Influence of Drying Methods on Bioactive Properties, Fatty Acids and Phenolic Compounds of Different Parts of Ripe and Unripe Avocado Fruits.

All drying processes increased oil content, antioxidant activity, total phenolic contents, and most of the phenolic compounds in the pulp, peel and seeds of both ripe fruits with varied degrees (p < 0.05). In addition, the processes reduced the oil contents, linoleic acids, 3,4-dihydroxybenzoic acid, (+)-catechin, and naringenin of the pulp, antioxidant activity of the peels and seeds, and 3,4-dihydroxybenzoic acid, (+)-catechin of the seeds and it enhanced all other parameters in the pulp, peel, and seeds of unripe fruits (p < 0.05). Comparing the phenolic profiles of avocado pulp, peels, and seeds of ripe and unripe fruits indicated that the peel and seeds are richer than the pulp and that is superior in unripe fruits than ripe ones. In addition, drying processes particularly microwave and air drying greatly enhanced the bioactive properties of ripe and unripe avocado fruits and could thus be used to elongate the shelf-life of avocado fruit products without major impact on the overall quality.


Drying of Avocado fruit parts
Avocado fruits were manually cleaned, pulp, peel and seeds were separated from the fruits and then about 100 g of each part was dried either by air drying at room temperature 24 for two week or microwave Arçelik ARMD 580, Turkey oven capable to generate at 720 W power at 2450 MHz or in an oven Nüve FN055 Ankara, Turkey, 55 L volume at 60 19 h . After drying, the fruit samples were cooled at room temperature, and then kept frozen at 25 under nitrogen in sealed bottles for further analyses.

Moisture content
The moisture contents of the samples were measured at 100 5 in an oven Nüve FN055 Ankara, Turkey according to AOAC method 9 .

Oil content
The oil content of avocado samples was determined according to AOAC 9 method. Total oil content of the samples was extracted with petroleum benzene in Soxhlet Apparatus for 5 h and the solvent was removed with a rotary vacuum evaporator at 50 .

Fatty acid composition
The sample oils were esterified according to ISO-5509 10 method. Commercial mixtures of fatty acid methyl esters were used as reference data for the relative retention times. Fatty acid methyl esters of oil samples were analyzed on gas chromatograph Shimadzu GC-2010 equipped with flame-ionization detector FID and capillary column Tecnocroma TR-CN100, 60 m 0.25 mm, film thickness: 0.20 µm . The temperature of injection block and dedector was 260 . Mobile phase was nitrogen with 1.51 mL/min flow rate. Total flow rate was 80 mL/min and split rate was also 1/40.

Preparation of the sample extracts
The extracts of avocado fruit parts were done according to Lopez-Cobo et al. 11 . Ground sample 1 g of each part was added to 20 mL of hexane and mixture of 10 mL methanol: water 80:20, v/v . The mixture was kept in ultrasonic water-bath for 15 min, followed by centrifugation at 6000 rpm for 15 min. The supernatant was removed. The steps were repeated twice and the lower phases were collected. The extract was concentrated at 37 under vacuum. The volume was made up to 10 mL with a mixture of methanol: water 50:50 and then filtered with 0.45 µm filter.

Total phenolic content
Total phenolics contents of avocado fruit parts were determined using the Folin-Ciocalteu FC reagent as applied by Yoo et al. 12 . Folin-Ciocalteu 1 mL and Na 2 CO 3 10 mL were added to extract and mixed with vortex. The deionised water was added until the final volume was 25 mL, and kept at dark for 1 h. The absorbance was measured at 750 nm in a spectrophotometer. The results are shown as mg gallic acid equivalent GAE /100 g of fresh weight.

Antioxidant activity
The free radical scavenging activity of the extracts was determined using DPPH 1,1-diphenyl-2-picrylhydrazyl according to Lee et al. 13 . The extract was mixed with 2 mL methanolic solution of DPPH. The mixture was shaken vigorously and allowed to stand at room temperature for 30 min and the absorbance was recorded at 517 nm by using a spectrophotometer. Antioxidant activity was calculated using following relation: 2.2.8 Determination of phenolic compounds A Shimadzu-HPLC equipped with a PDA detector and an Inertsil ODS-3 column was applid for the qualification and quantification of phenolic compounds were performed. The mobile phase was composed of 0.05 acetic acid A and acetonitrile B and 20 µL of the extract was injected and run at 1 mL/min at 30 for a total running time of 60 min. The peaks were measured at 280 and 330 nm using a PDA detector. The total running time per sample was 60 min.

Statistical analysis
The analyses of variance were performed using JMP version 9.0. Tukey s tests was applied to determine the significant variations among results of control, maturation and drying types p 0.05 . All analyses were carried out three times and the results are mean standard deviation MSTAT C of independent tissue values.

Results and Discussion
3.1 Effect of drying methods on moisture, oil, total phenolics and antioxidant activity The effect of drying methods on moisture, oil, total phenolics and antioxidant activity of unripe and ripe avocado Pinkerton fruit parts are shown in Table 1. Unripe and ripe avocado fruit parts pulp, peel and seed were dried by different methods air-, microwave-and oven drying . The moisture contents of fresh pulp 68.47 and 73.01 , peel 74.31 and 74.26 and seeds 57.81 and 59.11 of unripe and ripe fruits, respectively, were significantly p 0.05 reduced by all drying methods with the highest reduction being observed in oven-dried samples. Previous studies indicated that increasing the drying temperature significantly reduced the moisture contents of avocado seeds and pulp 14,15 . In peel and pulp of ripe fruits, all drying treatment significantly p 0.05 increased the oil contents compared to fresh samples. Mostert et al. 16 reported that both ripening and drying treatment enhanced the oil contents of avocado pulp. In addition, dos Santos et al. 17 stated that temperature had positive effect on the oil yield of avocado pulp. However, Moreno et al. 18 reported that drying Hass avocado at temperatures higher than 100 reduced the oil yield and attributed that to structural changes to idioblastic cells. With exception of unripe peels and seeds, drying processes increased the antioxidant activity in the pulp, peel, and seeds of ripe fruits and the pulp of unripe fruits p 0.05 with the highest activity being observed in air-dried pulp, peel and seeds of ripe fruits and conventional oven-dried pulp of unripe fruits. In addition, all drying treatments significantly enhanced the total phenolic contents of the pulp and peel of both ripe and unripe fruits compared to fresh samples. Among the treated samples, the highest total phenolic content was observed in all parts of ripe and the pulp of unripe avocado fruits dried with microwave oven and the peel and seeds unripe fruits subjected to air drying treatment. The enhancement of the antioxidant activity and total phenolic contents with drying treatments could be linked to the reduction of moisture content and consequently the concentration of the bioactive compounds. Krumreich et al. 19 studied the effect of oven drying under ventilation 40 and 60 and vacuum oven 60 of avocado pulp on the quality of extracted oil and they observed that drying the pulp at 60 resulted in high bioactive compounds and antioxidant activity of the oil. They attribute that to the inactivation of polyphenol degrading enzyme namely polyphenol oxidase at 60 , thereby preserving the phenolic compounds in the oil 19 . Figueroa et al. 20 reported that the highest total phenolic content of avocado peel was achieved with a conventional oven drying at 85 . Furthermore, Saavedra et al. 5 reported that increasing drying temperature from 45 to 60 increased the antioxidant capacity values of avocado seeds whereas it reduced that of the peels. Apparently, the oil contents, antioxidant activity, and total phenolic contents of avocado are greatly influenced by many factors such as ripening stage, fruit parts, drying process conditions type, temperature, and duration as well as other factors such as genotypes, growing conditions, and postharvest processing.

Effect of drying methods on fatty acids composition
Fatty acid composition of different fruit part oils of avocado Pinkerton cultivar extracted by Soxhlet system dried at the different drying systems air-, microwave-or oven-drying are presented in Table 2. Generally, the fatty acid profile is differed among the tested fruit parts of ripe and unripe fruits subjected to different drying treatments p 0.05 . With exceptions of oleic in unripe fruits and palmitic in ripe ones, drying treatments significantly p 0.05 enhanced the values of all fatty acids of the pulp with the highest values being observed in pulp dried using conventional oven. In the peel samples of unripe fruits, drying treatment significantly p 0.05 increased fatty acids,  Linolelaidic *mean±standard deviation; **Values in each row with different letters are significantly different (p < 0.05);***nonidentified except linolenic, with the enhancement of the majority of fatty acids was found in air-dried peels. Palmitic and arachidonic acid amounts of avocado fruit parts peel, pulp and seed varied depending on drying type and maturity p 0.05 . In addition, a partial decrease in some fatty acids was observed with drying. While oleic acid amounts of raw and ripe fruit pulps did not change much depending on the drying type, fluctuations were observed in the oleic acid amounts of shell and seed oils p 0.05 . Interestingly, microwave-and air-drying treatments significantly enhanced the level of linoleic acid in the seeds of both ripe and unripe fruits. In addition, all drying treatments increased the amounts of linoleic acid in the pulp of both ripe and unripe fruits, and in the peel and seeds of ripe fruits. The increase in linoleic acid following drying treatment is likely due to thermal degradation of cell matrix and decomposition of conjugated lipids and phospholipids and thereby releasing more free form of linoleic acid. Previously, various studies indicated that oleic, palmitic, linoleic are the major fatty acids in the pulp and seed oils of various avocado cultivars 1, 8, 21 24 . Krumreich et al. 19 studied the effect of drying on the fatty acids of oil extracted for the pulp of mature avocado fruits and observed that oven drying treatment at 60 influenced positively the fatty acid compositions of the oil. However, Santana et al. 8 reported that the fatty acid composition of avocado pulp oil was not affected by drying process; however, it was affected marginally with the ripening stage and peeling process. The variation in the fatty acid composition of avocado oils between these studies could be attributed to the differences in the, genetic makeup, the fruits parts pulp, peel, and seeds , maturity stage, postharvest practices, environmental conditions, drying conditions, and oil extraction methods.

Effect of drying methods on phenolic compounds
The profiles of phenolic compounds of the pulp, peel, and seeds of ripe and unripe avocado fruits as affected by different drying methods are shown in Tables 3, 4 and 5. The major phenolic compounds in the pulp of both ripe and unripe avocado fruits were -catechin, 1,2-dihydroxybenzene, and 3,4-dihydroxybenzoic acid Table 3 . With few exceptions, the quantities of all phenolic compounds were higher in the pulp of unripe fruits compared to that in ripe ones. While the -catechin contents of fresh, air-dried, microwave and oven-dried unripe avocado fruit peels were 203.04 mg / 100 g, 208.87 mg / 100 g, 36.71 mg / 100 g and 99.82 mg / 100 g, respectively, the -catechin contents of fresh, air-dried, microwave and ovendried ripe avocado fruit peels were 111.70 mg / 100 g, 79.60 mg / 100 g, 212.93 mg / 100 g and 118.01 mg / 100 g, respectively. There was an increase in the amount of catechin of ripe avocado peel dried in microwave and oven. Probably, this may be due to the decrease in the amount of -catechin in the pulp during maturation as a result of the biochemical reaction. In previous studies, the quantities of phenolic compounds of avocado pulp was differed between avocado cultivars from different countries and growing seasons 8,11,28,29 and the composition was different than that reported herein for Pinkerton cultivar. Santana et al. 8 reported that fruit ripening and unpeeling process enhanced the phenolic compounds of avocado oils. Regardless of drying process, the peel of both ripe and unripe avocado fruits contained considerable quantities of phenolic compounds Table 4 .
-Catechin and 1,2-dihydroxybenzene were found to be the dominant phenolic compounds in both unripe and ripe fruit peel extract.
-Catechin content of unripe fruit peel extract was significantly high when the peel was dried by air 208.87 mg/100 g while that of ripe peel extract was significantly high when the peel was dried by microwave 212.93 mg/100 g . A high level of quercetin was observed in unripe and ripe peel 1174.08 mg/100 g and 773.18 mg/100 g, respectively dried by microwave. Among the drying methods used, microwave drying greatly improved the quantities of phenolic compounds in the peels of both ripe and unripe avocados. Similarly, Figueroa et al. 20 reported that increasing drying temperature from 45 to 85 significantly increased the phenolic compounds in avocado peels. The phenolic compounds of unripe and ripe fruit seed extracts dried by different drying methods are shown in Table 5. In the seeds obtained from unripe fruits, air-drying treatment greatly enhanced the quantities of most phenolic compounds of the seeds compared to other drying treatments, except gallic and caffeic acids which were improved more by conventional oven drying and syringic acid by microwave drying. Interestingly, a higher level of quercetin 2369.94 mg/100 g content of unripe seeds was observed in air-dried seeds compared to than in fresh seeds 137.66 mg/ 100 g suggesting that such drying treatment enhanced its contents by more than 17 folds. In the seeds of ripe avocados, air drying treatment improved the contents of 3,4-dihydroxybenzoic, trans-ferulic, and trans-cinnamic acids, kaempferol and isorhamnetin and microwave drying improved that of apigenin 7 glucoside and resveratrol, whereas conventional drying increased the levels of all other phenolic compounds. Comparing the phenolic profiles of avocado pulp, peels, and seeds of ripe and unripe fruits indicated that the peel and seeds are richer than the pulp and that is superior in unripe fruits than ripe ones. The present results highlight the suitability of using the avocado by product, namely, peel and seed, of unripe fruits as sources of phenolic compounds after suitable drying processes. Saavedra et al. 5 reported that avocado peels and seeds contain large variety and high amounts of phenolic compounds and drying process improved the quantities of phenolic compounds. Variations in phenolic compounds between fruit parts might be associated with maturity stage but also may be due to environmental con-   ditions, postharvest management, geographical conditions and genetic background 25 27, 30 . Optimum harvest maturity and processes are the important factors determining the quality of avocado fruit.

Conclusion
This study represent one of the few studies on the combined impacts of avocado maturity stage and drying methods on the oil yield bioactive properties and fatty acid composition of the pulp, peel, and seeds. The results indicate that the avocado pulp, peel, and seeds are important sources of bioactive compounds and essential fatty acids at the ripe and unripe stages. With few exceptions, drying of the pulp, peel, and seeds significantly improved oil yield, bioactive properties and fatty acid composition of these parts of both ripe and unripe avocado fruits. The most suitable drying method for preserving the analyzed quality parameters was microwave-drying followed by air and oven. The present results highlight the suitability of using the avocado fruit pulp, peel and seed, of both ripe and unripe fruits as sources of bioactive compounds after suitable drying process.