Centella asiatica - Phytochemistry and mechanisms of neuroprotection and cognitive enhancement

Abstract

This review describes in detail the phytochemistry and neurological effects of the medicinal herb Centella asiatica (L.) Urban. C. asiatica is a small perennial plant that grows in moist, tropical and sub-tropical regions throughout the world. Phytochemicals identified from C. asiatica to date include isoprenoids (sesquiterpenes, plant sterols, pentacyclic triterpenoids and saponins) and phenylpropanoid derivatives (eugenol derivatives, caffeoylquinic acids, and flavonoids). Contemporary methods for fingerprinting and characterization of compounds in C. asiatica extracts include liquid chromatography and/or ion mobility spectrometry in conjunction with high-resolution mass spectrometry. Multiple studies in rodent models, and a limited number of human studies support C. asiatica's traditional reputation as a cognitive enhancer, as well as its anxiolytic and anticonvulsant effects. Neuroprotective effects of C.asiatica are seen in several in vitro models, for example against beta amyloid toxicity, and appear to be associated with increased mitochondrial activity, improved antioxidant status, and/or inhibition of the pro-inflammatory enzyme, phospholipase A2. Neurotropic effects of C. asiatica include increased dendritic arborization and synaptogenesis, and may be due to modulations of signal transduction pathways such as ERK1/2 and Akt. Many of these neurotropic and neuroprotective properties of C.asiatica have been associated with the triterpene compounds asiatic acid, asiaticoside and madecassoside. More recently, caffeoylquinic acids are emerging as a second important group of active compounds in C. asiatica, with the potential of enhancing the Nrf2-antioxidant response pathway. The absorption, distribution, metabolism and excretion of the triterpenes, caffeoylquinic acids and flavonoids found in C. asiatica have been studied in humans and animal models, and the compounds or their metabolites found in the brain. This review highlights the remarkable potential for C. asiatica extracts and derivatives to be used in the treatment of neurological conditions, and considers the further research needed to actualize this possibility.

Figure 1:

Centella asiatica plant, showing spade-like leaves, internode stolons and small flowers.

Figure 2.

Cytoscape network for compounds found in C. asiatica. The clustering relationship is based on structural similarities among 57 compounds using the Tanimoto algorithm. Seven main clusters are present: saponins, pentacyclic triterpenoids, sterols, sesquiterpenes, eugenol derivatives, caffeoylquinic acids (subclusters of mono- or di- caffeoylquinic acids) and flavonoids. A representative compound of each cluster is shown. The compound numbers displayed correspond to the Pubchem CIDs (Table 2), and the compound numbers 1–16 refer to the compounds given in Figure 4. ▲, indicates a node with a Tanimoto score of < 0.68. A Tanimoto score > 0.68 is statistically significant at the 95% confidence interval (Kim et al. 2012b).

Figure 3.

Structural sub-types of centelloids. a- ursane family and b- oleanane family. R₁-R₃= OH or H, R₄=H or oligosaccharide. Adapted from Azerad (Azerad 2016).

Figure 4.

Structures of compounds detected in aqueous extracts of C. asiatica.

Figure 5.

UPLC-MS analysis of an aqueous extract of C. asiatica. 10 μl Injection (5 mg ml⁻¹ of dry mass resuspended in 70% aqueous methanol). a Ion intensity map (of an aqueous extract of C. asiatica. In this 2D visualization, retention time is given on the x-axis, the m/z values are shown on the y-axis, and colors represents signal intensity. b Extracted ion chromatograms for 16 commonly found compounds. [M-H]⁻ (1,3,4: m/z 353.09; 2,5: m/z 289.07; 6,7,9–11: m/z 515.12; 8: m/z 609.15; 12: m/z 301.03; 13: m/z 973.50; 14: m/z 957.51; 15: m/z 503.34; 16: m/z 487.34). Chromatographic separation was conducted using a Shimadzu Nexera UPLC system equipped with Inertsil Phenyl-3 column (150 × 4.6 mm, 5 μm). Mobile phase A was water with 0.1 % formic acid, and mobile phase B was methanol with 0.1% formic acid. The gradient started with 5% B and was held for 1 min, followed by a 10 min linear gradient from 5 % to 30 %. The gradient was then stepped to 100% B at 23 min and held for 12 min and finally, stepped back to 5% B to equilibrate the column. The flow rate was 0.4 mL min⁻¹, and the column temperature was maintained at 45 °C. An AB Sciex Triple TOF 5600 mass spectrometer equipped with a TurboSpray electrospray ionization sourceoperated in the negative ionization mode was used. The instrument was operated in the information-dependent acquisition (IDA) mode using a collision energy of 40 V. Compound identity is based on accurate mass, isotopic pattern, retention time, MS/MS spectra and standard addition using authentic compounds. Structures of compounds 1 to 16 are given in Figure 4.

Figure 6.

UPLC-MS chromatogram of an ethanolic extract of C. asiatica. 10 μl Injection (5 mg ml⁻¹ of dry mass resuspended in methanol 70%). Shown is an overlay of extracted ion chromatograms for commonly found compounds. [M-H]⁻ (1,3,4: m/z 353.09; 6,7,9–11: m/z 515.12; 8: m/z 609.15; 12: m/z 301.03; 13: m/z 973.50; 14: m/z 957.51; 15: m/z 503.34; 16: m/z 487.34). Experimental conditions as described in the legend of Figure 5. Structures of compounds 1 to 16 are given in Figure 4.

Figure 7.

Mass spectra of isomeric mono-caffeoylquinic acids recorded in negative ionization mode showing differences in the intensity of MS fragment ions. Standards were injected by infusion (10 μl min⁻¹ in methanol:water 70:30 V/V, 1 mg L⁻¹). A Synapt G2 HDMS (Waters Corp., MA, USA) was used for ion detection. The cone voltage was 20 V and the transfer energy was set at 25 eV. a 3-O-caffeoylquinic acid, b 4-O-caffeoylquinic acid, c 5-O-caffeoylquinic acid, d Intensity comparison for major MS/MS product ions of CQAs. Data are in accord with Xie et al., 2011.

Figure 8.

ESI q-IMS-MS/MS analysis of an aqueous extract of C. asiatica (5 mg ml⁻¹ of dry mass resuspended in methanol 70%, 10 μl injection). a, 2D map visualization of drift time versus retention time, b, drift time distribution of ions, c and d, fragmentation spectrum and proposed transfer dissociation of madecassoside ([M-H]⁻ m/z 973.5) and asiaticoside ([M-H]⁻ m/z 957.5), respectively. Method: Acquity UPLC chromatographic conditions were as described in Figure 5. The LC system was coupled to a Synapt G2 HDMS (Waters Corp., MA, USA) used for detection in negative ion electrospray mode. Nitrogen was used as carrier gas for ion mobility experiments. Data acquisition range was m/z 50–1200. The cone voltage was 20 V. The T-wave ion mobility cell was operated at 800 m s⁻¹ and the wave amplitude was set at 35 V. Helium and nitrogen IMS carrier were both set at 80 ml min⁻¹.

Figure 9.

Morris water maze probe data of healthy aged male and female mice treated with a water extract of Centella asiatica (CAW, 2 mg/mL dissolved in the drinking water) for 2 weeks prior to, and during testing. CAW increased memory retention in both male and female healthy aged mice. Extract treated animals spent significantly more time in the target quadrant than control, untreated mice. (Adapted from Gray et al. J. Ethnopharmacology 2016)

Figure 10.

Morris water maze data of aged female Tg 2576 transgenic or wild type (WT) mice treated with a water extract of Centella asiatica (GKW, 2 mg/mL dissolved in the drinking water) for 2 weeks prior to, and during testing. C.asiatica treatment normalized memory deficits in the Tg2576 mouse model of Aβ accumulation, reducing the time and distance traveled by Tg2576 mice to find the hidden platform compared to untreated Tg2576 animals. (adapted from Soumyanath et al. 2012).