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European Journal of Histochemistry 2011; volume 55:e18 Luminescence and fluorescence families as Asteraceae, Apiaceae, Lauraceae, Pinaceae and others.9,10 They are usually pro- Correspondence: Federico Boschi, Department of of essential oils. Fluorescence duced by steam distillation, although extrac- Morphological-Biomedical Sciences, Section of imaging in vivo of wild tion methods such as mechanical expression, Anatomy and Histology, University of Verona, chamomile oil solvent extraction or supercritical fluid extrac- strada Le Grazie 8, 37134 Verona, Italy. tion are also used.11,12 Essential oils are usual- Tel. +39.045.8027155 - Fax: +39.045.8027163. E-mail: [email protected] F. Boschi, M. Fontanella, L. Calderan, ly lipophilic and not soluble in water, but according to the composition and the produc- A. Sbarbati Key words: optical imaging, essential oil, trans- tion method they can be dissolved in alcohols; dermal administration, fluorescence, phospho- Department of Morphological-Biomedical 13 moreover they are often volatile compounds. rescence, luminescence, in vivo. Sciences, Section of Anatomy and Terpenoids, the main components of the oils, Histology, University of Verona, Italy are synthesized from isoprene units and follow Acknowledgements: the authors wish to thank Dr. the chemical formula (C5H9-11)n. Mono- Maria Paola Cecchini for the helpful discussions (n=2) and sesquiterpene s (n=3)14 are the and the Fondazione Monte dei Paschi di Siena for most common compounds, followed by diter- the financial support. Abstract penes (n=4) and aromatic compounds (phe- Received for publication: 18 September 2010. nols, benzenoids, flavonoids).15-18 The yield and Accepted for publication: 6 May 2011. Essential oils are currently of great impor- the composition of products can be greatly tance to pharmaceutical companies, cosmetics influenced by plant organs (leaves, flowers, This work is licensed under a Creative Commons producers and manufacturers of veterinary stems), as well by the ecosystem in which the Attribution NonCommercial 3.0 License (CC BY- NC 3.0). products. They are found in perfumes, creams, plant is cultivated.19,20 Certainly, different bath products, and household cleaning sub- species of the same gender could show differ- ©Copyright F. Boschi et al., 2011 stances, and are used for flavouring food and ent compositions (e.g. lavandula officinalis, Licensee PAGEPress, Italy drinks. It is well known that some of them act lavandula lanata and lavandula stoechas), European Journal of Histochemistry 2011; 55:e18 on the respiratory apparatus. The increasing although the main components of terpenoid doi:10.4081/ejh.2011.e18 interest in optical imaging techniques and the and phenolic fractions are similar.15,16,21,22 development of related technologies have Concerning interactions with living sys- was to investigate the luminescent properties made possible the investigation of the optical tems, it is known that essential oils affect the of essential oils, and to see if they could be properties of several compounds. Luminescent respiratory system;1 it was recently shown, measured in living organisms. We were inter- properties of essential oils have not been using Magnetic Resonance Imaging, that they ested in transdermal administration, which extensively investigated. We evaluated the can act on the physical properties of the mucus mimics the diffusion of some drugs and of luminescent and fluorescent emissions of sev- in rat airways.23,24 In recent years optical imag- chemicals used in cosmetics. We investigated eral essential oils, in order to detect them in ing techniques have been developed thanks to the luminescence of several essential oils living organisms by exploiting their optical the technological evolution of light detectors, (after exposure to normal ambient illumina- properties. Some fluorescent emission data which allow the study of the optical properties tion and to sunshine) and their fluorescent were high enough to be detected in dermal of different compounds. One of the fundamen- emission. We also detected the fluorescent treatments. Consequently, we demonstrated tal mechanisms of interaction between light emission of wild chamomile oil from the skin how the fluorescent signal can be monitored and biological objects is luminescence, which of treated mice. This study was a proof of prin- for at least three hours on the skin of living is subdivided into fluorescence, corresponding ciple for research about the application of mice treated with wild chamomile oil. The to an allowed optical transition with a short essential oils in vivo, and aimed to develop a results encourage development of this tech- (nanosecond) lifetime, as well as phosphores- new approach to the study of the effects of nique to investigate the properties of drugs cence, corresponding to a forbidden transition organic compounds in living organisms. and cosmetics containing essential oils. with longer decay times.25 Fluorescence is a process in which the emission of light is induced by illumination of the subject.26 Phosphorescence may be an appropriate term Materials and Methods Introduction to be used in the context of luminescence from organic compounds involving triplet-to-singlet Fluorescence and luminescence images ® Essential oils are composed of different mix- transitions.27 According with the common use were acquired using VivoVision System IVIS tures of organic compounds extracted from we refer here with luminescence to the radia- 200 (Xenogen Corporation, Alameda, CA, plants, having many biological activities.1 Most tive processes except for fluorescence. It is USA), made up of a camera sensor back essential oils (55%), are used in the food known that luminescence in plants is a thinned, back illuminated grade CCD 1 (2.7 industry for the production of aroma extracts, response to stress and shocks;28,29 fluorescence ¥2.7 cm, 2048¥2048 pixels, pixel dimension 13.5 µm, cooled at 90°C) and a 150W Quartz while 20% are used as fragrances in perfumery may correlate with senescence30 and stress.31 halogen 3250° Kelvin lamp. Images were and cosmetics, or in pharmaceutical or natural However, the optical properties of essential acquired using Living Image 2.6 software products (5%); a consistent percentage of oils oils have been subject to very little investiga- (Xenogen Corporation). are separated into their component parts tion. Although luminescence and fluorescence (15%) for various uses.2 Recently there has cannot furnish direct information about the been a certain interest in their antibacterial chemical composition of essential oils, we sup- Compounds tested and antifungal properties or antioxidant activ- posed that they could be used to track the per- Wild Chamomile, Lavender (named here ities; some compounds have been isolated and sistence and diffusion of drugs or cosmetics Lavender 1), Marjoram, Melissa, Mentha, their pharmaceutical behaviour studied.3-8 containing essential oils on the skin of treated Oregano, Pine, Rosemary, and Tea Tree (pur- Essential oils are derived from as different living organisms. The aim of the present study chased from Dolisos, Pomezia, RM, Italy), [European Journal of Histochemistry 2011; 55:e18] [page 97] Original paper Lavender (named here Lavender 2) and Lemon experiment the animals had been fed an alfal- essential oils showed the maximum emission (purchased from Just, Grezzana, VR, Italy) fa-free diet to reduce tissue autofluores- with GFP excitation and GFP emission filters, were tested. For each compound 300 μL were cence.33 or GFP excitation and DsRed emission filters. placed in a non-fluorescent 96 multiwell plate The research was conducted in accordance Excitation light with a longer wavelength than and imaged using the optical instrument. with the regulations of the Italian Ministry of GFP produced lower fluorescent signal intensi- Health and to the European Communities ty. Using the GFP-GFP setup, the efficiency of Luminescence imaging Council (86/609/EEC) directives. all the compounds was in the range 0-6.5 10–5 p/s and the corresponding flux was in the Luminescent images of the compounds range 0-1.4 109 p/s. The greatest signal intensi- were acquired with the subsequent parame- ty with the GFP-GFP setup was found with ters: quartz halogen lamp off, field of view = Results Lavender 2 and Wild Chamomile. 12.8x12.8 cm, f/stop = 1, binning factor = 16, When narrow band emission filters were exposure time = 5 or 10 min; no excitation or applied, all the essential oils presented a emission filters were used. Luminescence imaging decrease of the signal from 560 nm to 660 nm. Essential oils were handled in a daylit labo- The data of the luminescent oil emissions ratory with blinds and placed in the dark room are shown in Figure 1. In the pre sunshine Fluorescence imaging in vivo of the instrument for 30 min before the first acquisition we found that the flux emitted luminescence acquisition (pre); then, the from the essential oils was in the range 1-5 103 The in vivo treatment with wild chamomile samples were exposed to sunshine for 5 min, p/s, with Melissa and Lavender 2 proving to be increased the emission coming from the skin after which three luminescent images (post1, the most luminescent compounds. In the first of the animals compared to the pre treatment post2 and post3) with exposure times of 10 image after sunshine exposure, almost all the image. Particularly, in Figure 3, it is possible to min were acquired consecutively. essential oils showed an increase in emission, compare the emission from the treated ani- followed by a continuous decrease in the sub- mals 3 h after the end of the treatment and the Fluorescence imaging sequent acquisitions. Lavender 2 and wild emission before treatment, with respect to the chamomile emitted the greatest signal intensi- control animal. Fluorescent images of the compounds were ty in all post images. The measurements of fluorescent emission acquired with the subsequent parameters: from a region of interest (ROI) drawn on quartz halogen lamp on, field of view = images and corresponding to the furless back 12.8x12.8 cm, f/stop = 2, binning factor = 4, Fluorescence imaging of the mice are presented in Figure 4.
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  • Growing and Knowing Lavender

    Growing and Knowing Lavender

    GROWING & KNOWING LAVENDER WRITTEN BY JOHN MASON CONTENTS CHAPTER 1 DIVERSITY AND CLASSIFICATION 8 The lavender family 9 Lavandula angustifolia syn L.vera, L.spica, L.officinalis (English lavender) 11 Lavandula stoechas ssp. stoechas (Spanish/Italian lavender) 11 Lavandula stoechas ssp stoechas ‘alba’ 11 Lavandula stoechas ssp. pendunculata - syn. L. stoechas ssp. canariensis 11 Lavandula viridis: 1m green/cream flowers, narrow green leaves. 11 Lavandula dentata var. candicans 11 Lavandula canariensis 12 Lavandula pubescens 12 Lavandula abrotanoides 12 Lavandula bipinnata 12 Lavandula gibsonii 12 Lavandula subnuda 12 Lavandula macra 12 Lavandula aristibracteata 12 Tim Upson’s Classification 13 Lavandula angustifolia 13 Lavandula lanata 13 Lavandula latifolia 13 Lavandula dentata var. dentata 13 Lavandula dentata var. candicans 13 Lavandula stoechas subsp. stoechas 13 Lavandula stoechas subsp. pedunculata 13 Lavandula stoechas subsp. sampaiana 13 Lavandula stoechas subsp. lusitanica 13 Lavandula stoechas subsp. luisieri 13 Lavandula stoechas subsp. atlantica 13 Lavandula stoechas subsp. maderensis 13 Lavandula stoechas subsp. cariensis 13 Lavandula viridis 13 Lavandula multifida 13 Lavandula canariensis 13 Lavandula pinnata 13 Lavandula buchii 13 Lavandula minutolii 13 Lavandula maroccana 13 Lavandula tenuisecta 13 Lavandula mairei 13 Lavandula antineae 13 Lavandula coronopifolia (syn. L. stricta) 13 Lavandula pubescens 13 Lavandula citriodora 13 Lavandula subnuda 13 Lavandula macra 13 Lavandula dhofarensis 13 Lavandula setifera 13 Lavandula