A simple solid injection device for the analyses of Bulbophyllum (Orchidaceae) volatiles Ubiratan F. da Silva a, b , Eduardo L. Borba c , JoaÄo Semir c , Anita J. Marsaioli a, * a Instituto de QuõÂmica, UNICAMP, CP 6154, Campinas 13081-970, SP, Brazil b Departamento de QuõÂmica, UFSM, Brazil c Instituto de Biologia, UNICAMP, Campinas 13081-970, SP, Brazil Received 14 May 1998 Abstract The volatile components of three orchid species (Bulbophyllum weddellii, B. ipanemense and B. involutum) were detected using GC/MS. Due to the minute dimension and amount of ¯owers, special traps and extraction methods were applied and the analyses were ®nally successful using a special solid sample injector which was constructed in our own laboratory. The chemical composition comparison revealed the high similarity between the volatiles present in B. weddellii and B. involutum, which is coherent with the pollinator sharing. On the other hand, the volatiles of B. ipanemense, a species pollinated by other pollinator species, are dierent from the two previous species. # 1998 Published by Elsevier Science Ltd. All rights reserved. Keywords: Bulbophyllum weddellii; B. ipanemense; B. involutum; Orchidaceae; GC/MS; Volatiles; Solid injector 1. Introduction The orchid family is widespread across the world and over 25,000 species are known to exist (Dressler, 1993). The variety of shapes, colors and scents present in Orchidaceae is seldom found in other plant families. They are pollinated by moths, carrion-¯ies, bees, but- ter¯ies, birds, etc. and there is a de®nite relationship between the ¯oral scent and the pollinators (van der Pijl & Dodson, 1966). Three Brazilian orchid species, Bulbophyllum weddellii, B. involutum and B. ipane- mense, native of Minas Gerais state, Brazil, were extensively studied from the ¯oral biology and phe- nology point of view, revealing that the three species are pollinated by females of Pholeomyia (Diptera: Milichiidae) with a high degree of speci®city (Borba, 1997). The above observations have attracted our attention as they might indicate that the orchid scents play a sig- ni®cant role in this orchid-pollinator relationship (Dressler, 1968; Janzen, 1971; Faegri & van der Pijl, 1979). We have thus focused on the chemical analysis of the volatile components of these three Bulbophyllum species. 2. Results and discussions The ®rst part of our investigations concerned the optimization of the ¯ower scent trapping and we have used B. weddellii. Based on previous experiments (Kaiser), the headspace technique was our ®rst choice. A glass vessel enclosing the orchid, a battery-operated pump and a trap containing Porapak Q was our major device. The trapped volatiles were desorbed from the trap with bidistilled solvent (dichloromethane and methanol 1.5 mL) and directly injected in the GC/MS, after reducing the volume of the solvent to 0.1 mL. After several attempts we came to the conclusion that this methodology was not ecient for these analyses due to loss of some rather volatile components and de- composition of others. High pressure Soxhlet extrac- tion with CO 2 (J and W High Pressure Soxhlet Extractor) and microhydrodistillation (Ferracini et al., 1995) were also performed, but in the three method- ologies we had to face analogous problems: the insig- ni®cant amount of extracted material, minute dimensions of the ¯owers and decomposition of the ¯oral scent within a few hours. We came ®nally to the conclusion that our analyses would never be correct unless the use of solvents was avoided. We have thus looked for a direct loading technique (Senanayake et al., 1976) which should be applicable Phytochemistry 50 (1999) 31±34 0031-9422/98/$ - see front matter # 1998 Published by Elsevier Science Ltd. All rights reserved. PII: S0031-9422(98)00459-2 PERGAMON * Author to whom correspondence should be sent.