In vitro propagation, carotenoid, fatty acid and tocopherol content of Ajuga multiflora Bunge

Abstract

The effect of plant growth regulators on shoot proliferation from shoot tip explants of Ajuga multiflora was studied. The highest number of shoots (17.1) was observed when shoot tip explants were cultured on Murashige and Skoog (MS) medium fortified with 8.0 µM 6-Benzyladenine (BA) and 2.7 µM α-naphthaleneacetic acid (NAA). The mean number of shoots per explant was increased 1.6-fold in liquid medium as compared with semi-solid medium. Maximum rooting (100 %) with an average of 7.2 roots per shoot was obtained on MS basal medium. Rooted plantlets were successfully acclimatised in the greenhouse with 100 % survival rate. Composition of carotenoids, fatty acids and tocopherols was also studied from leaves of greenhouse-grown plants and in vitro-regenerated shoots of A. multiflora. The greatest amounts of carotenoids, fatty acids and tocopherols were obtained from leaves of in vitro-regenerated shoots cultured on MS basal medium, followed by leaves of greenhouse-grown plants and leaves of in vitro-regenerated shoots cultured on MS basal medium with 2.0 µM BA or thidiazuron. The most abundant carotenoid in A. multiflora leaves was all-E-lutein (89.4-382.6 μg g^-1 FW) followed by all-E-β-carotene (32.0-156.7 μg g^-1 FW), 9'-Z-neoxanthin (14.2-63.4 μg g^-1 FW), all-E-violaxanthin (13.0-45.9 μg g^-1 FW), all-E-zeaxanthin (1.3-2.5 μg g^-1 FW) and all-E-β-cryptoxanthin (0.3-0.9 μg g^-1 FW). α-Tocopherol was the predominant tocopherol in A. multiflora leaves. Linolenic acid (49.03-52.59 %) was detected in higher amounts in A. multiflora leaf samples followed by linoleic acid (18.95-21.39 %) and palmitic acid (15.79-18.66 %).

Keywords: 6-Benzyladenine; Ajuga multiflora; Bioactive compounds; Shoot proliferation; Shoot tip; Thidiazuron.

Fig. 1

In vitro propagation of A. multiflora; a, b multiple shoots’ (green and purple shoots) induction on MS medium fortified with 8.0 µM BA; c shoot proliferation on MS medium fortified with 8.0 µM BA and 2.7 µM NAA; d rooting on MS basal medium; e, f acclimatised plants

Fig. 2

Shoot proliferation in a balloon-type bubble bioreactor containing MS liquid medium fortified with 8.0 µM BA and 2.7 µM NAA. a After 7 days of culture; b after 28 days of culture

Fig. 3

Effect of liquid and semi-solid culture media on shoot proliferation of A. multiflora after 45 days of culture

Fig. 4

HPLC chromatograms (UV, 450 nm) of carotenoids in leaf tissues of A. multiflora. 1 all-E-violaxanthin (RT: 6.6); 2 9′-Z-neoxanthin (RT: 7.5); 3 all-E-lutein (RT: 12.5); 4 all-E-zeaxanthin (RT: 14.5); 5 all-E-β-cryptoxanthin (RT: 22.7); 6 α-carotene (RT: 29.3); and 7 All-E-β-carotene (RT: 32.5); Chl chlorophyll

Fig. 5

HPLC chromatograms (UV, 295 nm) of tocopherols in leaf tissues of Ajuga multiflora. 1 δ-tocopherol (RT: 7.3); 2 γ-tocopherol (RT: 8.3); 3 α-tocopherol (RT: 9.4)