001, ** = p < 0.01, * = p < 0.05 versus Ctrl; °°° = p < 0.001, versus oxidized controls (0). Statistical analyses are performed with one-way ANOVA and Bonferroni post test.

Figure 5

Viability, expressed as % of the control (Ctrl), induced using incubation for 96 h with different amounts (10–2000 μg/mL) of A. melanocarpa UAM_M berry extract in A375 human melanoma cells and human HaCaT keratinocytes (MTT assay). Data are presented as mean and standard deviation (n = 12). *** = p < 0.001, ** = p < 0.01 versus respective Ctrl (statistical analyses were performed using one-way ANOVA and Bonferroni post test). For each concentration group: ^### = p < 0.001, ^## = p < 0.01, ^# = p < 0.05, for A375 cells versus HaCaT cells (Student’s unpaired t-test with Welch’s correction).