(e) Pharmacological activity: neuroprotective and neuromodulating function

Experiment design (Experiment model; Assay; Incubation period; Test subject)	Extract	Plant part	Plant Source	Extract dose; Route of administration; Positive control	Result	Reference
in vitro; OGD–reoxygenation and hypoxic neuronal death, Cell viability assay; Mouse primary cortical neurons, cerebral astrocytes, cerebral endothelial cells	80% Ethanol; Soaking	Leaf	Singapore	5 μg/mL	Extract suppressed post-hypoxic HDACs activation and reduce OGD-caused neuronal death	[75]
in vitro; OGD–reoxygenation; 24 h Human SH-SY5Y neuroblastoma cells	80% Ethanol; Soaking	Leaf	Singapore	100 μg/mL	Extract modulated cPLA2 expression induction in SH-SY5Y cells by HDAC inhibitors, MS-275, MC-1568, TSA and inhibited HAT activity.	[74]
in vitro; OGD–reoxygenation; 12 h and 24 h; Mouse primary cortical neurons	80% Ethanol; Soaking	Leaf	Singapore	1.6, 6.25 μg/mL	Extract inhibited levels of cPLA2 mRNA expression in primary cortical neurons subjected to 0.5 h OGD injury	[74]
in vitro; OGD; CCK-8; up to 12 h; neutron pre-treated with extract 1 h before OGD; Mouse primary cortical neurons from E15.5 Balb/c mouse embryos	80% Ethanol; Soaking	Leaf	Malaysia	6.25 μg/mL	Extract treated neurons showed significant increment in cell viability	[76]
in vitro; OGD–reoxygenation; CCK-8, MMP, apoptosis analysis, transient transfection and chromatin reporter assay; 0.5 h OGD+ 4–24 h Reoxygenation; Neurons treated with extract (1 h before, on the onset or after OGD-reoxygenation); Mouse primary cortical neurons from E15.5 Balb/c mouse embryos	80% Ethanol; Soaking	Leaf	Malaysia	0.075–20 μg/mL	Neurons treated with extract before, at the onset or after OGD showed increment in dose dependent manner but protective effect of extract was lesser when applied after OGD.   Neurons treated with 6.25 μg/mL extract at the onset of OGD has reduced MMP breakdown, apoptotic death, and pro-apoptotic (caspase-3, PARP-1) and has higher anti-apoptotic (14-3-3ε, p-Bad, Bcl-2) markers. Neurons treated with 2.5–10 μg/mL extract at the onset of OGD suggested a dose dependent increase effect in PPAR-γ mRNA level.	[76]
in vitro; Anti-spasmotic effect on acetycholine induced rat's bladder tissue contraction; Isolated tissues of rat's bladder	Ethanol	Leaf, stem	Malaysia	Mebeverine hydrochloride and detrusitol	Stem at 120 mg/mL: 79.77%  Leaf at 100 mg/mL: 26.62%  Detrusitol at 200 μg/mL: 89.86%, Mebeverine at 10 mg/mL: 80.74% of contraction inhibition	[93]
in vivo; Ellman assay-Acetylcholinesterase activity in brain, liver, kidney, heart; Pretreatment for 14 days (daily treatment (once)); Balb/C mice (male, 25 ± 5 g)	Methanol, Air dry, Maceration	Leaf	Seremban Malaysia	250, 500, 1000 mg/kg bw	All doses of extracts caused the higher acetylcholinesterase activity in liver, kidney, and heart compared to control group while they did not caused any difference in brain	[77]
in vivo; MCA occlusion stroke model; Reperfusion infarct volume, behaviour assessment, MRI; extract treated 1 h before, immediate or 3–24 h after 30 min MCA occlusion or 1 d after 15 min MCA acclusion; Long- Evans rats (male, 7-8 week-old)	80% Ethanol; Soaking	Leaf	Malaysia	10–60 pg (i.c.v., after 30 min MCA occlusion); 24 mg/kg body wt (i.p, 1 h before or 3–24 h after 30 min or 24 h after 15 min MCA occlusion)	I.C.V. treatment of 40 pg extract reduced apoptotic neuronal death, showed a maximal decrease of cerebral infarct volume at 1 day reperfusion. PPAR-γ, C/EBPβ mRNA levels, anti- apoptotic 14-3-3ε, p-Bad, and Bcl-2 levels were upregulated. Pro-apoptotic cleaved PARP-1 and cleaved caspase 3 protein levels were downregulated. Extract improved the functional outcomes based on the behavior observation at day 7 after ischemia (elevated body swing test and Bederson's postural reflex task) and at day 14 after ischemia (better ladder rung walking test). Extract via i.p treatment also significantly reduced infarct volume except 24 h after 30 min MCA occlusion. Behavior test conducted also showed better functional outcomes and smaller infarct volume via MRI.	[76]