Polyphenol-Enriched Extracts of Prunus spinosa Fruits: Anti-Inflammatory and Antioxidant Effects in Human Immune Cells Ex Vivo in Relation to Phytochemical Profile
Abstract
:1. Introduction
2. Results
2.1. LC-PDA-ESI-MS3 Metabolite Profiling
2.1.1. Phenolic Acids and Aldehydes
2.1.2. Flavonoids
2.1.3. Anthocyanins
2.2. Quantitative Profile of the Extracts/Fractions
2.3. Influence on Cells Viability
2.4. Antioxidant Effect: Influence on ROS Production by Neutrophils
2.5. Inhibition of ELA-2 Release by Neutrophils
2.6. Inhibition of IL-8 and TNF-α Production by Human Neutrophils
2.7. Influence on TNF-α, IL-6 and IL-10 Production by Human PBMCs
3. Discussion
4. Materials and Methods
4.1. Plant Material
4.2. Extracts Preparation
4.3. Phytochemical Profiling
4.3.1. Qualitative LC-MS/MS Profiling
4.3.2. Quantitative Profiling
4.4. Antioxidant and Anti-Inflammatory Activity in Cellular Models
4.4.1. Isolation of Human Neutrophils and PBMCs
4.4.2. Neutrophils and PBMCs Viability
4.4.3. ROS Generation by Human Neutrophils
4.4.4. ELA-2 Release by Human Neutrophils
4.4.5. Secretion of IL-8, IL-6, TNF-α and IL-10 by Immune Cells
4.5. Statistics and Data Analysis
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
Sample Availability
Abbreviations
BFF | n-butanol fraction of MEF |
CHA | chlorogenic acid (5-O-caffeoylquinic acid) |
CYG | cyanidin 3-O-β-d-glucopyranoside |
DEFF | diethyl ether fraction of MEF |
DEX | dexamethasone |
dw | dry weight |
EAFF | ethyl acetate fraction of MEF |
ELA-2 | elastase 2 (neutrophils elastase) |
fMLP | N-formyl-l-methionyl-l-leucyl-l-phenylalanine |
FRAP | ferric reducing antioxidant power |
fw | fresh weight |
GAE | gallic acid equivalents |
IBD | inflammatory bowel disease |
IL | interleukin |
LOD | limits of detection |
LOQ | limits of quantitation |
LPS | bacterial lipopolysaccharide |
MEF | methanol-water (75:25, v/v) extract of fresh fruits |
PB2 | procyanidin B2 |
PBMCs | peripheral blood mononuclear cells |
QU | quercetin |
ROS | reactive oxygen species |
TAC | total content of anthocyanins |
TFL | total content of flavonoids |
TNF-α | tumour necrosis factor α |
TPA | total content of phenolic acids |
TPC | total phenolic content (Folin–Ciocalteu assay) |
TPH | total phenolic content (sum of individual phenolics by HPLC) |
TTC | total content of tannin-type proanthocyanidins |
WRF | water residue of MEF after fractionated extraction |
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MEF | DEFF | EAFF | BFF | WRF | |
---|---|---|---|---|---|
Total contents: | |||||
TPC (GAE) | 87.57 ± 3.54 b | 126.49 ± 1.41 a | 123.63 ± 3.68 a | 68.23 ± 0.12 c | 64.59 ± 0.61 c |
TPH | 28.56 ± 0.58 d | 81.83 ± 0.80 b | 104.02 ± 1.92 a | 43.17 ± 1.14 c | 6.07 ± 0.14 e |
TPA | 19.67 ± 0.33 d | 35.15 ± 1.12 b | 91.26 ± 2.16 a | 29.62 ± 1.10 c | 4.79 ± 0.08 e |
TAC | 4.64 ± 0.11 b | n.d. | n.d. | 9.17 ± 0.33 a | 0.96 ± 0.06 c |
TFL | 4.25 ± 0.21 c | 41.11 ± 0.41 a | 12.21 ± 0.32 b | 4.38 ± 0.17 c | 0.32 ± 0.02 d |
TTC (PB2) | 44.53 ± 1.93 a | n.d. | n.d. | 8.02 ± 0.23 c | 28.36 ± 0.41 b |
Individual compounds: | |||||
Avicularin (50) | 1.32 ± 0.14 c | 2.52 ± 0.02 b | 2.98 ± 0.14 a | 0.41 ± 0.06 d | n.d. |
Guaiaverin (48) | n.d. | 1.05 ± 0.05 a | 0.61 ± 0.06 b | n.d. | n.d. |
Hyperoside (42) | 0.11 ± 0.005 c | 0.94 ± 0.07 b | 1.51 ± 0.03 a | n.d. | n.d. |
Isoquercitrin (45) | 0.09 ± 0.004 c | 0.21 ± 0.01 b | 0.97 ± 0.07 a | 0.12 ± 0.01 c | n.d. |
Reinutrin (47) | n.d. | 0.38 ± 0.02 a | 0.24 ± 0.02 b | n.d. | n.d. |
Rutin (44) | 1.60 ± 0.02 c | 0.29 ± 0.04 d | 2.25 ± 0.27 b | 2.93 ± 0.11 a | 0.22 ± 0.03 d |
Quercitrin (52) | 0.15 ± 0.01 c,d | 1.66 ± 0.04 b | 1.91 ± 0.11 a | 0.13 ± 0.01 d | n.d. |
Quercetin (62, QU) | n.d. | 27.45 ± 0.09 a | 1.25 ± 0.13 b | n.d. | n.d. |
Cyanidin 3-O-glucoside (19, CYG) | 1.96 ± 0.12 b | n.d. | n.d. | 4.46 ± 0.12 a | 0.55 ± 0.06 c |
Cyanidin 3-O-rutinoside (23) | 1.39 ± 0.07 b | n.d. | n.d. | 2.28 ± 0.10 a | 0.41 ± 0.05 c |
Peonidin-3-O-glucoside (26) | 0.99 ± 0.04 b | n.d. | n.d. | 1.68 ± 0.07 a | n.d. |
Protocatechuic acid (2) | n.d. | 5.22 ± 0.11 a | 0.63 ± 0.02 b | n.d. | n.d. |
p-Hydroxybenzoic acid (8) | n.d. | 0.51 ± 0.06 a | 0.47 ± 0.01 a | n.d. | n.d. |
Vanillic acid (13) | n.d. | 3.37 ± 0.19 a | n.d. | n.d. | n.d. |
p-Coumaric acid (30) | n.d. | 0.52 ± 0.03 a | n.d. | n.d. | n.d. |
Neochlorogenic acid (7) | 15.56 ± 0.30 c | 10.93 ± 0.84 d | 49.62 ± 1.51 a | 24.47 ± 1.07 b | 3.40 ± 0.15 e |
Chlorogenic acid (14, CHA) | 0.94 ± 0.02 b | 0.62 ± 0.02 c | 4.75 ± 0.03 a | 0.39 ± 0.01 d | n.d. |
Cryptochlorogenic acid (18) | 1.57 ± 0.07 d | 4.85 ± 0.10 b | 28.74 ± 0.37 a | 3.63 ± 0.12 c | n.d. |
Vanillin (24) | n.d. | 5.57 ± 0.38 a | 0.55 ± 0.04 b | n.d. | n.d. |
Solvent B (Acetonitrile) | Solvent C (Tetrahydrofuran) | ||
---|---|---|---|
Time (min) | Concentration (%, v/v) | Time (min) | Concentration (%, v/v) |
0–1 | 3.0 (isocratic elution) | 0–8 | 2.5 (isocratic elution) |
1–25 | 3.0 → 20.5 (linear gradient) | 8–9 | 2.5 → 6.5 (linear gradient) |
25–45 | 20.5 → 50.0 (linear gradient) | 9–19 | 6.5 (isocratic elution) |
45–50 | 50.0 (isocratic elution) | 19–24 | 6.5 → 9.0 (linear gradient) |
50–52 | 50.0 → 3.0 (return to initial condition) | 24–50 | 9.0 (isocratic elution) |
52–60 | 3.0 (equilibration) | 50–52 | 9.0 → 2.5 (return to initial condition) |
52–60 | 2.5 (equilibration) |
Analyte | λ (nm) | Regression (Linear Model) | r | Linear Range (μg/mL) | F-Test | LOD (μg/mL) | |
---|---|---|---|---|---|---|---|
Equation | n | ||||||
Protocatechuic acid | 260 | y = 15884.45x | 6 | 0.9998 | 0.64–63.7 | 19979.99 | 0.398 |
p-Hydroxybenzoic acid | 260 | y = 25596.21x | 6 | 0.9997 | 0.63–62.8 | 13094.52 | 0.365 |
Vanillic acid | 260 | y = 16884.29x | 6 | 0.9997 | 0.56–55.5 | 15385.86 | 0.373 |
Vanillin | 280 | y = 18673.22x | 6 | 0.9999 | 0.67–66.8 | 32063.03 | 0.389 |
Cyanidin 3-O-β-d-glucopyranoside | 520 | y = 17908.10x | 6 | 0.9990 | 0.52–52.3 | 4091.50 | 0.339 |
Cyanidin 3-O-rutinoside | 520 | y = 10946.63x | 6 | 0.9997 | 0.51–51.0 | 12466.71 | 0.325 |
Peonidin 3-O-β-d-glucopyranoside | 520 | y = 12604.30x | 6 | 0.9956 | 0.48–48.5 | 906.70 | 0.340 |
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Magiera, A.; Czerwińska, M.E.; Owczarek, A.; Marchelak, A.; Granica, S.; Olszewska, M.A. Polyphenol-Enriched Extracts of Prunus spinosa Fruits: Anti-Inflammatory and Antioxidant Effects in Human Immune Cells Ex Vivo in Relation to Phytochemical Profile. Molecules 2022, 27, 1691. https://doi.org/10.3390/molecules27051691
Magiera A, Czerwińska ME, Owczarek A, Marchelak A, Granica S, Olszewska MA. Polyphenol-Enriched Extracts of Prunus spinosa Fruits: Anti-Inflammatory and Antioxidant Effects in Human Immune Cells Ex Vivo in Relation to Phytochemical Profile. Molecules. 2022; 27(5):1691. https://doi.org/10.3390/molecules27051691
Chicago/Turabian StyleMagiera, Anna, Monika Ewa Czerwińska, Aleksandra Owczarek, Anna Marchelak, Sebastian Granica, and Monika Anna Olszewska. 2022. "Polyphenol-Enriched Extracts of Prunus spinosa Fruits: Anti-Inflammatory and Antioxidant Effects in Human Immune Cells Ex Vivo in Relation to Phytochemical Profile" Molecules 27, no. 5: 1691. https://doi.org/10.3390/molecules27051691