Alnus genus (Betulaceae)

Almost all the species belonging in Alnus genus have been traditionally used in Ayurveda, Unani, and Chinese folk medicine; traditional Korean medicine utilised them for haemorrhage, burn injuries, fever, diarrhoea and alcoholism; contemporary indigenous healers also use primarily the bark of Alnus species for the preparation of various therapeutical teas (Turner and Hedba 1990).

Different parts of the plants like stems, bark, seeds, leaves, roots, fruits, tree cones, buds and blossoms are known to possess therapeutical activity and they are characterised by a remarkable number of compounds such as triterpenoids, saponins, flavonoids, diarylheptanoids, phenols, steroids and tannins (Sati et al. 2011).

These molecules exhibited antioxidant (Dinic et al. 2014), anti-inflammatory (Lai et al.

2012), antiviral (Tung et al. 2010), cytotoxic (León-Gonzales et al. 2014, Novaković et al. 2014, Choi et al. 2008), inhibitory activity against nuclear factor kappa activation, nitric oxide and tumour necrosis factor-production, human umbilical vein endothelial cells, farnesyl protein transferase, cell-mediate low density lipoprotein oxidation, and HIV -1 - induced cytopathic effect in MT-4 cells (Sati et al. 2011).

2.3.6.1. Antioxidant activity

Evaluation of the antioxidant activity of two diarylheptanoids, platyphylloside 5(S)-1,7-di(4-hydroxyphenyl)-3-heptanone-5-O-β-D-glucopyranoside and its analogue, 1,7-di(4-hydroxyphenyl)-5-O-β-D-[6-(E-p-coumaroylglucopyranosyl)]heptane-3-one, both isolated from the bark of Alnus glutinosa L. has been carried out by Dinic et al.

(Dinic et al. 2014). The published results indicated that neutralization of reactive oxygen species is an important mechanism of diarylheptanoid action, although these compounds exert a considerable anticancer effect. Therefore, the authors concluded that

30

these compounds may serve as protectors of normal cells during chemotherapy without significantly diminishing the effect of the applied chemotherapeutics.

Antioxidant and antimicrobial activities of methanolic extracts from the leaves and barks of three Alnus species (Alnus glutinosa L., Alnus incana L. and Alnus viridis Chaix) was evaluated by Dahija et al. Antioxidant activity of the extracts was determined using the DPPH radical scavenging method. All of the extracts showed antioxidant activity higher than that of thymol, which was used as a positive control (Dahija et al. 2014).

The ethanolic extract prepared from the leaves of Alnus formosana Burk showed notable antioxidant activity in in vitro assays using DPPH, hydroxyl and superoxide free radicals (Lee et al. 2006). This study did not include the identification of the antioxidant constituents, but other papers reported the presence of oregonin (2) as the main compound in Alnus formosana bark extracts (Lee et al. 2005), which previously has been proved to be a potent antioxidant (Keserű and Nógrádi 1995).

Oregonin (2) and hirsutenone (1) (see structures in section 6.3.1.) isolated from Alnus japonica Thunb. inhibited LDL peroxydation in human cells in vitro with much higher activity than the positive control probucol. This effect suggests therapeutical potential in arteriosclerosis and related diseases (Lee et al. 2005).

2.3.6.2. Anticancer activity

The protective effects towards doxorubicin damaging activity of two diarylheptanoids isolated from the bark of Alnus glutinosa: platyphylloside (i22), 5(S)-1,7-di(4-hydroxyphenyl)-3-heptanone-5-O-β-D-glucopyranoside and its newly discovered analog 5(S)-1,7-di(4-hydroxyphenyl)-5-O-β-D -[6-(E-p-coumaroylglucopyranosyl)]heptane-3-one were studied by Dinic et al. HaCaT cells were employed, which are non-cancerous human keratinocytes commonly used for skin regenerative studies. Diarylheptanoids significantly antagonised the effects of doxorubicin by lowering the sensitivity of HaCaT cells to this drug. (S)-1,7-di(4-hydroxyphenyl)-5-O-β-D -[6-(E-p-coumaroylglucopyranosyl)]heptane-3-one prevented doxorubicin-induced cell death by activating autophagy. Both the diarylheptanoids protected HaCaT cells against doxorubicin-induced DNA damage. They significantly promoted migration and affected F-actin distribution. These results indicate that chemo-protective effects of

31

diarylheptanoids may occur at multiple subcellular levels. Therefore, these diarylheptanoids could be considered as protective agents for non-cancerous dividing cells during chemotherapy (Dinic et al. 2015).

A study of secondary metabolites from the bark of Alnus glutinosa led to the isolation of fourteen diarylheptanoids: oregonin (2), platyphylloside, rubranoside A, rubranoside, B hirsutanolol, hirsutenone (1), hirsutanonol-5-O-β-D-glucopyranoside, platyphyllonol-5-O-β-D-xylopyranoside, aceroside VII, alnuside A, alnuside B, 1,7-bis-(3,4-dihydoxyphenyl)-5-hydroxy-heptane-3-O-β-D-xylopyranoside, (5S)-1-(4-hydroxyphenyl)-7-(3,4-dihydroxyphenyl)- 5-O-β-D-glucopyranosyl-heptan-3-one, and (5S)-1,7-bis-(3,4-dihydroxyphenyl)-5-O-β-D

-[6-(3,4-dimethoxycinnamoyl-glucopyranosyl)]-heptan-3-one. All isolated compounds were analysed for in vitro protective effect on chromosome aberrations in peripheral human lymphocytes in 1-2 µg/ml concentrations, using Amifostine as positive control. The majority of them, exerted pronounced effect in decreasing DNA damage. The authors observed that those compounds that possessed pronounced protective activity had a 3-keto group in the heptanoid chain and also catechol moieties. The protective effect strongly correlated with the antioxidant activity of the compounds (Novakovic et al. 2014).

2.3.6.3. Anti-inflammatory activity

Diarylheptanoids from Alnus hirsuta Spach. exhibited inhibitory activity on the production of nitric oxide, on reactive oxygen species and on the expression of various pro-inflammatory molecules (Weicheng et al. 2011).

Previous studies of Alnus formosana Burk’s leaves revealed a rich amount of diarylheptanoid glycosides, including oregonin (2) (Lee et al. 2006) that demonstrated to be a significant COX-2 inhibitor (Lee et al. 2000); active against LPS-induced NO production (Lai et al. 2012) and also considered an antioxidant agent.

2.3.6.4. Immunosuppressive activity

The effect of hirsutenone (1) isolated form Alnus japonica on the processes involved in the induction and maintenance of atopic dermatitis was investigated by Joo et al.

Hirsutenone (1) inhibited the proliferation of T- and B-cells in a dose dependent

32

manner. The authors assumed that the main mechanism of action involves the inhibition of the dephosphorilation of NFATc2, a transcription factor responsible for the emergence of atopic dermatitis. The immunosuppressive effect of hirsutenone (1) was comparable to that of cyclosporine, a well-known calcineurin inhibitor. These results suggested that the effect of hirsutenone (1) on the cytokines produced by the T-cells originates from calcineurin inhibition (Joo et al. 2010).

2.3.6.5. Other effects

An extensive research was carried out on the anti-adipogenic effect of A. hirsuta f.

sibirica: following the isolation of a new diarylheptanoid, the study indicated its inhibitory effect on the induction of peroxysome proliferator activated receptor γ protein expression and on the adipocyte differentiation in 3T3-L1 cells (Lee et al. 2013).

Anti-influenza activity was reported about platyphyllon isolated from the bark of Alnus japonica that showed antiviral effects against KBNP-0028 (H2N2) (Tung et al. 2010).