Headspace Gas Chromatography Research Articles

Determination of ethanol in hand sanitizers by a self-built headspace device and gas chromatography

Introduction: Headspace sampling coupled with gas chromatography (HS-GC) has been proven to be a very useful technique for the determination of volatile and semivolatile components in complex matrices by avoiding tedious sample preparation. In this study, the HS system was fabricated at low cost, exploiting unused GC sensor-heaters. Methods: The accuracy and stability of the self-built HS were evaluated, and the HS-GC-FID system was applied to the determination of ethanol in hand sanitizer gels. The HS operating conditions, including temperature and equilibrium time, were optimized using a central composite face-centered design (CCFC) and MODDE 5.0 software. Samples incubated at 80 °C for 25 min gave the best results with respect to sensitivity and reproducibility (RSD < 3%). Results: The calibration curve showed high linearity (R2 > 0.998) in the range of 0.25 to 2.5% (w/w) ethanol, using acetonitrile 1% (w/w) as an internal standard. The recovery ranged between 104.1 and 108.3% using spiked samples on real matrices. A small survey of 11 commercially available samples collected in the local markets showed that only one of them did not meet the standards set by the FDA.

A simple and universal headspace GC-FID method for accurate quantitation of volatile amines in pharmaceuticals.

Volatile amines are reagents commonly used in pharmaceutical manufacturing of intermediates, active pharmaceutical ingredients (APIs), and drug products as participating regents for chemical reactions and optimization of product yield. Due to their compound specific daily allowable intake, residual volatile amines are required by regulatory agencies to be monitored and controlled in pharmaceutical products intended for human consumption. However, the accurate quantification of residual volatile amines in pharmaceutical entities can often be challenging as these analytes may chemically react and/or interact with the sample matrix. Herein, we describe a simple and universal headspace gas chromatography with flame ionization detection (HS-GC-FID) method capable of separating 14 commonly used volatile amines. The chemical activity of the volatile amines with the API matrix were mitigated by using 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) as an additive to reduce matrix effects in conventional high-boiling diluents. The addition of DBU drastically improved the detectability and method accuracy of the residual volatile amines in an acidic API, namely, Ketoprofen®. Additionally, DBU was employed as a GC deactivation reagent to ensure interfacial adsorption of the analytes to GC components were reduced, thereby improving method precision. Method validation showed acceptable linearity, limit of detection, limit of quantitation, solution stability, precision, and robustness. Separation specificity, evaluated by observing the chromatographic resolution of the volatile amines with one-another and against a set of 23 common residual solvents, were shown to be acceptable for most peak pairs.

Head Space-GC Analysis of N, N-Diethyl-M-Toluamide (DEET) in Human Postmortem Specimens: A Forensic Case Report

In India, self-poisoning through pesticides is an increasingly common form of suicide. Although deleterious insect repellents (sub-classification of pesticides) are regulated or interdicted, but significant data related to misuse can be procured. Meanwhile, the emergence of novel commercially manufactured products containing insect repellant as a prime ingredient without public and scientific apprehension regarding detrimental impacts has exacerbated the circumstances. Forensic examination of the viscera of the deceased discerned that the death was attributable to Diethyltoulamide intoxication. As no conspiracy behind the death was evident, hence it was pellucid to comprehend that the insect-repellant product was self-administered. Headspace GC analysis of viscera showed noteworthy results in concern to the identification of Diethyltoulamide. This study attempts to identify the most appropriate pesticide and determine whether it is present in biological matrices. The samples were received from the hospital and processed in the Forensic Science Laboratory for testing on possible poisoning. The pesticides Malathion, Dicholorovos, Baygon, and Lindane were utilised as a kind of control. The extraction of pesticides from various biological matrices was done using the solvent extraction method.

Study on drug substance degradation under headspace GC conditions of residual solvent analysis.

Isobutyraldehyde (IBA) was detected in drug substance (DS) containing an amino acid group using a headspace-gas chromatography (HS-GC) method. High-performance liquid chromatography-mass-spectrometry (HPLC-MS) data from an HS vial confirmed that IBA was a degradant. The HS-GC method was modified to minimize IBA by keeping the HS oven temperature lower than 80 °C.

Analysis of the formation mechanism of volatile and non-volatile flavor substances in corn wine fermentation based on high-throughput sequencing and metabolomics

The purpose of this study was to reveal the relationship between core microorganisms and flavor substances in the fermentation process of corn wine. Microbial diversity, volatile and non-volatile flavor substances were detected by high-throughput sequencing (HTS), headspace solid phase micro-extraction gas chromatography-mass spectrometry (HS-SPME/GC–MS) and gas chromatography time of flight mass spectrometry (GC-TOF-MS). High performance liquid chromatography (HPLC) was used to detect organic acids in corn wine fermentation, and its physiochemical properties were tracked. The results showed that physiochemical factors changed obviously with fermentation time. Bacillus, Prevotella_9, Acinetobacter and Gluconobacter were the predominant bacterial. Rhizopus and Saccharomyces were the dominant fungi. Acetic acid and succinic acid were important organic acids in corn wine. According to variable importance of projection (VIP) > 1 and P < 0.05, 24 volatile flavor substances with significant difference were screened out from 52 volatile flavor substances. Similarly, 25 non-volatile flavor substances with significant differences were screened out from the 97 reliable metabolites identified by 223 chromatographic peaks. Eight key metabolic pathways were enriched from 25 non-volatile flavor substances according to path influence values > 0.1 and P < 0.05. Based on Two-way Orthogonal Partial Least Squares (O2PLS) model and Pearson correlation coefficient, Saccharomyces, Rhizopus, uncultured_bacterium, Aneurinibacillus, Wickerhamomyces and Gluconobacter may be the potential volatile flavor-contributing microorganism genus in corn wine. The Pearson correlation coefficient showed that Saccharomyces was significantly positively correlated with malic acid, oxalic acid, valine and isoleucine, and Rhizopus was positively correlated with glucose-1-phosphate and alanine. These findings enhanced our understanding of the formation mechanism of flavor substances in corn wine and provided the theoretical basis for stabilizing flavor quality of corn wine.

Method Development and Validation for the estimation of Class-2 Residual Solvents in Valacyclovir by HS-GC

A simple headspace gas chromatographic (HS-GC) is a novel approach for the simultaneous detection of residual solvents such as ethanol has been devised and validated. Valacyclovir in ethanol, acetone, isopropyl alcohol, toluene, and acetic acid utilizing nitrogen as the carrier gas at a 4 mL/min flow rate with DB-620 column (80mx0.22mm) 1.8m inner diameter by using a flame ionization detector (FID). The headspace gas chromatographic technique provides an acceptable symmetry and resolution for all these residual solvents. The suggested method was shown to be satisfactory for determining the residual solvents of five distinct types. The method's validation findings showed that it is specific, exact, accurate, rugged, linear, and robust, with recoveries ranging from 80-120%. The %RSD (Relative Standard Deviation) for six injections should be NMT-10%. The correlation coefficient R2 ≥ 0.999.

Olive Oil Produced from Olives Stored under CO2 Atmosphere: Volatile and Physicochemical Characterization.

In this study, an alternative debittering technique for olives, invented and patented by Prof. Vassilis Dourtoglou, was employed. Olive fruits (Olea europaea cv. Megaritiki) were stored under CO2 atmosphere immediately after harvest for a period of 15 days. After the treatment, a sensory evaluation between the olives stored under CO2 and those stored under regular atmospheric conditions (control) was performed. Additionally, the CO2-treated olives were used for the cold press of olive oil production. The volatile profile of the olive oil produced was analyzed using headspace solid-phase microextraction (HS-SPME) and gas chromatography coupled to mass spectrometry (GC-MS). A total of thirty different volatile compounds were detected. The volatile characteristics of olive oil are attributed, among others, to aldehydes, alcohols, esters, hydrocarbons, alkanes, and terpenes. The volatile compounds' analysis showed many differences between the two treatments. In order to compare the volatile profile, commercial olive oil was also used (produced from olives from the same olive grove with a conventional process in an industrial olive mill). The antioxidant activity, the content of bioactive compounds (polyphenols, α-tocopherol, carotenoids, and chlorophylls), and the fatty acids' profile were also determined. The results showed that the oil produced from CO2-treated olives contains different volatile components, which bestow a unique flavor and aroma to the oil. Moreover, this oil was found comparable to extra virgin olive oil, according to its physicochemical characteristics. Finally, the enhanced content in antioxidant compounds (i.e., polyphenols) not only rendered the oil more stable against oxidation but also better for human health. The overall quality of the olive oil was enhanced and, as such, this procedure holds great promise for future developments.

Comparison of Quantitative Values of Headspace Gas Chromatography--Mass Spectrometry and a Formate Quantification Kit in Blood Formate Quantification.

Methanol poisoning is caused by the toxicity of formate, a by-product of methanol metabolism. Measurement of blood formate concentrations is required for emergency treatment and investigation of the cause of death. In this study, we measured concentrations of formate in the plasma of a patient with methanol poisoning using headspace gas chromatography--mass spectrometry (HS-GC--MS) and a formate assay kit. Results showed a discrepancy as the quantitative values of the kit were higher than those of HS-GC--MS. Metabolic profiling of low-molecular-weight organic compounds in patient plasma samples showed that the concentrations of lactate were correlated with the values obtained using the kit. We observed a progression when lactate and lactate dehydrogenase were added to the kit reaction simultaneously, even in the absence of formate. Moreover, disulfiram, an aldehyde dehydrogenase inhibitor, suppressed the values of patient plasma samples in the formate assay kit, implying that formate production from remaining methanol in patient plasma samples via formaldehyde occurred during the kit reaction. The reactions of the kit with lactate and methanol were undesirable for accurate measurement of formate concentration in the sample. However, considering that elevated concentrations of lactate and remaining methanol both cause acidosis and are dangerous to the body, cross-reactions with lactate and methanol in the formate assay kit may be acceptable for rapid diagnosis in facilities where HS-GC--MS and other physical and chemical equipment are unavailable.

Identification of characteristic aroma and bacteria related to aroma evolution during long-term storage of compressed white tea.

Compressed white tea (CWT) is a reprocessed tea of white tea. Long-term storage has greatly changed its aroma characteristics, but the material basis and transformation mechanism of its unique aroma are still unclear. In this study, flavor wheel, headspace gas chromatography ion mobility spectroscopy, chemometrics, and microbiomics were applied to study the flavor evolution and important aroma components during long-term storage of CWT, and core functional bacteria were screened. During long-term storage, the aroma of CWT gradually changed from sweet, fruity and floral to stale flavor, woody and herbal. A total of 56 volatile organic compounds (VOCs) were identified, 54 of which were significantly differences during storage. The alcohols content was the highest during 1-5 years of storage, the esters content was the highest during 7-13 years of storage, and the aldehydes content was the highest during 16 years of storage. Twenty-nine VOCs were identified as important aroma components, which were significantly correlated with 6 aroma sub-attributes (P < 0.05). The functional prediction of bacterial community reminded that bacterial community could participate in the transformation of VOCs during storage of CWT. Twenty-four core functional bacteria were screened, which were significantly associated with 29 VOCs. Finally, 23 characteristic differential VOCs were excavated, which could be used to identify CWT in different storage years. Taken together, these findings provided new insights into the changes in aroma characteristics during storage of CWT and increased the understanding of the mechanism of characteristic aroma formation during storage.

Non-targeted screening and trimethylamine determination in Tilletia foetida-infected wheat using HS-SPME-GC-MS

Common bunt disease of wheat in China is caused mainly by Tilletia foetida. However, reliable approaches for determining disease-associated biochemical markers are rarely reported. Here, a headspace-solid-phase microextraction coupled with headspace GC-tandem mass spectrometry (HS-SPME-GC-MS) was used to screen volatile substances in infected wheat, and an optimal chemical marker was selected to establish analytical methods for disease determination. Non-targeted screening of 13 volatile compounds unique to diseased wheat allowed a metabolite with rotten fish-like smell, trimethylamine (TMA), to be selected as the inspection marker. Subsequently, two analytical methodologies, HS-SPME-GC-MS and headspace gas chromatography with flame ionization detection (HS-GC-FID), were established to determinate the TMA content in wheat. The linear relationship, recovery and reproducibility of the methods were validated. The limit of detection (LOD) was 0.02 µg/kg for the former method, 5000-fold lower than that for the latter. When analysing samples, HS-SPME-GC-MS showed excellent sensitivity and allowed for the determination of 0.05% infected kernels among whole wheat grains. Therefore, TMA determination using HS-SPME-GC-MS is an effective alternative method to detect wheat common bunt disease occurring at extremely low infection rate.

Comparative Evaluation of Carboxyhemoglobin Quantification in Postmortem Whole Blood by CO-Oximetry and Headspace Gas Chromatography with Flame Ionization Detection and Atom Absorption Spectrophotometry.

A comparative evaluation of two methods used for carboxyhemoglobin (COHb) determination in postmortem whole blood was performed: carbon monoxide (CO)-oximetry measuring at 128 wavelengths and headspace gas chromatography with flame ionization detection (HS-GC--FID) where CO was determined after catalytic reduction of CO to CH4 and Fe was determined by atom absorption spectrophotometry (AAS, 248.3 nm). An aliquot of 100 µL whole blood was loaded into the CO-oximetry module. In the HS-GC--FID analysis, to 1.0 mL of whole blood, 3.0 mL of saponin solution was added, mixed and centrifuged. To 20 mL HS vials, 400 µL of the supernatant was added and the vials were immediately sealed. One milliliter of potassium hexacyanoferrat (III) solution was added through the HS septum and mixed. The samples were incubated at 70°C for 5 min. CO was separated using He as carrier gas and a CP-Molsieve 5 Å PLOT capillary column. Fe was determined using 400 µL of the saponin supernatant diluted to 10 mL by water. During a period of ∼3years, 124 postmortem whole blood samples were analyzed. Bland-Altman method comparison showed satisfactory agreement and no significant bias between the methods for the whole saturation range (5 to 85% COHb). Five samples, all with %COHb >40, showed deviations of more than 10% COHb in absolute terms. One sample, in the lower COHb range <10%, was false negative on the CO-oximetry method. The between-assay accuracy, reported as bias, at 60% COHb was -0.8% and -9.0%, and precision, reported as relative standard deviation, was 1.6% and 7.7%, for the CO-oximetry and HS-GC--FID-AAS methods, respectively. Both methods obtained satisfactory results in proficiency testing rounds, with z-scores <±2 (n = 11). This study showed that the CO-oximetry method based on the 128-wavelength principle and the HS-GC--FID-AAS method are comparable and satisfactory for %COHb determination in postmortem whole blood.

Gas chromatography for β-caryophyllene determination in St. John’s wort infused oil

Hypericum Perforatum L., commonly known as St. John’s wort, is one of the best-known medicinal herbs. St. John’s wort infused oils are mostly used externally, but can be taken orally for stomach ulcers, are used as a food supplement. Therefore, it is important to develop methods that allow a rapid and reliable determination of β-caryophyllene, which is one of the main active components of St. John’s wort. In this work, two gas chromatographic techniques, static headspace gas chromatography (SHS-GC) and solid phase microextraction, combined with gas chromatography-mass spectrometry (SPME-GC-MS), were examined for the determination of β-caryophyllene in St. John’s wort infused oils. A comparison of these two techniques revealed that SPME-GC-MS can be successfully applied for the analysis. The other method, SHS-GC, is simple and fast, but because of its high detection limit, it can only be applied to oil samples with β-caryophyllene concentration higher than 0.1 mg/g.

Residual Solvents by GC-HS Procedure for Etoricoxib: Method Development and Validation Report

The intent of this research paper was to describe a “headspace gas chromatography (HGC)” procedure development and its completely validation for the analysis of residuals of methanol (MTL), isopropyl alcohol (IPL), t-butanol (TBL), toluene (TLE) and dimethylformamide (DFL) simultaneously in Etoricoxib (EIB). The experimentations are done on HGC system fitting with flame ionization type detector employing DB-624 silica fused capillary column (stationary phase) and nitrogen gas (mobile phase). The injector port and detector port temperatures were kept at 200 oC and 260 oC, respectively. N-methyl pyrrolidone was diluent. The MTL, IPL, TBL, TLE, and DFL detection (LOD) and quantitation (LOQ) values were much smaller than their ICH specification level concentrations. The linear corelation evaluated through range of LOQ to 150% of ICH specification level concentrations for MTL, IPL, TBL, TLE, and DFL of ICH. The regression coefficients for MTL, IPL, TBL, TLE, and DFL were ≥0.9950, and the diagrams of theoretic residuals concentration versus gotten peak response are linear. The HGC procedure proposed was represented by great accuracy, precision, ruggedness and specificity. For a minimum of 48 hr, the EIB sample with MTL, IPL, TBL, TLE and DFL is stable while managed to keep at ambient temperature. The current developed and completely validated HGC procedure can run effectively for EIB residual solvents (MTL, IPL, TBL, TLE and DFL) assessing in active pharma ingredient production.

Emission characteristics of volatile organic compounds from regenerated PET fibers based on the headspace gas chromatograph

AbstractThe emission behaviors of volatile organic compounds (VOCs) can be characterized by the key parameters: the initial concentration (m0), partition coefficient (K) and the diffusion coefficient (D). The MHE‐GC method was validated to determine the emission characteristics of VOCs (Benzene, Toluene, Ethylbenzene, P‐xylene, M‐xylene, Styrene, and O‐xylene) in regenerated PET fibers. This work has studied the effects of fiber size, sample weight incubation time and incubation temperature on the detection of these VOCs. It showed finer fiber sample with less sample facilitated the emission of VOCs in fibers above detectable limit. The optimized headspace incubation time, fiber sample weight, and headspace incubation temperature were selected as 60 min, about 0.8 g and 110 °C, respectively. In addition, incubation temperature has a negative effect on the gas/fiber partition coefficient. The fibers with greater fineness showed higher activation energy, which revealed a positive release of VOCs from fibers. The relative standard deviation (RSD/%) of detection was 3%–10%. The present method is reliable and suitable for use in testing of the VOCs content in fiber‐related materials.

Diffusion of Carbon Monoxide and Hydrogen Cyanide to Muscles and Blood-An Experimental Study.

Postmortem carbon monoxide (CO) and hydrogen cyanide (HCN) diffusion under ambient conditions was assessed in a human cadaver model. The main objective of this study was to determine whether the postmortem diffusion of HCN and CO greatly affected the determination of HCN, carboxyhemoglobin (COHb), and carboxymyoglobin (COMb). Layered samples of blood, musculocutaneous, and muscular specimens were collected from the adult cadavers and placed in the tight chambers designed for the purpose of this experiment. The specimens were treated with CO and HCN for 24 h. COHb and COMb were determined using headspace gas chromatography (GC) with an O-FID detector while the HCN values were assessed using a GC headspace with an NPD detector. It was shown that the skin substantially limited the diffusion of CO which penetrated the superficial layers of the muscle very slightly, all the while not affecting the blood level of COHb in the 4.5 cm layer of the muscle located underneath. There were no differences regarding the CO diffusion between superficially charred and thermally coagulated compared to that observed in intact integuments. In addition, the cutaneous sample deprived of the adipose layer was not shown to be a barrier to the moderate diffusion of CO into the blood layer below. HCN was found to easily diffuse from the skin to the blood vessels (vein specimens), and partial charring and thermocoagulation of the superficial muscular layer favored the diffusion of cyanides into the tissues. Similarly to CO, HCN diffusion to the blood and muscles was greatly limited by the adipose layer.

Determination of carbonate content in barite ore by headspace gas chromatography.

This paper reports a method for determining the carbonate content in barite ore using headspace gas chromatography. Based on the acidification reaction, the carbonate in the barite ore was converted to CO2 in a closed headspace vial. When the carbonate content was significant, the pressure caused changes in the CO2 and O2 signals and affected the measurement accuracy. It was found that carbonate content is proportional to the intensity ratio of the CO2 to O2 signals. Thus, the carbonate content in barite ore can be measured indirectly using a theoretical model. The results showed that the carbonate in 3g of barite ore sample with a particle size of 74μm could react completely with a hydrochloric acid solution (2mol/L) at 65°C for 5min. The method described herein had good precision (relative standard deviation<4.14%) and accuracy (relative differences<6.12%). Further, the limit of quantification was 0.07mol/L. Owing to its simplicity and speed, this method can be used for the batch determination of carbonate content in barite ore.

Essential oil from Sabina chinensis leaves: A promising green control agent against Fusarium sp.

Sabina chinensis is a woody plant with important ecological functions in different regions of China, but its essential oils (EO) against plant pathogenic fungi remain largely undetermined. The purpose of our study was to assess the chemical composition and antifungal activity of S. chinensis EO based on optimization of the extraction process. In this study, an actionable and effective model with the experimental results and identified optimum conditions (crushing degree of 20 mesh, liquid-solid ratio of 10.1:1, immersion time of 9.1h) was established successfully to achieve an extraction yield of 0.54%, which was basically consistent with the theoretical value. A total of 26 compounds were identified using headspace gas chromatography-mass spectrometry (GC-MS) and showed that the major constituent was β-phellandrene (26.64-39.26%), followed by terpinen-4-ol (6.53-11.89%), bornyl acetate (6.13-10.53%), etc. For Petri plate assays, our experiments found for the first time that S. chinensis EO revealed high and long-term antifungal activity against the tested strains, including Fusarium oxysporum and Fusarium incarnatum, at EC50 values of 1.42 and 1.15 µL/mL, which especially reached approximately 76% and 90% growth inhibition at a dose of 0.2 µL/mL, respectively. Furthermore, the antifungal activity of EO from different harvest periods showed remarkable variation. The orthogonal partial least-squares discriminant analysis (OPLS-DA) method revealed 11 metabolites with chemical marker components, and 5 of its potential antifungal activities, terpinen-4-ol, α-terpineol, α-elemol, γ-eudesmol, and bornyl acetate, were strongly correlated with the mycelial inhibition rate. In total, this study explored the antifungal activity of EO against root rot fungus as a potential fungicide and provided valuable information into developing potential products from natural agents.

Volatilome evolution during storage and in vitro starch digestibility of high-power ultrasonication pretreated wholegrain Oryza sativa L.

The potential of high-power ultrasonication (HPU) to enhance the physicochemical stability of bran-containing cereal products has been demonstrated, but the information concerning how the wholegrain volatilome and key quality-related chemical reactions evolve responding to HPU remains scarcely reported. The objective of this work was to examine the headspace volatile fingerprinting features of sonicated wholegrain brown rice (WBR; 400 W, 28 kHz, 30 min) following an accelerated storage testing (37 °C, 20 days), and simultaneously to identify the key chemical reactions induced by ultrasonication. A total of 70 aroma compounds were identified by the untargeted headspace GC–MS, including 9 alkanes, 6 alkenes, 15 aldehydes, 6 furans, 12 ketones, 9 alcohols and 13 miscellaneous compounds. Multivariate statistical analysis demonstrated that HPU pretreatments before a storage process significantly influenced the volatilome evolution, as revealed by a clear classification between sonicated and unsonicated grains. Supervised orthogonal partial least squares discriminant analysis identified the volatiles including acetic acid, pentanoic acid, hexanal, ethyl hexanoate and 2-pentyl-furan as HPU-related markers, inferring that HPU mainly modified the chemical reactions involving lipid decomposition, free fatty acids oxidation and esterification. This was further confirmed by targeted monitoring of lipid peroxidation products, with 13.22–14.84 % of MDA contents reduced (p < 0.05) in sonicated samples after storage. Besides, the ultrasonic effects resulted in a slight improvement of in vitro starch digestibility of WBR samples depending on the rice ecotype. This investigation demonstrated the potential of HPU pretreatments for prolonging the oxidative stability of WBR grains, without significantly compromising digestion properties.

Fast Catalyst Ratio Estimate in Gas Phase Polyethylene Dual Catalyst System

AbstractOne approach used in the industry to improve the properties of polyethylene is to use multi‐reactor with a single catalyst or multiple catalysts in a single reactor. In the latter case, two catalysts with distinct kinetics are selected to achieve the desired product properties. Such mixed catalyst systems enable tailored and advantageous properties at the cost of more challenging process control, because the ratio of the two catalysts serves as an additional manipulated variable. A fast method to estimate the ratio of active catalysts using headspace gas chromatography measurements is proposed here. In this method, a small perturbation in the feed rate is introduced to induce transient responses in the gas phase concentration. Ideally, with known responses from each individual catalyst, the active catalyst ratio can be estimated. To demonstrate this concept, a process model is developed in Aspen Plus. A set of dynamic simulation is performed to understand the responses of each catalyst and the mixed catalyst system, to changes in feed comonomer concentration. The results demonstrate that this method has significantly faster responses compared to feedback from bulk polymer properties and induces minimal process upset or product off‐spec due to small perturbations in a short period of time.

Scented at day and night: diel variation in the floral scent chemistry of the threatened tree Amburana cearensis (Leguminosae) and effects of this variation on its pollinators

The reproductive success of most angiosperms depends on pollinators. A combination of floral traits, including olfactory cues, are involved in attracting these animals. The emission and chemical composition of floral volatile organic compounds (VOCs) is often associated with the sensory preferences of the most frequent floral visitors. However, the relationship between VOCs and nocturnal and diurnal pollinators is poorly known in tropical dry forests. Our study describes the dynamics of floral scent chemistry throughout the anthesis of Amburana cearensis (Leguminosae) and its association with floral visitors in the Caatinga dry forest, one of the largest expanses of seasonally dry tropical forest (SDTF) in the world. The species is a threatened tree with high socioeconomic importance in the Caatinga and other SDTFs of South America (exploited for timber and medicinal uses). Its flowers emit an intense, pleasant and sweet fragrance throughout anthesis, starting at ∼16:00 h and ending at ∼15:00 h of the following day (lasting 23 h). We used dynamic headspace sampling and gas chromatography coupled with mass spectrometry (GC-MS) to collect and analyze the floral scent within a natural population. The fragrance of A. cearensis was characterized by the dominant presence of monoterpenes, especially linalool and (Z)-linalool oxide (furanoid) and (Z)-linalool oxide (pyranoid), as well as (E,E)-α-farnesene and an aromatic benzenoid, benzyl alcohol. The relative proportions of floral VOCs emitted by A. cearensis changed during anthesis, with a marked presence of linalool in daytime samples (day: 45% / night: 27%) and furanoid (Z)-linalool oxide at night (day: 12% / night: 29%). The dominance of benzyl alcohol, linalool and its oxides in the floral scent of A. cearensis is possibly related to the attraction of nocturnal and diurnal Lepidoptera as pollen vectors. Indeed, we observed six species of butterflies and seven species of moths pollinating the flowers of A. cearensis in 41 h of observations. However, other terpenoids in the fragrant blend possibly contribute to the attraction of additional diurnal floral visitors, including bees, which are also effective pollinators. The continuous scent production throughout anthesis, and the changes in floral scent with a peak of linalool production during the day, may be related to the wide spectrum of pollinators of A. cearensis. This association between floral visitors and dynamics of VOCs presentation may therefore increase the reproductive success of the species, which only blooms every two years and is strictly self-incompatible.