Volatile compounds of arazá fruit (Eugenia stipitata McVaught)

The arazá (Eugenia stipitata McVaugh) is native from the Peruvian part of the Amazonian forest. The fruit is a sphere, characterized by its intense aroma and acid flavor and green to yellow peel at maturity. The edible part, a creamy-white pulp, is eaten fresh or is used to prepare juices, marmalades, ice creams and liquors. Volatile compounds were isolated from arazá by continuous liquid-liquid extraction using pentane-dichloromethane [1 : 1 (v/v)] for 8 h, and analyzed by GC-FID and GC-MS using HP-Innowax fused silica column. The concentrated extract showed aroma notes resembling the flavor of fresh fruit, described as sweet-green-fruity. A total amount of 7.9 mg of volatile compounds per kilogram of fresh fruit was obtained. In total, 27 esters, 20 terpenes, five alcohols, six carbonyls, five acids, three hydrocarbons, two lactones and two sulfur-compounds were identified. Of the 70 components identified, 53 are reported for the first time in this fruit. Esters (54.8 % of the total composition) were the most abundant compound class. Ethyl octanoate, ethyl dodecanoate and ethyl decanoate were found to be the major constituents. Other significant compounds were: ethanol, 1-hexanol, globulol, 2-methylbutanoic acid, hexanoic acid, octanoic acid, 3-methyl-2-buten-1-ol and 2-furfural. INTRODUCTION Colombia has a natural diversity of tropical fruits with distinctive exotic flavors appealing to the producer that they could be an important source of income. However, the volatile composition responsible for their flavors has not yet been characterized widely. Among them, arazá (Eugenia stipitata McVaught), belonging to the Myrtaceae family, is an indigenous Amazonian tree widespread in different regions of Colombia. This fruit is also known as araçaboi in Brazil or as pichi or sororia in Peru. The fruit, characterized by its intense aroma and acid flavor, is round, about 12-15 cm in diameter, 800 g maximum weight, and green to yellow peel at maturity. The edible part, a creamy-white pulp, is eaten fresh or is used to prepare juices, marmalades, ice creams and liquors.1-3 As far it is known, there are only two reports on the composition of the volatile compounds of this fruit.4,5 In both works, 30 and 65 volatiles were identified, respectively. The main purpose of this study was to identify additional arazá fruit (Eugenia stipitata McVaught) compounds that may contribute to its delicate flavor. Revista CENIC Ciencias Químicas, Vol. 38, No. 3, 2007. 364 MATERIALS AND METHODS Fresh mature arazá fruits were picked from bushes grown in Caquetá, Colombia, and transported by airplane to the laboratory within 24 h after harvest. The fruits were allowed to ripen at room temperature. After separation of the skin and seeds the pulp was gently bended in a commercial blender. The blended pulp was immediately subjected to extraction. Isolation of volatile compounds was made by the following procedure: an aliquot of blended pulp (1 kg) was diluted with distilled water (1 L) and centrifuged at 10 000 r /min for 20 min . Decanol (0.25 mg) was added as internal standard before the liquid-liquid extraction. The supernatant was continuously extracted with pentane-dichloromethane [1 : 1 (v/v)] for 8 h . The organic phase was dried over anhydrous sodium sulfate and concentrated to 0.2 mL on a Kuderna-Danish evaporator with a 15-cm Vigreux column. Extractions were made by triplicate. An HP 6890 GC with a FID, equipped with an HP-Innowax fused silica column (60 m X 0.25 mm X 0.25 μm film thickness) was employed. The column temperature was programmed as follows: 50 oC hold 4 min, to 220 oC at 4 oC/min, then hold 10 min. Nitrogen carrier gas was used at a flow rate of 1 mL/min . The injector and detector were maintained at 230 oC . Sample injection volume was 1 μL with a split ratio of 1 : 10. Linear retention indices were calculated using n-paraffin standards.6 An HP 6890 Series II equipped with a mass selective detector HP5973N and the same capillary column and temperature program as in GC-FID technique was used. Helium carrier gas was used at a flow rate of 1 mL/min . Mass spectra were recorded in the electron-impact mode at 70 eV by 1.8 scans/s . Detection was performed in the scan mode between 30 and 400 Daltons. Compounds were identified by comparing their spectra to those of authentic standards, those in NIST library or literature7-9 and also, in many cases, by comparison of their GC linear retention indices to those of standard compounds. Quantitative analysis was made by the internal standard method from the electronic integration of the FID peak areas without the use of correction factors. RESULTS AND DISCUSSION The volatile compounds of arazá fruit were obtained by liquid-liquid extraction and analyzed by GC-FID and GC-MS. A valid aroma concentrate was prepared by using an established procedure with an acceptable extraction efficiency (> 80 % recovery) and low danger of artifact formation.10-12 The concentrated extract showed aroma notes resembling the flavor of fresh fruit, described as sweet-green-fruity. Table 1 presents identified compounds with their concentrations. Quantitations were based upon GC-FID peak integration data, so accuracy is potentially limited by a number of factors, including coelution of two or more components and differences in FID response factors among compounds. The quantitative data (Table 1) shows that totally 7.9 mg of volatile compounds per kilogram of fresh fruit were obtained. In total, 27 esters, 20 terpenes, five alcohols, six carbonyls, five acids, three hydrocarbons, two lactones and two sulfur-compounds were identified. Of the 70 components identified, 53 are reported for the first time. According to class of compounds, esters dominate the volatiles profile. These compounds that constitute over 54.8 % of the total volatiles include many ethyl and hexyl esters. Of them, ethyl octanoate, ethyl dodecanoate and ethyl decanoate were found to be the major ones. In one previous result about arazá fruit, the amount of esters was low,4 whereas in the other report they were in significant amounts. These discrepancies may be related to the stage of ripeness of the fruit when sampled, different cultivars or geographical regions and the isolation method. Two identified lactones, δdecalactone and γ-dodecalactone, were reported for the first time in arazá fruit. In the terpene group, many monoterpene and sesquiterpenes were identified, with the major representatives being globulol and Table 1. Volatile constituents of arazá fruit. d n u o p m o C n o i t a c i f i t n e d I 2 I R 3 t n u o m A ) g k / g μ ( e t a t e c a l y h t E 1 C G , S M 4 2 8 3 4 l o n a h t E 1 C G , S M 8 9 8 6 6 5 α e n e n i P C G , S M 2 1 0 1 t e t a o n a t u b l y h t E 1 C G , S M 1 2 0 1 0 1 e t a o n a t u b l y h t e m 2 l y h t E 1 C G , S M 0 4 0 1 4 1 β e n e n i P C G , S M 5 1 1 1 t e n e c r y M C G , S M 0 4 1 1 t l a n e t n e p 2 l y h t e M 2 1 S M 2 5 1 1 0 6 e t a o n a x e h l y h t E 1 C G , S M 0 2 2 1 t l o 1 n e t u b 2 l y h t e M 3 1 C G , S M 1 3 2 1 5 9 2 ) E ( β e n e m i c O C G , S M 1 4 2 1 8 3 e t a t e c a l y x e H 1 C G , S M 1 6 2 1 0 5 1 e n e l o n i p r e T C G , S M 0 7 2 1 t e n o n a t u b 2 y x o r d y H 3 1 C G , S M 5 7 2 1 4 5 e t a o n a p o r p l y x e H C G , S M 1 2 3 1 4 5 e t a o n a t u b o s i l y x e H 1 C G , S M 5 2 3 1 2 3 l o n a x e H 1 C G , S M 0 4 3 1 3 1 3 l o n e x e H 3 ) Z ( C G , S M 1 6 3 1 3 2 1 e t a o n a t c o l y h t e M 1 C G , S M 8 7 3 1 5 2 e n a c e d a r t e T 1 1 C G , S M 5 9 3 1 3 4 1 e t a o n a t u b l y h t e m 2 l y x e H 1 C G , S M 8 0 4 1 3 8 1 e t a o n a t c o l y h t E C G , S M 1 2 4 1 1 3 0 1 e t a t e c a o i h t l y h t e m 2 l y h t E 1 S M 5 2 4 1 2 6 1 l a r u f r u F 2 1 C G , S M 9 3 4 1 0 0 3 α e n e a p o C C G , S M 5 7 4 1 t e n a c e d a t n e P 1 1 C G , S M 9 9 4 1 8 2 Revista CENIC Ciencias Químicas, Vol. 38, No. 3, 2007. 365 1 Reported for the first time in this fruit. 2 Identification: MS mass spectra, GC comparison of retention indices with standards. 3 Linear retention indices reported on HP-Innowax capillary column. t Represents less than 10 μg/kg . Table 1. (continued) d n u o p m o C n o i t a c i f i t n e d I 2 I R 3 t n u o m A ) g k / g μ ( e t a o n a t u b y x o r d y h 3 l y h t E C G , S M 2 0 5 1 3 4 β e n e l o b a s i B 1 C G , S M 8 0 5 1 t e t a o n a n o n l y h t E 1 C G , S M 9 1 5 1 9 2 e t a o n e t c o 2 ) E ( l y h t E 1 C G , S M 2 3 5 1 5 2 e t a o n a p o r p o i h t l y h t e m 3 l y h t E 1 C G 5 3 5 1 5 2 l a r u f r u f 2 l y h t e M 5 1 C G , S M 8 5 5 1 0 0 1 l o n a t c o l y h t e M 2 1 C G , S M 5 6 5 1 1 4 β e n e l l y h p o y r a C C G , S M 1 7 5 1 t e t a o n a x e h l y x e H 1 C G , S M 2 9 5 1 8 5 1 l a n a t n e p l y h t e m 2 y x o r d y H 3 1 S M 2 0 6 1 t e t a o n a c e d l y h t E C G , S M 1 2 6 1 6 1 5 e t a o n a x e h l y n e x e H 3 ) Z ( 1 C G , S M 8 3 6 1 5 2 e t a t e c a l y l l e n o r t i C 1 C G , S M 8 5 6 1 t α e n e l u m u H C G , S M 9 8 6 1 t d i c a c i o n a t u b l y h t e M 2 1 C G , S M 1 9 6 1 0 5 2 e n a c e d a t p e H 1 1 C G , S M 9 9 6 1 6 2 D e n e r c a m r e G C G , S M 0 1 7 1 t γ e n e l o r u u M 1 C G , S M 3 1 7 1 5 2 β e n e l a h c a m i H 1 C G , S M 6 1 7 1 5 2 e t a o n a c e d n u l y h t E 1 C G , S M 1 2 7 1 6 1 ) Z , E ( α e n e s e n r a F 1 C G , S M 2 2 7 1 0 5 e t a o n a t c o l y x e H 1 C G , S M 0 0 8 1 1 4 1 e t a o n a c e d o d l y h t E 1 C G , S M 8 1 8 1 2 9 7 d i c a c i o n a x e H C G , S M 0 3 8 1 1 9 2 e t a o n a x e h l y r u f r u F 1 C G , S M 5 4 8 1 3 3 n a r u f l y o n a x e H 2 1 C G 7 4 8 1 3 3 e t a o n a p o r p l y h t e l y n e h P 2 1 C G , S M 2 5 8 1 0 0 3 e t a t e c a l y p o r p l y n e h P 2 1 C G , S M 6 2 9 1 5 4 e t a o n a c e d a r t e t l y h t E 1 C G , S M 7 2 0 2 1 9 1 d i c a c i o n a t c O C G , S M 5 4 0 2 8 0 2 e t a o z n e b l y h t e l y n e h P 2 1 C G , S M 6 5 0 2 9 7 l o l u b o l G 1 C G , S M 1 6 0 2 6 1 2 l o n e l u h t a p S 1 C G , S M 0 7 0 2 5 2 e t a o z n e b l y n e x e H 2 ) E ( 1 C G , S M 8 7 0 2 3 3 e t a t e c a l y m a n n i C ) E ( 1 C G , S M 9 9 0 2 0 0 1 γ l o m s e d u E 1 C G , S M 5 3 1 2 t l o n i d a C T 1 C G , S M 8 3 1 2 3 8 δ e n o t c a l a c e D 1 C G , S M 5 4 1 2 5 2 l o l o r u u M T 1 C G , S M 1 5 1 2 8 5 α l o l o r u u M 1 C G , S M 5 6 1 2 0 1 d i c a c i o n a c e D 1 C G , S M 8 5 2 2 3 3 γ e n o t c a l a c e d o D 1 C G , S M 6 1 3 2 5 5 e t a o n e c e d a t c o 9 ) E ( l y h t e M 1 S M 8 3 4 2 5 8 d i c a c i o n a c e d o D 1 C G , S M 8 3 6 1 1 4 (E,Z)-α-farnesene. Germacrene D, reported in a previous study as the most abundant compound,4 was only detected in traces in the present study. Alcohols and carbonyls compounds represented 17.0 and 6.9 % of the total volatiles. Of them, ethanol, 1-hexanol and 3-methyl-2-buten1-ol were the major alcohols, whereas 2-furfural was the most abundant carbonyl compound. The presence of some acids (6.8 %) and n-paraffins (2.5 %) should not significantly contribute to fruit flavor, due to the high thresholds of these compounds.13 Two sulfur-compounds not reported previously, ethyl 2-methylthioacetate and ethyl 3-methylthiopropanoate were identified. On the other hand, methyl 3methylthipropanoate which was previously reported,5 was not found in the present study. Certainly, many of the found compounds (Table 1) should make important contributions to the overall arazá flavor, particularly the esters, with their relative low threshold.13 Many of the esters found in arazá which seems to be strong contributors to tropical fruit aromas have also been found in a variety of other tropical and subtropical fruits. Ethyl and hexyl esters similar to those found in this study have been identified as important contributors of tropical fruit flavors in papaya,14 passion fruit,15 guava,16 pineapple,17 lulo fruit,18 and mango.19