Lasianthera africana P. Beauv. (Icacinaceae) is a traditional Nigerian medicinal plant used for treatment of ulcers, diarrhea, parasitic infections and diabetes. This study was aimed at characterizing the bioactive principles extractable from the flavonoid-rich fraction of L. africana leaves (LAFRF), and to evaluate its effects on renal and cardiac functions. Isolation, and purification of the LAFRF was achieved using standard methods. The in vitro antioxidant activity was evaluated on DPPH* and ferric reducing antioxidant potential (FRAP). The total flavonoids (281.05 AE 7.44 mg QE/g), were identified, structurally characterized and quantified using high resolution ultra-performance liquid chromatography, in tandem with quadrupole-time-of-flight electrospray ionization mass spectrometer (UPLC-PDA-QTOF-ESI-MS/MS). Fifty Wistar rats of both sexes (110-130 g), were distributed into 10 groups (n ¼ 5). Groups 1 and 2 served as the normal and CCl 4 controls respectively. Groups 3A-6B constituted the preventive and curative studies. The effects of the LAFRF at 3, 10, and 30 mg/kg body weight on urea and creatinine concentrations, lactate dehydrogenase (LDH), and creatine kinase (CK) activities of CCl 4 -intoxicated rats were assessed. The LAFRF displayed remarkable in vitro antioxidant property by scavenging the DPPH*, with an IC 50 of 5.40 AE 0.00 μg/ml which is more potent than the scavenging activity of the ascorbic acid (IC 50 of 7.18 AE 0.00 μg/ml), and also effectively reduced Fe 3þ to Fe 2þ when compared to gallic acid. The UPLC-PDA-QTOF-ESI-MS/MS fingerprint of the LAFRF indicated presence of quercetin (758983.6 mg/kg), rutin (17540.4 mg/kg), luteolin (126524.3 mg/kg), isorhamnetin (197949.0 mg/kg), and other non-phenolic compounds. The LAFRF significantly (p < 0.05) improved renal function, and normalized cardiac enzyme activities in vivo. The ability of the LAFRF to scavenge the DPPH and Fe 3þ radicals, improve renal and cardiac functions following CCl 4 intoxication shows its potential in the development of alternative therapy for combating oxidative stress-related complications.
Various aromatic compounds that are structurally analogous to lignin were tested as possible/preferred substrates for purified laccase from newly isolated white rote fungi,
Trametes polyzona
WRF03. The pH optima were tested using different substrates and kinetic studies were conducted at these pH optima. The pH optima in the presence of ABTS, α-naphthol, o-dianisidine, and catechol were 4.5 but 5.0 and 5.5 in the presence of guaiacol and pyrogallol, respectively. The initial velocities obtained from the kinetic study were analyzed using Graph Pad Prism 7 and Lineweaver-Burk plot to obtain kinetic constants (
k
m
and
Vmax
) which were used to calculate substrate specificity. Amongst all the substrates tested, ABTS had the highest specificity-constant (181.51 M
−1
s
−1
), and therefore, the most preferred substrate was followed by α-naphthol,
o
-dianisidine, guaiacol, pyrogallol, and catechol. Resorcinol, orcinol, and veratryl alcohol did not display any considerable chemical shift in the presence of
Trametes polyzona
WRF03 laccase. Also, oxidation of phenolic substrates appeared to be dependent on the nature of the substituent groups and their relative position on the aromatic nucleus. Since most of these substrates are structural analogs of lignin and many recalcitrant environmental pollutants, the enzyme may find application in delignification, treatment of wastewater containing dyes, and polycyclic aromatic hydrocarbons (PAHs).
Context
Bryophyllum pinnatum
(Lam.) Oken (Crassulaceae) is used traditionally to treat many ailments.
Objectives
This study characterizes the constituents of
B. pinnatum
flavonoid-rich fraction (BPFRF) and investigates their antioxidant and anticholinesterase activity using
in vitro
and
in silico
approaches.
Materials and methods
Methanol extract of
B. pinnatum
leaves was partitioned to yield the ethyl acetate fraction. BPFRF was isolated from the ethyl acetate fraction and purified. The constituent flavonoids were structurally characterized using UPLC-PDA-MS
2
. Antioxidant activity (DPPH), Fe
2+
-induced lipid peroxidation (LP) and anticholinesterase activity (Ellman’s method) of the BPFRF and standards (ascorbic acid and rivastigmine) across a concentration range of 3.125–100
μg/mL were evaluated
in vitro
for 4 months. Molecular docking was performed to give insight into the binding potentials of BPFRF constituents against acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE).
Results
UPLC-PDA-MS
2
analysis of BPFRF identified carlinoside, quercetin (most dominant), luteolin, isorhamnetin, luteolin-7-glucoside. Carlinoside was first reported in this plant. BPFRF significantly inhibited DPPH radical (IC
50
= 7.382 ± 0.79 µg/mL) and LP (IC
50
= 7.182 ± 0.60 µg/mL) better than quercetin and ascorbic acid. Also, BPFRF exhibited potent inhibition against AChE and BuChE with IC
50
values of 22.283 ± 0.27 µg/mL and 33.437 ± 1.46 µg/mL, respectively compared to quercetin and rivastigmine. Docking studies revealed that luteolin-7-glucoside, carlinoside and quercetin interact effectively with crucial amino acid residues of AChE and BuChE through hydrogen bonds.
Discussion and conclusions
BPFRF possesses an excellent natural source of cholinesterase inhibitor and antioxidant. The material could be further explored for the potential treatment of oxidative damage and cholinergic dysfunction in Alzheimer’s disease.
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