Volatile Compounds of Viola odorata Absolutes: Identification of Odorant Active Markers to Distinguish Plants Originating from France and Egypt

Absolutes isolated from Viola odorata leaves, valuable materials for the flavor and fragrance industry, were studied. Violets are mainly cultivated in France and Egypt and extracted locally. The absolutes of the two origins showed different olfactory profiles both in top and heart notes, as evidenced by sensory analysis. The aims of this study were i) to characterize the volatile compounds, ii) to determine the odorant‐active ones, and iii) to identify some markers of the plant origin. Two complementary analytical methods were used for these purposes, i.e., headspace solid‐phase microextraction (HS‐SPME) using different fiber coatings followed by GC/MS analysis and gas chromatography – olfactometry/mass spectrometry (GC‐O/MS) applied to violet leaf extracts. From a total of 70 identified compounds, 61 have never been reported so far for this species, 17 compounds were characterized by both techniques (with seven among them known from the literature), 23 compounds were solely identified by HS‐SPME GC/MS (among them only two being already mentioned as components of violet absolutes in the literature), and, finally, 30 compounds were only identified by GC‐O/MS. According to the HS‐SPME GC/MS analyses, ethyl hexanoate and (2E,6Z)‐nona‐2,6‐dienol were specific volatile compounds of the sample with French origin, while (E,E)‐hepta‐2,4‐dienal, hexanoic acid, limonene, tridecane, and eugenol were specific of the samples with Egyptian origin. Additional compounds that were not detected by HS‐SPME GC/MS analysis were revealed by GC‐O analyses, some of them being markers of origin. Pent‐1‐en‐3‐ol, 3‐methylbut‐2‐enal, 2‐methoxy‐3‐(1‐methylethyl)pyrazine, 4‐ethylbenzaldehyde, β‐phenethyl formate, and 2‐methoxy‐3‐(2‐methylpropyl)pyrazine revealed to be odorant markers of the French sample, whereas cis‐rose oxide, trans‐rose oxide, and 3,5,5‐trimethylcyclohex‐2‐enone were odorant markers of the Egyptian samples.     

Electroantennogram responses of a parasitic wasp, Microplitis croceipes, to host-related volatile and anthropogenic compounds

The parasitic wasp Microplitis croceipes (Cresson) (Hymenoptera: Braconidae) showed its own characteristic electroantennogram (EAG) response profiles to 13 host‐related (cis‐3‐hexenol, α‐pinene (R)‐(+)‐limonene (S)‐(–)‐limonene, trans‐β‐ocimene (±)‐linalool, (–)‐trans‐caryophyllene, α‐humulene, nerolidol, trans‐nerolidol, cis‐nerolidol, methyl jasmonate and indole) and four anthropogenic (2‐diisopropylaminoethanol, 2,2′‐thiodiethanol, 2‐methyl‐5‐nitroaniline and cyclohexanone) volatile compounds. These profiles were similar between males and females except for 2‐diisopropylaminoethanol, which elicited significantly larger EAG responses in males. Among the compounds tested, cis‐3‐hexenol, linalool and cyclohexanone elicited the largest EAG responses. EAG responses were not influenced by the age of wasps between 1 and 13 days after emergence. EAG responses were dose‐dependent, and highly EAG‐active compounds elicited significant EAG responses with less than 10 μg of the compounds at source. Quantification of compounds released from an odour cartridge indicates that release rate is highly dependent on the chemical nature of stimuli, showing up to 10 000‐fold differences in the amount released between different compounds when the same amount was loaded in the odour cartridge. Wasps having undergone a behavioural training regime to be attracted to either cyclohexanone or methyl jasmonate did not show any differences in EAG responses from those of untrained wasps.     

Volatiles released from bean plants in response to agromyzid flies

Liriomyza sativae Blanchard and Liriomyza huidobrensis (Blanchard) (Diptera: Agromyzidae) are two invasive flies in China that have caused economical damage on vegetables and ornamental plants. In this article, we report the profiles of emitted volatiles from healthy, mechanically damaged, and leafminer-damaged bean, Phaseolus vulgaris L., plants. Among 25 emitted volatiles identified, (E)-2-hexen-1-al, (3E)-4,8-dimethyl-1,3,7-nonatriene (DMNT), (Z)-3-hexenyl acetate, (Z)-3-hexen-1-ol, (syn)- and (anti)-2-methylpropanal oxime, (syn)-2-methylbutanal oxime, linalool, and (E,E)-α-farnesene were consistently released from damaged bean plants. Combined amounts of these nine compounds made up more than 70% of the total volatiles emitted from each treatment. No qualitative differences in volatile emission were found between bean plants damaged by the two fly species; however, amounts of several major compounds induced by L. huidobrensis damage were significantly higher than those from plants damaged by L. sativae. The mechanically damaged plants released a higher proportion of green leaf volatiles than plants in the other treatments, whereas leafminer-damaged plants produced more terpenoids and oximes. Furthermore, the volatile profiles emitted from plants, damaged by adult leafminers, by second instar larvae, and even the plants with empty mines left by leafminer larvae (the pupal stage) were significantly different. The identification of volatile oximes released from damaged plants was confirmed and is discussed in a behavioral and biological control context.Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorized users.     

The volatile organic compounds of introduced and native dung and carrion and their role in dung beetle foraging behaviour

1. The decomposition of biological material produces a plethora of volatile organic compounds (VOCs), which are implicated in the foraging behaviour of coprophagous and necrophagous insects. Dung beetles (Coleoptera: Scarabaeidae: Scarabaeinae) have an acute olfactory system used to locate food resources. Accordingly, identification of food resource VOCs potentially used in food location is integral to understanding dung beetle foraging ecology. 2. In this study, volatile emissions from dung and carrion of native and introduced animals in New Zealand were analysed using solid‐phase microextraction (SPME) and gas chromatography‐mass spectrometry (GC‐MS). Volatile profiles were compared via principal component analyses (PCAs) and cluster solutions based on attractiveness using canonical discriminant analysis (CDA). 3. A total of 115 compounds were detected from 21 food types. Statistical analyses showed that dung and carrion volatile profiles clustered according to attractiveness to the dung beetle Saphobius edwardsi, and that different dung types formed distinct clusters and grouped separately from carrion. 4. This study suggests that volatile profiles emitted by food resources used by dung beetles are complex, producing distinct odours, which potentially mediate foraging decisions.     

Chemical composition and antimicrobial activity of volatiles from Degenia velebitica, a European stenoendemic plant of the Brassicaceae family

Free and glucosidic bound leaf volatiles of Degenia velebitica were isolated and fractionated simultaneously into H₂O‐soluble, H₂O‐insoluble, and highly volatile compounds by hydrodistillation–adsorption (HDA) and analyzed by GC/MS. Among the 24 constituents identified, the main compounds obtained by the HDA method were S‐ and/or N‐atom containing compounds, i.e., 6‐(methylsulfanyl)hexanenitrile ( 10 ; 26.78%), dimethyl trisulfide ( 6 ; 26.35%), 3,4,5‐trimethylpyrazole ( 17 ; 13.33%), hex‐5‐enenitrile ( 2 ; 10.11%), dimethyl tetrasulfide ( 8 ; 4.93%), and pent‐4‐enyl isothiocyanate ( 7 ; 4.45%). In addition, O‐glycosidically bound volatiles and free volatiles were isolated by solvent extraction. Sixteen volatile O‐aglycones and twelve free volatile components were identified. The main O‐aglycones were eugenol ( 19 ; 24.15%), 2‐methoxy‐4‐vinylphenol ( 11 ; 11.50%), and benzyl alcohol ( 20 ; 9.49%), and the main free volatiles were (9Z,12Z)‐octa‐9,12‐dienic acid (38.35%), hexadecanoic acid (22.64%), and phytol (5.80%). The H₂O‐soluble volatile fraction obtained by HDA, containing mostly glucosinolate degradation products and 3,4,5‐trimethylpyrazole ( 17 ), was evaluated for antimicrobial activity by determining inhibition zones with the diffusion method as well as minimal inhibitory concentrations (MIC) and minimal microbicidal concentrations (MMC) with the micro‐dilution method. The fraction expressed activity against the tested Gram‐positive and Gram‐negative bacteria as well as against yeast, with MIC values equal to or lower than 16.7 μg/ml.     

Aroma Profile of Rubus ulmifolius Flowers and Fruits During Different Ontogenetic Phases

The chemical composition of spontaneous volatile emission from Rubus ulmifolius flowers and fruits during different stages of development was evaluated by HS‐SPME‐GC/MS. In total, 155 chemical compounds were identified accounting 84.6 – 99.4% of whole aroma profile of flowers samples and 92.4 – 96.6% for fruit samples. The main constituents were α‐copaene, β‐caryophyllene, germacrene D, (E,E)‐α‐farnesene, 1,7‐octadien‐3‐one,2‐methyl‐6‐methylene, tridecane, (E)‐2‐hexenol acetate, (E)‐3‐hexenol acetate and cyperene. The results give a chemotaxonomic contribution to the characterization of the VOCs emitted from flowers and fruits during their ontogenic development.     

Phytochemical Findings Evidencing Botanical Origin of New Propolis Type from North‐West Argentina

Propolis samples from north‐west Argentina (Amaicha del Valle, Tucumán) were evaluated by palynology, FT‐IR spectra, and RP‐HPTLC. In addition, the volatile fraction was studied by HS‐SPME‐GC/MS. The botanical species most visited by Apis mellifera L. near the apiaries were collected and their RP‐HPTLC extracts profiles were compared with propolis samples. In addition, GC/MS was performed for volatile compounds from Zuccagnia punctata Cav. (Fabaceae). FT‐IR spectra and RP‐HPTLC fingerprints of propolis samples showed similar profiles. In RP‐HPTLC analyses, only Z. punctata presented a similar fingerprint to Amaicha propolis. The major volatile compounds present in both were trans‐linalool oxide (furanoid), 6‐camphenone, linalool, trans‐pinocarveol, p‐cymen‐8‐ol, and 2,3,6‐trimethylbenzaldehyde. Potential variations for the Amaicha del Valle propolis volatile fraction as consequence of propolis sample preparation were demonstrated.     

Chemical Composition and Character Impact Odorants in Volatile Oils from Edible Mushrooms

The aim of this study was to investigate the chemical composition and the odor‐active components of volatile oils from three edible mushrooms, Pleurotus ostreatus, Pleurotus eryngii, and Pleurotus abalonus, which are well‐known edible mushrooms. The volatile components in these oils were extracted by hydrodistillation and identified by GC/MS, GC‐olfactometry (GC‐O), and aroma extract dilution analysis (AEDA). The oils contained 40, 20, and 53 components, representing 83.4, 86.0, and 90.8% of the total oils in P. ostreatus, P. eryngii, and P. abalonus, respectively. Odor evaluation of the volatile oils from the three edible mushrooms was also carried out using GC‐O, AEDA, and odor activity values, by which 13, eight, and ten aroma‐active components were identified in P. ostreatus, P. eryngii, and P. abalonus, respectively. The most aroma‐active compounds were C₈‐aliphatic compounds (oct‐1‐en‐3‐ol, octan‐3‐one, and octanal) and/or C₉‐aliphatic aldehydes (nonanal and (2E)‐non‐2‐enal).     

Antifungal Activity of Volatile Compounds Produced by an Edible Mushroom Hypsizygus marmoreus against Phytopathogenic Fungi

Volatile compounds with antifungal activity produced by edible mushrooms have potential as biological control agents to combat fungal diseases and reduce fungicide use in agriculture. Here we investigated the antifungal activity of volatile compounds produced by the edible mushroom Hypsizygus marmoreus (TUFC 11906) against eight phytopathogenic fungi. The results showed that volatile compounds from the mycelia and culture filtrates (CFs) of H. marmoreus had antifungal activity against some phytopathogenic fungi. Among them, the mycelial growth and conidial germination of Alternaria brassicicola were significantly inhibited by 60 and 100%, respectively. Moreover, the volatile compounds from CFs inhibited the lesion formation of A. brassicicola on detached cabbage leaves by 94%. The volatile compounds had higher antifungal activity against A. brassicicola than other fungi. With the removal of the volatile compounds from conidia of A. brassicicola, the conidia began to germinate, which indicates fungistatic activity of the compounds. The volatile compounds were isolated from the CFs of H. marmoreus, and the major volatile compound with antifungal activity was estimated to be 2‐methylpropanoic acid 2,2‐dimethyl‐1‐(2‐hydroxy‐1‐methylethyl)propyl ester. As the volatile compound produced by H. marmoreus is a product of an edible mushroom and has fungistatic activity against some phytopathogenic fungi, especially A. brassicicola, it may be possible to use the compounds as a novel safe agent for protecting crops in the field and during storage.     

Variation of the Chemical Composition and Antimicrobial Activity of the Essential Oils of Natural Populations of Tunisian Daucus carota L. (Apiaceae)

The essential oils of Daucus carota L. (Apiaceae) seeds sampled from ten wild populations spread over northern Tunisia were characterized by GC‐FID and GC/MS analyses. In total, 36 compounds were identified in the D. carota seed essential oils, with a predominance of sesquiterpene hydrocarbons in most samples (22.63–89.93% of the total oil composition). The main volatile compounds identified were β‐bisabolene (mean content of 39.33%), sabinene (8.53%), geranyl acetate (7.12%), and elemicin (6.26%). The volatile composition varied significantly across the populations, even for oils of populations harvested in similar areas. The chemometric principal component analysis and the hierarchical clustering identified four groups, each corresponding to a composition‐specific chemotype. The in vitro antimicrobial activity of the isolated essential oils was preliminarily evaluated, using the disk‐diffusion method, against one Gram‐positive (Staphylococcus aureus) and two Gram‐negative bacteria (Escherichia coli and Salmonella typhimurium), as well as against a pathogenic yeast (Candida albicans). All tested essential oils exhibited interesting antibacterial and antifungal activities against the assayed microorganisms.     

Volatile compounds with characteristic odour in moso‐bamboo stems (Phyllostachys pubescens Mazel ex Houz. De ehaie)

Introduction – Moso‐bamboo (Phyllostachys pubescens) is well known as an edible shoot in Asia, and the stems of moso‐bamboo are used as tableware due to its characteristic odour. Despite the pleasant odour of bamboo stems, no detailed analysis of the volatile compounds has been reported. Objective – To clarify the potent odourants contributing to the characteristic aroma of the bamboo, the aroma extract dilution analysis (AEDA) method was performed through gas chromatography olfactometry (GC‐O) analysis. In addition, relative flavour activity (RFA) was calculated, in which both the flavor dilution (FD) factor and weight percentage of each compound are involved. Results – Eighty‐nine compound in bamboo stems oil were identified by GC and GC‐MS. The main components of the oil were palmitic acid (16.5%), (E)‐nerolidol (10.2%) and indole (8.1%). In sensory analysis, 18 aroma‐active compounds were detected by aroma extract dilution analysis (AEDA). The most intense aroma‐active compounds were eugenol (sweet, clove‐like, green) and (E)‐2‐nonenal (green). Conclusion – The results of the sniffing test, RFA and FD factor indicated that (E)‐2‐nonenal and eugenol were estimated to have a bamboo‐like aroma, and aldehyde compounds, such as a phenylacetaldehyde (floral) and C9–C10 unsaturated aldehydes, make the aroma of bamboo. Copyright © 2010 John Wiley & Sons, Ltd.     

Patterns in Volatile Emission of Different Aerial Parts of Caper (Capparis spinosa L.)

We analyzed the spontaneous volatile emission of different aerial parts of the caper (Capparis spinosa L.) by HS‐SPME‐GC/MS. We identified 178 different compounds of which, in different proportions based on the sample type, the main ones were (E)‐β‐ocimene, methyl benzoate, linalool, β‐caryophyllene, α‐guaiene, germacrene D, bicyclogermacrene, germacrene B, (E)‐nerolidol, isopropyl tetradecanoate, and hexahydrofarnesyl acetone. The multivariate statistical analyses seem to point out that the parameter leading the emission patterns is the function of the analyzed sample; the flower samples showed differences in the emission profile between their fertile and sterile portions and between the other parts of the plant. The green parts emission profiles group together in a cluster and are different from those of seeds and fruits. We also hydrodistilled fully bloomed caper flowers, whose volatile oil showed significant differences in the composition from those of other parts of the plant reported.     

Variability and stability of tea weevil‐induced volatile emissions from tea plants with different weevil densities,photoperiod and infestation duration

Abstract After herbivore attack, many plants emit herbivore‐induced plant volatiles (HIPVs). HIPVs can attract carnivores and/or repel herbivores, thereby mediating tritrophic plant–herbivore–carnivore interactions. HIPVs act as chemical information between organisms; hence, their variability and stability are vital. In the present study, variations in the volatile emissions, from the tea plant Camellia sinensis (O. Ktze) damaged by the tea weevil Myllocerinus aurolineatus (Voss) (Coleoptera: Curculionidae), with weevil densities, photoperiod and infestation duration, were investigated. The volatiles induced by high‐density weevils were more abundant in composition and amount than those induced by low‐density weevils, whether at noon, night or after weevil removal. The induced volatile emissions were similar on the second and third day after infestation, and the emissions of the major induced compounds displayed diurnal cycles. Linalool, (E,E)‐α‐farnesene, and benzyl nitrile were emitted mainly at noon, whereas 1,3,8‐p‐menthatriene and (E)‐β‐ocimene were maximally emitted at night. Given the different emission dynamics, significant differences were found between noon‐ and night‐induced volatiles. In summary, tea plants damaged by different weevil densities emitted a relatively stable signal at a particular time. This stability could be attributed to the similarities under the two densities of the main induced volatile compounds, their relative ratios and the emission dynamics of the induced volatiles.     

Enantiomer Distribution of Major Chiral Volatile Organic Compounds in Selected Types of Herbal Honeys

In this article, volatile organic compounds in 14 honey samples (rosemary, eucalyptus, orange, thyme, sage, and lavender) were identified. Volatile organic compounds were extracted using a solid phase microextraction method followed by gas chromatography connected with mass spectrometry analysis. The studied honey samples were compared based on their volatile organic compounds composition. In total, more than 180 compounds were detected in the studied samples. The detected compounds belong to various chemical classes such as terpenes, alcohols, acids, aldehydes, ketones, esters, norisoprenoids, benzene and furane derivatives, and organic compounds containing sulfur and nitrogen heteroatom. Ten chiral compounds (linalool, trans‐linalool oxide, cis‐linalool oxide, 4‐terpineol, α‐terpineol, hotrienol, and four stereoisomers of lilac aldehydes) were selected for further chiral separation. Chirality 26:670‐674, 2014. © 2014 Wiley Periodicals, Inc.     

The Volatile Profiles of a Rare Apple (Malus domestica Borkh.) Honey: Shikimic Acid‐Pathway Derivatives,Terpenes, and Others

The volatile profiles of rare Malus domestica Borkh . honey were investigated for the first time. Two representative samples from Poland (sample I) and Spain (sample II) were selected by pollen analysis (44–45% of Malus spp. pollen) and investigated by GC/FID/MS after headspace solid‐phase microextraction (HS‐SPME) and ultrasonic solvent extraction (USE). The apple honey is characterized by high percentage of shikimic acid‐pathway derivatives, as well as terpenes, norisoprenoids, and some other compounds such as coumaran and methyl 1H‐indole‐3‐acetate. The main compounds of the honey headspace were (sample I; sample II): benzaldehyde (9.4%; 32.1%), benzyl alcohol (0.3%; 14.4%), hotrienol (26.0%, 6.2%), and lilac aldehyde isomers (26.3%; 1.7%), but only Spanish sample contained car‐2‐en‐4‐one (10.2%). CH₂Cl₂ and pentane/Et₂O 1 : 2 (v/v) were used for USE. The most relevant compounds identified in the extracts were: benzaldehyde (0.9–3.9%), benzoic acid (2.0–11.2%), terpendiol I (0.3–7.4%), coumaran (0.0–2.8%), 2‐phenylacetic acid (2.0–26.4%), methyl syringate (3.9–13.1%), vomifoliol (5.0–31.8%), and methyl 1H‐indole‐3‐acetate (1.9–10.2%). Apple honey contained also benzyl alcohol, 2‐phenylethanol, (E)‐cinnamaldehyde, (E)‐cinnamyl alcohol, eugenol, vanillin, and linalool that have been found previously in apple flowers, thus disclosing similarity of both volatile profiles.     

Influence of volatile compounds from herbivore‐damaged soybean plants on searching behavior of the egg parasitoid Telenomus podisi

Parasitoids use herbivore‐induced plant volatiles (HIPVs) to locate their hosts. However, there are few studies in soybean showing the mechanisms involved in the attraction of natural enemies to their hosts and prey. The objective of this study was to evaluate the influence of volatile organic compounds (VOCs) of soybean, Glycine max (L.) Merr. (Fabaceae) (cv. Dowling), that were induced after injury caused by Euschistus heros (Fabricius) (Hemiptera: Pentatomidae), on the searching behavior of the egg parasitoid Telenomus podisi Ashmead (Hymenoptera: Scelionidae). Four HIPVs from soybean, (E,E)‐α‐farnesene, methyl salicylate, (Z)‐3‐hexenyl acetate, and (E)‐2‐octen‐1‐ol, were selected, prepared from standards at various concentrations (10^?6 to 10^?1 m ), and tested individually and in combinations using a two‐choice olfactometer (type Y). Telenomus podisi displayed a preference only for (E,E)‐α‐farnesene at 10^?5 m when tested individually and compared to hexane, but they did not respond to the other compounds tested individually at any concentration or when combinations of these compounds were tested. However, the parasitoids stayed longer in the olfactometer arm with the mixture of (E,E)‐α‐farnesene + methyl salicylate at 10^?5 m than in the arm containing hexane. The results suggest that (E,E)‐α‐farnesene and methyl salicylate might help T. podisi to determine the presence of stink bugs on a plant. In addition, bioassays were conducted to compare (E,E)‐α‐farnesene vs. the volatiles emitted by undamaged and E. heros‐damaged plants, to evaluate whether (E,E)‐α‐farnesene was the main cue used by T. podisi or whether other minor compounds from the plants and/or the background might also be used to locate its host. The results suggest that minor volatile compounds from soybean plants or from its surroundings are involved in the host‐searching behavior of T. podisi.     

Chemical Composition and Aroma Evaluation of Essential Oils from Evolvulus alsinoides L.

The aim of this study was to investigate the chemical composition and the odor‐active compounds of the essential oils from Evolvulus alsinoides, which is a well‐known edible and medicinal plant. The volatile compounds in the oils were identified by hydrodistillation (HD) and solvent‐assisted flavor evaporation (SAFE) in combination with GC, GC/MS, GC/O (=olfactometry), aroma extract dilution analysis (AEDA), and relative flavor activities (RFA values). The most abundant compound in the HD oil was cis‐α‐necrodol (12.62%), an irregular monoterpene with a cyclopentane skeleton, which is very unusual in the plant kingdom. In the SAFE oil, the main components included 2‐butoxyethanol (9.01%), benzyl alcohol (8.01%), and γ‐butyrolactone (7.37%). Through sensory analysis, 21 aroma‐active compounds were identified by GC/O. The most intense aroma‐active compounds in the HD oil were hexan‐1‐ol and γ‐nonalactone, both of which showed high RFA values. α‐Methyl‐γ‐butyrolactone and dimethyl sulfone contributed more strongly to the aroma of the SAFE oil. These results imply that the essential oils of E. alsinoides deserve further investigation in the food industry.     

Chemical Composition and Antimicrobial Activity of the Volatile Fractions from Leaves and Flowers of the Wild Iraqi Kurdish Plant Prangos peucedanifolia Fenzl

The volatile fractions isolated from Prangos peucedanifolia Fenzl leaves and flowers were investigated for their phytochemical composition and biological properties. Flower and leaf hydrodistillation afforded 3.14 and 0.49 g of yellowish oils in 1.25 and 0.41% yields, respectively, from dry vegetable materials. According to the GC‐FID and GC/MS analyses, 36 (99.35% of the total oil composition) and 26 compounds (89.12%) were identified in the two oils, respectively. The major constituents in the flower volatile fraction were β‐pinene (35.58%), α‐pinene (22.13%), and β‐phellandrene (12.54%), while m‐cresol (50.38%) was the main constituent of the leaf volatile fraction. The antimicrobial activity was evaluated against several bacterial and fungal strains, on the basis of the minimum inhibitory concentration (MIC) by the micro‐ and macrodilution methods. The two volatile fractions showed moderate antifungal and antibacterial activities, especially against Trichophyton rubrum (MIC of 2×10³ μg/ml), Streptococcus mutans, Streptococcus pyogenes, and Staphylococcus aureus (MIC≤1.9×10³ μg/ml for all).     

Electroantennogram responses of aphid nymphs to plant volatiles

Abstract. Electroantennogram (EAG) responses of immature aphids were investigated for 30 plant volatile compounds in third‐ and fourth‐stadium nymphs of the black bean aphid, Aphis fabae. The nymphs were destined to develop into adult alate (winged) virginoparae. The EAG response profiles were similar to those previously reported in the adults. Among the compounds tested, hexanonitrile elicited the largest EAG responses in both nymphal stadia, corresponding to previously reported results with adults. Six‐carbon aliphatic compounds showed relatively higher EAG activities in the nymphs but, in contrast, (E)‐2‐hexenal, benzaldehyde, α‐pinene and β‐pinene, and citronellal elicited relatively smaller EAG responses in nymphs than adults. Although overall EAG response profiles were similar between the third and the fourth stadia for the majority of the volatiles, four aldehyde compounds, hexanal (E)‐2‐heptenal, 2‐hydroxybenzaldehyde and citronellal, showed relatively higher EAG activities in the third than in the fourth stadium. The present study indicates that aphid nymphs possess a functional olfactory receptor system before the antennae are fully developed morphologically and physiologically.