Biotechnology Advances 31 (2013) 166–174 Contents lists available at SciVerse ScienceDirect Biotechnology Advances journal homepage: www.elsevier.com/locate/biotechadv Research review paper In vitro culture of lavenders (Lavandula spp.) and the production of secondary metabolites Sandra Gonçalves, Anabela Romano ⁎ Institute for Biotechnology and Bioengineering, Centre of Genomics and Biotechnology (IBB/CGB), Faculty of Sciences and Technology, University of Algarve, Campus de Gambelas, Ed. 8, 8005-139 Faro, Portugal article info abstract Article history: Lavenders (Lavandula spp., Lamiaceae) are aromatic ornamental plants that are used widely in the food, perfume Received 31 May 2012 and pharmaceutical industries. The large-scale production of lavenders requires efficient in vitro propagation tech- Received in revised form 21 September 2012 niques to avoid the overexploitation of natural populations and to allow the application of biotechnology-based Accepted 22 September 2012 approaches for plant improvement and the production of valuable secondary metabolites. In this review we dis- Available online 27 September 2012 cuss micropropagation methods that have been developed in several lavender species, mainly based on meristem proliferation and organogenesis. Specific requirements during stages of micropropagation (establishment, shoot Keywords: Cell suspensions multiplication, root induction and acclimatization) and requisites for plant regeneration trough organogenesis, Meristem proliferation as an important step for the implementation of plant improvement programs, were revised. We also discuss differ- Organogenesis ent methods for the in vitro production of valuable secondary metabolites, focusing on the prospects for highly Secondary metabolites scalable cultures to meet the market demand for lavender-derived products. Shoot multiplication © 2012 Elsevier Inc. All rights reserved. Rooting Contents 1. Introduction ............................................................. 166 2. Importance and applications ...................................................... 167 3. In vitro propagation .......................................................... 167 3.1. Micropropagation of Lavandula spp. by meristem proliferation ................................... 168 3.1.1. Establishment of in vitro cultures ............................................ 168 3.1.2. Shoot multiplication .................................................. 168 3.1.3. Root induction ..................................................... 168 3.1.4. Acclimatization .................................................... 169 3.2. Plant regeneration via organogenesis .............................................. 169 4. Production of secondary metabolites .................................................. 170 5. Metabolic pathways of essential oil biosynthesis and metabolic engineering studies . ........................... 170 6. Concluding remarks .......................................................... 171 Acknowledgment .............................................................. 172 References ................................................................. 172 1. Introduction The lavenders (Lavandula spp.) are flowering plants of the mint Abbreviations: ABA, Abscisic acid; Ads, Adenine hemisulfate; BA, 6-Benzylaminopurine family (Lamiaceae) that are endemic to the Mediterranean region, the ′ or 6-Benzyladenine; CPPU, N-(Chloro-4-pyridyl)-N -phenylurea; IAA, Indole-3-acetic acid; Arabian Peninsula, the Canary Islands and India. There are 39 species, IBA, Indole-3-butyric acid; MS, Murashige and Skoog (1962) medium; NAA, α- Naphthaleneacetic acid; PGR, Plant growth regulator; TDZ, Thidiazuron. numerous hybrids and nearly 400 registered cultivars (Upson and ⁎ Corresponding author. Andrews, 2004), many of which have aromatic and medicinal proper- E-mail address: [email protected] (A. Romano). ties that are highly valued in the fragrance, pharmaceutical, food and 0734-9750/$ – see front matter © 2012 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.biotechadv.2012.09.006 S. Gonçalves, A. Romano / Biotechnology Advances 31 (2013) 166–174 167 flavor industries. Lavenders are also popular ornamental and decorative Table 1 plants (Touati et al., 2011). Main compounds of the essential oils from some of the most important Lavandula spp. Effective protocols for the propagation of Lavandula spp. are required Speciesa Main compounds Reference to produce large quantities of plants for the ornamentals market or for Lavandula angustifolia Linalol and linalool acetate D'Auria et al. (2005) the extraction of valuable metabolites while avoiding the exploitation Mill. of wild populations. Lavenders may be propagated sexually or asexually, Lavandula latifolia Med. Linalol, 1,8-cineole and camphor Munõz-Bertomeu et although reproduction through seeds is often slow and the plants vary al. (2007a) b significantly in terms of growth rate and essential oil composition. Vege- Lavandula×intermedia Linalol, linalool acetate, camphor Desautels et al. and 1,8-cineole (2009) tative propagation is necessary to produce genetically homogeneous in- Lavandula pedunculata Camphor and 1,8-cineole Zuzarte et al. (2010) dividuals, but multiplication via cuttings is slow and labor-intensive, and (Miller) Cav. root induction is often inefficient. Additional factors such as climate, Lavandula pinnata L. fil. α- and β-phellandrene Figueiredo et al. water availability and susceptibility to diseases may also influence the (1995) Lavandula stoechas L. Fenchon, camphor, myrtenyl Giray et al. (2008) content and composition of essential oils (Chawla, 2009; Pierik, 1987). acetate and 1,8-cineole The limitations discussed above can be overcome using in vitro Lavandula viridis L'Hér 1,8-Cineole and camphor Nogueira and propagation methods under controlled environmental conditions, fa- Romano (2002) cilitating the rapid multiplication of superior clones and the extrac- a Species names as used by the authors. tion of valuable metabolites throughout the year without seasonal b A sterile cross between L. latifolia and L. angustifolia. constraints (Zuzarte et al., 2010). Furthermore, genetic engineering is only possible in conjunction with in vitro culture techniques (Kothari et al., 2010), which could provide lavender producers with The medicinal properties of lavender essential oils were recently opportunities to improve the quality and quantity of essential oils reviewed by Woronuk et al. (2011). These oils can be absorbed into the and thus the economic potential of Lavandula spp. (Dronne et al., human body via three routes: through the respiratory system, by direct 1999a). The last review to consider the propagation of lavenders by contact with the skin and by oral ingestion (Perry and Perry, 2006). Es- tissue culture was published more than 20 years ago, prior to the de- sential oils are widely used in aromatherapy and massage, and many velopment of in vitro culture techniques with a greater emphasis on benefits are claimed for these practices (Cavanagh and Wilkinson, micropropagation (Segura and Calvo, 1991). We therefore summarize 2002). Lavender oils are thought to encourage sleep (Field et al., 2008; more recent work focusing on the application of in vitro propagation Hallschmid et al., 2004) and are widely used to reduce anxiety techniques to Lavandula spp. and the use of in vitro plant cells and tis- (Bradley et al., 2007; Dobetsberger and Buchbauer, 2011; Kritsidima et sues for the production of valuable secondary metabolites. al., 2009). Lavender essential oils have been evaluated for the treatment of dementia (Smallwood et al., 2001)andseveralLavandula spp. are thought to produce oils with antimicrobial, anticholinesterase and anti- 2. Importance and applications oxidant activities (Costa et al., 2012; Hanamanthagouda et al., 2010). In addition to volatiles, lavenders produce further active metabolites such Lavenders are widely used as ornamental and melliferous plants as phytosterols, phenolic acids and flavonoids, which act as antioxidants (Upson and Andrews, 2004) and can also help in the reforestation of (Costa et al., 2011; Georgiev et al., 2009; Nitzsche et al., 2004; Spiridon et fire-damaged areas (González-Coloma et al., 2011). However, the eco- al., 2011). Lavender extracts have been also reported with properties nomic value of Lavandula spp. predominantly relates to the properties suitable for the management of central nervous system disorders of their essential oils, which are strictly regulated by international ISO (Alnamer et al., 2012; Costa et al., 2011). standards and are used both cosmetically and therapeutically for centu- Lavender extracts are also used in the food industry because of their ries. Lavender essential oils are extensively used in the manufacturing health-promoting and nutraceutical effects. Hsu et al. (2007) found of soaps, perfumes, food flavors and other products, as pleasant fra- that aqueous extracts of L. angustifolia Vera and Lavandula stoechas grances or as antimicrobial agents (Cavanagh and Wilkinson, 2002; contained a potent tyrosinase inhibitor and were suitable as food Woronuk et al., 2011). Evidence for the therapeutic use of lavenders bleaching agents, whereas Kovatcheva-Apostolova et al. (2008) found can be traced back to the ancient Romans and Greeks, but a recent in- that the addition of Lavandula vera extract to minced chicken reduced crease in the popularity of alternative medicines has renewed interest lipid oxidation and the loss of α-tocopherol