Quality, Safety and Biological Studies on Campylanthus glaber Aerial Parts

Abstract

In Cabo Verde, several endemic species are used in traditional medicine. However, no scientific studies have been conducted on the quality, efficacy, and safety of most of these plants. This study focused on establishing the botanical and chemical identification parameters required for a quality monograph of Campylanthus glaber Benth. aerial parts, a medicinal plant of Cabo Verde traditionally used to treat fever and muscular pain. In addition, in vitro antioxidant and antihyperglycemic activity, cytotoxicity, and genotoxicity were assessed for this medicinal plant. Optical microscopy, LC/UV-DAD-ESI/MS, and colorimetric assays were used for botanical, chemical, and biological studies, respectively. Cytotoxicity was assessed by the MTT assay with HepG2 cells, and genotoxicity by the Ames test. Microscopically, the xeromorphic leaf of C. glaber presents a thick cuticle (13.6-25.5 µm), thick-walled epidermal cells, anomocytic-type stomata, glandular trichomes (stalk length = 49.4-120.8 µm), and idioblasts containing calcium oxalate microcrystals. The chemical screening of aqueous and hydroethanolic extracts of this medicinal plant revealed the presence of organic acids, iridoids, phenylethanoids, and flavonoids as the main classes of marker compounds, with malic acid, citric acid, and verbascoside being the main marker compounds identified. Both extracts showed similar LC/UV-DAD/ESI-MS qualitative profiles and DPPH radical scavenger activity (IC₅₀ = 130.9 ± 1.4; 134.3 ± 3.1 µg/mL). The hydroethanolic extract inhibited both α-amylase and α-glucosidase enzymes in a dose-dependent manner. Both extracts showed no cytotoxicity (up to 1000 µg/mL) by the MTT assay and no genotoxic potential with or without metabolic activation up to 5 mg /plate. The results obtained are an important contribution to the monographic quality assessment of C. glaber aerial parts and suggest that this medicinal plant may be safe and potentially used as an herbal drug raw material for pharmaceutical purposes.

Keywords: Campylanthus glaber; antihyperglycemic; antioxidant; herbal drug; herbal medicine; quality control; safety; verbascoside.

Figure 1

Macroscopic characteristics of Campylanthus glaber aerial parts; (a) C. glaber dried aerial parts; (b) C. glaber flowering branch; (c) Bunch of dried aerial parts of C. glaber (alecrim-bravo) sold in Santiago Island market (Praia market); scale bars: 1 cm. Photographies by Katelene Lima (a,b) and Maria Cristina Duarte (c).

Figure 2

Macroscopic characteristics of Campylanthus glaber, aerial parts: (a) general aspect of the stem branches with the presence of trichomes (arrow) in the younger branch nodes; (b) cross-section section of the stem branch with details of the cork region and vascular cylinder; (c) leaf abaxial view; (d) leaf adaxial view with detail of groove (arrow) formed by revolute margin covered by trichomes. Abbreviations: cc: central cylinder; ck: cork; vc: vascular cylinder; tr: trichomes.

Figure 3

Microscopic characteristics of the Campylanthus glaber stem cross-section. (a) General aspect of the epidermis (arrow (ep)), cork, parenchyma, sclerenchyma (arrow (sc)), phloem, xylem, and central cylinder (arow(cc)) (from outside to inside). Details of (b) cork with four layers of quadrangular-shape cells; (c) parenchyma and groups of sclereids; (d) groups of sclereids forming a ring around the phloem; (e) xylem vessels organized in 2–8 cells; (f) central cylinder large thick-walled cells; (g) trichomes located in the epidermis. Abbreviations: cc: central cylinder; ck: cork; ep: epidermis; p: parenchyma; ph: phloem; sc: sclereids; xl: xylem. Scale bars: (a) = 100 µm; (b–g) = 50 µm.

Figure 4

Microscopic characteristics of Campylanthus glaber leaf. (a) Cross-section of the midrib showing the cylindrical mesophyll with detail of stomata (arrow (st)) and groove (arrow (gr)); (b) detail of trichome (arrow) located in the groove formed by the curved margins and palisade parenchyma; (c) vascular bundles (arrows) embedded in the aquifer parenchyma; (d) xylem vessels (arrow) and phloem; (e) idioblasts (arrow) with calcium oxalate microcrystals; (f) epidermal cells (arrows (ep)) and anomocytic-type stomata (arrows (st)); (g) trichomes located on the leaf margins with detail of trichomes with swollen base (arrow). Abbreviations: ap: aquifer parenchyma; ec: epidermal cells; gr: groove; id: idioblast; pp: palisade parenchyma; ph: phloem; st: stomata; vb: vascular bundle; xv: xylem vessel. Scale bars: (a) = 200 µm; (b,c) = 100 µm; (d) = 20 µm; (e–g) = 50 µm.

Figure 5

Microscopic characteristics of C. glaber aerial part powder. (a) Fragmented leaf epidermal cells with stomata and idioblasts with microcrystals; (b) protective trichome; (c) fragment of pitted vessels; (d) fragment of the stem central cylinder.

Figure 6

LC/UV-DAD profile of CgAE and CgE with UV spectra and retention times of the main marker classes of compounds identified: organic acids (peaks (a,b)), iridoid glucosides (peaks (c–e,g)), caffeoyl phenylethanoids glycosides ((f,j,l–p)) flavonoids (peaks (h,k)), and an unknown compound (i). Abbreviations: CgAE: Campylanthus glaber aqueous extract; CgEE: Campylanthus glaber ethanolic extract.

Figure 7

Viability of HepG2 after incubation (48 h) with CgAE and CgEE extracts assessed by MTT reduction assay. Data arpresented as percentage of solvent control (dashed line) and mean ± standard deviation; n = 2–3. * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001.