Development of White Cabbage, Coffee, and Red Onion Extracts as Natural Phosphodiesterase-4B (PDE4B) Inhibitors for Cognitive Dysfunction: In Vitro and In Silico Studies

Abstract

Human cognition fundamentally depends on memory. Alzheimer's disease exhibits a strong correlation with a decline in this factor. Phosphodiesterase-4 B (PDE4B) plays a crucial role in neurodegenerative disorders, and its inhibition is one of the promising approaches for memory enhancement. This study aimed to identify secondary metabolites in white cabbage, coffee, and red onion extracts and identify their molecular interaction with PDE4B by in silico and in vitro experiments. Crushed white cabbage and red onion were macerated separately with ethanol to yield respective extracts, and ground coffee was boiled with water to produce aqueous extract. Thin layer chromatography (TLC)-densitometry was used to examine the phytochemicals present in white cabbage, coffee, and red onion extracts. Molecular docking studies were performed to know the interaction of test compounds with PDE4B. TLC-densitometry analysis showed that chlorogenic acid and quercetin were detected as major compounds in coffee and red onion extracts, respectively. In silico studies revealed that alpha-tocopherol (binding free energy (∆G_bind) = -38.00 kcal/mol) has the strongest interaction with PDE4B whereas chlorogenic acid (∆G_bind = -21.50 kcal/mol) and quercetin (∆G_bind = -17.25 kcal/mol) exhibited moderate interaction. In vitro assay showed that the combination extracts (cabbage, coffee, and red onion) had a stronger activity (half-maximal inhibitory concentration (IC₅₀) = 0.12 ± 0.03 µM) than combination standards (sinigrin, chlorogenic acid, and quercetin) (IC₅₀ = 0.17 ± 0.03 µM) and rolipram (IC₅₀ = 0.15 ± 0.008 µM). Thus, the combination extracts are a promising cognitive enhancer by blocking PDE4B activity.

Figure 1

TLC profile of coffee, onion, chlorogenic acid, and quercetin along with their spectrums (b) and (d). TLC plates indicate (a(i)) and (c(iii)) (at 254 nm); (a(ii)) and (c(iv)) (at 366 nm); CE = coffee extract; OE = onion extract; Cl = chlorogenic acid; Qu = quercetin; AUC = area under the curve; Rf = retention factor.

Figure 2

The 3D caption of native and known ligands with native ligand, and the 2D caption of native ligand and known ligands with amino acid residues. (a) = redocked native ligand 3D interaction with amino acid residues; (b) = native ligand 2D interaction with amino acid residues; (c) = docked known ligand (ZINC000043194322) 3D interaction with amino acid residues; (d) = known ligand (ZINC000043194322) 2D interaction with amino acid residues. Trp = tryptophan; Phe = phenylalanine; Asp = aspartic acid; Gln = glutamine; Met = methionine; Ser = serine; Ile = isoleucine; His = histidine; H = hydrogen; N = nitrogen; F = fluorine; O = oxygen; H₂O = water.

Figure 3

The 3D caption of rolipram and quercetin with amino acid residues, and the 2D caption of rolipram and quercetin with amino acid residues. (a) = rolipram 3D interaction with amino acid residues; (b) = rolipram 2D interaction with amino acid residues; (c) = quercetin 3D interaction with amino acid residues; (d) = quercetin 2D interaction with amino acid residues. Phe = phenylalanine; Asp = aspartic acid; Thr = threonine; Ile = isoleucine; His = histidine; N = nitrogen; H = hydrogen; O = oxygen; OH = hydroxyl group.

Figure 4

The 3D caption of chlorogenic acid and alpha-tocopherol with native ligand, and the 2D caption of chlorogenic acid and alpha-tocopherol with amino acid residues. (a) = chlorogenic acid 3D interaction with amino acid residues; (b) = chlorogenic acid 2D interaction with amino acid residues; (c) = alpha-tocopherol 3D interaction with amino acid residues; (d) = alpha-tocopherol (PubChem CID 1742129) 2D interaction with amino acid residues. Phe = phenylalanine; Asn = asparagine; Glu = glutamic acid; Asp = aspartic acid; Gln = glutamine; Met = methionine; Thr = threonine; Ile = isoleucine; Val = valine; His = histidine; H = hydrogen; O = oxygen; OH = hydroxyl group; H₂O = water.

Figure 5

GI tract and brain permeation prediction of the 41 top-ranked secondary metabolites in white cabbage, red onion, and coffee by brain or the intestinal estimated permeation predictive model (BOILED-Egg) method. GI = gastrointestinal, BBB = blood-brain barrier, HIA = human intestinal absorption, PGP+ = P-glycoprotein positive, PGP- = P-glycoprotein negative, TPSA = topological polar surface area, secondary metabolites: 1 = sinigrin, 5 = gluconapin, 8 = glucobrassicanapin, 9 = quercetin, 10 = catechin, 11 = kaempferol, 13 = cyanidin, 14 = chlorogenic Acid, 15 = p-coumaric acid, 16 = caffeic acid, 17 = gallic acid, 18 = ascorbic acid, 21 = caffeine, 22 = trigonelline, 24 = myricetin, 25 = taxifolin, 26 = laricitrin, 27 = isorhamnetin, 30 = tectorigenin, 31 = ferulic acid, 32 = protocatechuic acid, 33 = onionin (A) 34 = cycloalliin, 35 = isoalliin, 36 = methiin, 37 = propiin, 38 = allicin, 39 = S-methyl-L-cysteine, 40 = S-ethylcysteine, 41 = S-propyl-L-cysteine, 42 = S-allylcysteine, 43 = S-propylmercaptocysteine, 44 = dipropyl trisulfide, 45 = diallyl disulfide, 46 = diisopropyl disulfide, 47 = methyl propenyl disulfide, 48 = 6-methyl-4,5-dithia-1-heptene, 49 = cyanidin 3-glucoside, 52 = malvidin 3-glucoside, 53 = peonidin-3-glucoside, and 54 = alliin.

Figure 6

Nonlinear regression curves between the activity (%) of nine tested samples against PDE4B1 and their concentrations (Log (µM)). (a) = sinigrin (IC50 = 0.24 ± 0.01 µM); (b) = white cabbage extract (IC50 = 0.27 ± 0.009 µM); (c) = quercetin (IC50 = 0.25 ± 0.01 µM); (d) = red onion extract (IC50 = 0.22 ± 0.02 µM); (e) = chlorogenic acid (IC50 = 0.20 ± 0.03 µM); (f)=coffee extract (IC50 = 0.18 ± 0.03 µM); (g) = combination (sinigrin, quercetin, and chlorogenic acid) standard (IC50 = 0.17 ± 0.03 µM); (h) = combination (white cabbage, red onion, and coffee) extract (IC50 = 0.12 ± 0.03 µM); (i) = rolipram (a PDE4B inhibitor) (IC50 = 0.15 ± 0.008 µM). PDE4B1 = phosphodiestrase-4B1; No Cpd = no compound.

Figure 7

Mechanism of action of cabbage, coffee, and red onion extracts. GPCR = G-protein coupled receptor, GDP = guanosine diphosphate, GTP = guanosine triphosphate, AC = adenylyl cyclase, ATP = adenosine triphosphate, cAMP = cyclic adenosine monophosphate, 5′-AMP = 5-adenosine monophosphate, PKA = protein kinase (A) BDNF = brain-derived neurotrophic factor, CREB = cAMP-response element binding, pCREB = phosphorylated cAMP-response element binding.