ABSTRACT
Baby banana (Musa acuminata AA Simmonds cv. Bocadillo) is cultivated in Colombia and other tropical regions. The hyperpigmentation phenomenon is the occurrence of green veins through the peels which causes that fruits are rejected in the international marketing. The aim of this study was the analysis of the hyperpigmentation phenomenon in relation to the production of volatile compounds in Baby banana peel and pulp. Volatiles profiles from pulp and peel of hyperpigmentated and control ripe samples were analyzed by GC-FID and GC-MS. The results suggest that there are no qualitative and quantitative differences in the volatile compounds between control and hyperpigmentated fruits.
Introduction
The Baby banana fruit (Musa acuminata AA Simmonds cv. Bocadillo) is cultivated commercially in Colombia, Ecuador, Costa Rica, Venezuela and Kenya, but Colombia is the main country in Latin-America which has developed the marketing for exporting fruit to Europe and USA. The Baby banana is known in English as “Lady banana”, “Finger banana”, “Date banana”, “Fig banana” or “Sucrier”, depending on the region where it is cultivated.
Since climatic changes influence rainfalls, temperature and water availability, the traders of this tropical fruit in Colombia confirmed the presence of an irregular ripening in several fruit peels during the post-harvest stage that it was called hyperpigmentation. This is expressed by the occurrence of visible small green bands in the peels during the ripening which means that chlorophyll degradation was abnormal in comparison with control bananas. Therefore, those Baby banana fruits with hyperpigmentation were not suitable for export because they did not meet the overall standards of quality and both farmers and traders were affected economically (Castro-Benitez, Citation2015).
The knowledge on chlorophyll degradation has grown considerably nowadays due to relevant reports about chlorophyll catabolites in higher plants and fruits (Kräutler et al., Citation2012), but the identification of enzymes and intermediates involved in the metabolic pathways of volatile compounds is not enough to have a deep knowledge about possible relations among both biosynthetic process (Osorio et al., Citation2010). No previous information about the influence of the hyperpigmentation phenomenon on the volatile compounds profiles of Baby banana has been reported. Volatile compounds generation may exhibit a normal pattern even if the fruits suffer the hyperpigmentation phenomenon.
Fruit aroma is recognized as an important indicator that reflects the quality of fruit flavor and determines its acceptance by the consumer. Therefore, the objective of this study was the analysis of the hyperpigmentation phenomenon in relation to the production of volatile compounds in Baby banana peel and pulp.
Materials and Methods
Samples
Two batches of banana fruits, hyperpigmentated and control (1 kg each one), selected with a similar ripening degree (color of the peel green – trace of yellow), were harvested in a commercial plantation located in Antioquia, Colombia from the 2016 crop season and immediately transported to the laboratory by Nativa Produce SAS Co. (). Three batches of five ripe fruit pulps or peels in each were cut and rapidly homogenized in a rotor-stator blender Ultra-Turrax T18 Basic (Carl Zeiss, Oberkochen, Germany), at 25,000 min−1 during 5 min. These batches were used for subsequent analyses.
Figure 1. Examples of hyperpigmentated (left side) and control (right side) Baby banana fruits.
Chemicals and Reagents
Pure reference standards for GC analyses were acquired from Sigma–Aldrich (St. Louis, MO), except 2-pentyl acetate, 2-pentyl 3-methylbutanoate, and 2-heptyl butanoate, which were synthesised by esterification in the laboratory (Becker et al., Citation1973). Hexyl (E)-3-hexenoate was generously supplied by Dallant S.A. (Barcelona, Spain). A n-alkane solution (C8–C32) was purchased from Sigma–Aldrich (St. Louis, MO). To this solution, a mixture of C5–C7 n-alkanes was added. Anhydrous sodium sulfate, sodium chloride, diethyl ether and methyl nonanoate were acquired from Merck (Darmstadt, Germany).
Standard Chemical Analysis
Soluble solids, total acidity (as malic acid) and pH value were performed on the fruit pulp according to standard methods (AOAC, Citation2006).
Solid-phase Microextraction Analysis (SPME)
Volatile compounds from the headspace of peel or pulp homogenates were isolated using 1-cm length 50/30 μm DVB/CAR/PDMS fibers (Supelco Park, Bellefonte, PA, USA). This fiber was selected for the analysis because it provided the most authentic odor for banana (Pino and Febles, Citation2013; Pino et al., Citation2017). Fibers were conditioned prior to each sampling step and immediately used after conditioning. SPME extraction conditions were: 3 g of stirred homogenate, 3 ml of Milli-Q water and 1 g NaCl contained in a 15 ml-vial sealed with a PTFE-lined screw cap, and 40°C for 30 min. Extraction were made by triplicate for each batch.
Isolation of Volatile Compounds by Solvent-assisted Flavour Evaporation (SAFE)
Homogenized pulp (200 g) was mixed with 300 ml of Milli-Q water (previously saturated with NaCl) and the mixture was extracted with recently redistilled diethyl ether (2 × 100 ml) on an autoshaker at room temperature for 10 min. After centrifugation (6000 min−1; 5 min), the supernatants were combined and the residue was discarded. The organic layer was isolated in a separatory funnel, and the aqueous layer was discarded. The organic phase was subjected to SAFE (Engel et al., Citation1999) at 40 °C, and the organic phase was dried over anhydrous Na2SO4 and concentrated to 0.6 ml using a Vigreux column (15 cm x 1 cm i.d.) and then to 0.2 ml with a gentle nitrogen stream. The odor of the isolate was evaluated to ensure that it closely matched the profile of the ripe Baby banana. Extraction were made by triplicate for each batch.
GC-FID and GC-MS Analysis
A Konik 4000A gas chromatograph (Konik, Barcelona) with a flame ionization detector (FID) was used. The capillary columns were DB-5 ms and DB-Wax (each, 30 m x 0.25 mm x 0.25 μm; J & W Scientific, Folsom, CA, USA). The oven temperature was held at 50 °C for 2 min and then raised to 250 °C at 4 °C min−1 and held for 10 min. Hydrogen was used as a carrier gas at a flow rate was 1 ml min−1. Injector and detector temperatures were 250 °C. For the SAFE extracts 1 μl was injected in 1:10 split mode and for SPME extracts splitless mode (2 min) was applied. The retention times of a series of n-alkanes (C5–C32) was used to calculate the retention indices for all identified compounds and for reference standards. Quantitative analyses of the compounds were performed by relative area percents. All data were obtained by triplicate.
GC-MS analyses were performed on a Hewlett–Packard 6890N series II (Agilent, Palo Alto, CA) gas chromatograph with the same fused capillary columns, temperature program and Helium carrier gas flow rate as in GC-FID. EIMS, electron energy, 70 eV; ion source and connecting parts temperature, 250 °C. The acquisition was performed in scanning mode (mass range m/z 35–400 u). Compounds were preliminarily identified by use of NIST 05, Wiley 6, NBS 75k, Adams 2001, and in-house Flavorlib libraries, and then the identities of most were confirmed by comparison of their linear retention indices with those of reference standards or with published data (Adams, Citation2007).
Statistical Analysis
The data from triplicate analyses from each batch were processed for statistical analysis by the t-Student’test.
Results and Discussion
General Characteristics of the Fruits
The general characteristics of the pulp from hyperpigmentated and control fruits are shown in . No significant differences were found between both types of fruit pulps.
Table 1. General characteristics of pulp from hyperpigmentated and control fruits.
Analysis of Fruit Peel and Pulp by SPME
The analysis of volatile compounds of peel and pulp from hyperpigmentated and control fruits showed 51 compounds, distributed mainly as esters, alcohols, ketones and aldehydes ().
Table 2. Volatile compound (area %) of peel and pulp from hyperpigmentated and control fruits isolated by SPME.
Major compounds in peels and pulps were 3-methyl butanoate, 3-methylbutyl 3-methylbutanoate, (Z)-4-hepten-2-yl butanoate, 3-methylbutyl acetate and 2-pentyl butanoate. No significant differences were found between both types of peels and pulps.
Analysis of the Pulp by SAFE
SAFE extraction allows the isolation of volatile compounds from the matrix diluted in water using high vacuum distillation at low temperature, thus avoiding potential flavor modification or artefacts due to the formation of volatile compounds at elevated temperatures (Majcher and Jelen, Citation2009). The extraction takes a long time, but reproducibility is good and compared with other isolation methods, like SPME, SAFE is more effective in extracting less volatile and polar components. Thus, it was selected to provide a representative analysis of the overall flavor of Baby banana. Each isolation method has advantages and drawbacks depending on the food matrix studied, so it is essential to recover an isolate that is as representative as possible of the product. For this purpose the representativeness of the odor of the samples was checked by sensory analysis and the isolates closely matched the profile of the ripe Baby banana.
The results reported in show that the use of SAFE method allowed the identification of 120 volatile compounds extracted from the pulp of hyperpigmentated and control fruits. The composition of volatile compounds included 61 esters, 19 alcohols, 13 ketones, 5 acids, 3 aldehydes, and 19 miscellaneous compounds. Esters seemed to be the dominant family in terms of total amount and number in Baby banana and they possess fruity-banana notes (Pino and Febles, Citation2013; Pino et al., Citation2017; Thaiphanit and Anprung, Citation2010). Among the identified compounds, 3-methylbutyl butanoate, 3-methylbutyl 3-methylbutanoate and 3-methylbutyl acetate were the major constituents.
Table 3. Volatile compound (area %) of pulp from hyperpigmentated and control fruits isolated by SAFE.
From a quantitative point of view, no significant differences were found in the relative quantities presented in depending on the fruits were hyperpigmentated or not. This means that the phenomenon has no effect on the flavor of the fruits because the sensory key compounds are in similar amounts in both control and hyperpigmentated fruits.
Conclusions
The results suggest that there are not qualitative and quantitative differences in the composition of volatile compounds between control and hyperpigmentated Baby banana fruits. The hyperpigmentation phenomenon is clearly related only to natural pigments on the peel, but it has no effect on the flavor of the fruit. Hence, these results should be taking in mind for future examination of the actual quality standards that reject hyperpigmentated fruits.
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