Characterization of the oils from the pulp and seeds of avocado ( cultivar : Fuerte ) fruits By

La semilla y pulpa del aguacate (Persea americana Mill), cultivar Fuerte, cultivado en la región del nordeste de Brasil fueron analizados para determinar su composición centesimal. El contenido de lípidos en pulpa y semillas, en la base húmeda fueron 15.39% y 1.87%, respectivamente. Los aceites extraídos de la pulpa y de las semillas por hexano representan similares índices de refracción (1.4608 y 1.4592), densidad específica (0.9272 y 0.9300) e índice de peróxidos (1.40 y 1.37) pero diferentes índices de acidez (2.45 y 4.12), iodo (77.6 y 69.4) y saponificación (178.3 y 231.6), respectivamente. El análisis de cromatografía gaseosa revela la presencia de 22 y 27 ácidos grasos en aceite de pulpa y de semillas, respectivamente. El ácido palmítico fue el principal ácido saturado, presente en concentraciones de 21,3% en pulpa y 20,8% en aceites de semillas. La principal diferencia entre estos aceites fue su contenido de ácidos grasos mono-insaturados (C18:1 -64.3% en aceite de pulpa y 15.4% de semillas) y poli-insaturado (C18:2 y C18:3 -9.14% y 0.46% respectivamente, en aceite de pulpa y 34.39% y 5.81% respectivamente, en aceite de semillas).


INTRODUCTION
The avocado fruit belongs to the family of Lauraceae which includes approximately 150 species, majority of which are habitat of tropical America.The fruit distinguishes itself from other fruits due to its high lipid content and is consumed as food, and as ingredient in the formulations of pharmaceutical and cosmetic products.The oil content of the fruit depends upon its ecological origin and on the cultivar, as for example, in Guatemalan and Mexican cultivars, the oil content varies from 10 to 13% and from 15 to 25%, respectively (Biale and Young 1971) while in the fruits from Carrebean, a low fat (2.5 to 5%) has been reported (Hatton et al. 1964).According to Soares et al. (1992) adulteration of olive oil with avocado oil to a level of 35% is undetectable on the basis of flavor as the avocado oil possesses similar characteristics to that of olive oil (Gutfinger & Letan 1974;  Freitas et al. 1993).Though some work has been published on the physico-chemical properties (Tango et  al. 1972), and the fatty acids composition (Mazliak 1971;  Tango et al. 1972; Itoh et al. 1975, Martinez et al. 1988,  Southwell et al. 1990,) of the pulp oil obtained from some important avocado cultivars, and of the seed oil (Mazliak 1971), no comparative study on the characterization of the pulp and seed oils of the fruits of the same avocado cultivar is so far available.Furthermore, the fatty acid composition of the avocado oil, besides the cultivar and stage of ripening, depends upon the geographical growth location (Ahmad and Barmore, 1980).The present work was therefore undertaken to compare the physical and physicochemical properties of the oils extracted from the pulp, and seed of the avocado (cultivar: Fuerte), fruits, as well as to compare their fatty acid composition.

EXPERIMENTAL
Preparation of the pulp and seed samples of avocado fruits: Avocado fruits (Lauraceae spp.), cultivar Fuerte were obtained from a farm located in the vicinity of the city of João Pessoa in the state of «Paraíba» in Brazil.The fruits, were stored at room temperature (approx.28 o C) till they become ripe.

Grasas y Aceites
The pulp and seeds were removed manually from the fruit and the pulp was homogenised in a domestic blender.The seeds were cut into small pieces and triturated in a mill.The pulp and seed samples were filled in polyethylene bags and stored in a refrigerator.
Proximate analysis of pulp and seeds: Moisture, protein, lipid, ash and, crude fibre contents were determined following the standard AOAC (1990) methods.Total carbohydrate content was calculated from the difference.Ten different samples were analysed in duplicate.
Extraction of the oil: The homogenized avocado pulp and triturated seeds were dried in an oven for 24 hours at 50 o C. The dried mass was triturated again in a mill and passed through the sieves of 40 mesh sizes.The oil was obtained by extraction of the dried powders with solvent hexane.Three lots for each oil were prepared.
Physical and physicochemical properties of oil: The specific gravity and refractive index were determined at room temperature (28 o C) using specific density bottle and refractometer, respectively.For determination of acid, peroxide, iodine and saponification values, standard AOAC (1990) methods were used.Three different samples of oil were analysed in duplicate.
Fatty acid composítion of seed oil: Fatty acids were transformed to their methyl esters (FAME) following the method of Hartman and Lago (1973) and were determined by using a gas chromatograph HP 5890 Series III (Hewlett Packard) equipped with a flame ionisation detector.1.5 µl of the FAME sample was injected and GC separation was carried out on BP-INNOwax capillary column (Hewlett Packard; 30m length, 0.25 mm i.d. and 0.25 µm film thickness).The carrier gas (helium) head pressure was maintained at 11.5 psi and the column flow rate was 1ml/min.The oven temperature was held initially at 120 o C for 1 min, later increased at 8 o C/min to 210 o C and then maintained at 210 o C for 45 min.The temperatures of the injection port and of detector were 250 o C and 280 o C, respectively.FAME were positively identified by matching their retention time data and mass spectra with those of the authentic standards obtained from various firms (Sigma; Nu-Chek-Prep, USA) which were also run under identical analytical conditions using high resolution GC-MS system (GCQ of Finnigan Mat).

RESULTS AND DISCUSSION
The proximate analysis of the avocado pulp and seeds is presented in Table I.Besides moisture, the lipid was the principal constituent of the pulp present in concentrations of nearly 15.4%.For cultivar Fuerte, Tango et al. (1972) reported a higher lipid content (22.1 %), while Freitas et al. (1993 ) reported a recovery of 20% lipid from the pulp of the same cultivar.Lipid concentrations ranging from 15 to 30% in Fuerte, Bacon and Hass cultivars were also reported by Biale and Young (1971).In comparison with the pulp, the seeds contained only 1.87% lipids.The protein content of the pulp and seeds were 1.01 and 1.87% respectively.For California and Florida fruits, the protein content ranged from 1.21 to 2.26% as reported by Hall et al. (1955) and Slater et al. (1975).Other constituents such as ash, fibre and carbohydrates were present in smaller quantities in pulp than in the seeds.
Table II shows the data on some physical and physico-chemical properties of pulp and seed oils extracted with hexane.The refractive index, specific gravity and peroxide values of the pulp and seed oils were very close to each other.These indices/values for the pulp oil were similar to the ones reported by Soares et al (1991).However, there was a considerable difference in the saponification values of pulp and seed oils, being 178.3 and 231.6, respectively.Tango et al. (1972) reported a range of 167.6 to 190.1 for saponification value.The iodine values of the pulp (77.6) and seed (69.4) oils differed very little from the reported values of 76.5 to 99.7 for the pulp oil of different avocado cultivars (Tango et al. 1972).A high iodine value (133) for the pulp oils of Bacon, Fuerte and Hass cultivars was reported by Martinez et al. (1988).Fatty acid composition of seed oil: The fatty acid composition of the pulp and seed oil is presented in Table III.In earlier reports (Mazliak 1971), eight fatty acids were reported in the pulp oil of cultivar Fuerte, while SouthweIl et al. (1990) reported the presence of only six fatty acids.However in this study, we could identify as many as 22 fatty acids in the pulp oil.A large majority of these fatty acids in the pulp oil has not been reported earlier.Only C 16:0 , C 16:1 , C 18:0 , C 18:1 , C 18:2 fatty acids (Martinez et al. 1988; Soares et al. 1991) together with C 18:3 (Frega et al. 1990Southwell et al. 1990) and C 14:0 , C 20:0 in trace amounts (Mazliak, 1971) were reported in the pulp oil of different avocado cultivars.In all the cultivars C 18:1 (oleic acid) was aIways the principal fatty acid followed by C 16:0 (palmitic) and C 18:2 (linoleic).Other fatty acids reported to be present in trace concentrations were C 14:0 (myristic) C 18:0 (stearic), C 18:3 (linolenic) and C 20:0 (arachidic) acids (Gutfinger and Letan, 1974, Itoh et al. 1975, Mazliak 1971, and Tango et al. 1972).Similar to these reports we also observed that the major fatty acid in the pulp oil was C 18:1 followed by C 16:0 and C 18:2 however, the concentrations of these fatty acids in the pulp oil were different from the cited reports.This quantitative difference is justifiable in view of the fruits from different geographical origin and other factors such as maturity and harvest practices.According to the Mazliak (1971) the concentration of the fatty acids C 18:1 , C 16:0 and C 18:2 in the Fuerte mesocarp oil was 67-72, 13-16.7 and 10.4-11.4%, respectively while Martinez et al. (1988) reported a range of 60-65, 15-19 and 11-12%, respectively for the same fatty acids in the oil from Becon, Fuerte, and Hass cultivars.Besides, they also reported the presence of C 16:1 (palmitoleic acid) in the concentration of 7.5 to 8.6%  against the value of 3-5. 1 % reported by Mazliak (1971).However, a higher concentration (72.2%) of C 18:1 fatty acid in avocado pulp oil was reported by SouthweIl et al. (1990).In our study, the principal fatty acids and their concentrations in the pulp oil were:C 18:1 -64.43%,C 16:0 -21.31%,C 18:2 -9.15% and C 16:1 -2.39% (Table III).In addition to these, C 6:0 , C 8:0 , C 13:0 , C 14:1 , C 15:1 , C 17:1 , C: 18:0 , C 18:3 , C 20:0 , C 20:1 , C 22:1 and C 24:0 were present in concentrations varying from 0.06 to 0.76%, and C 7:0 , C 9:0 , C 14:0 , C 15:0 , C 17:0 , C 22:0 in trace quantities (less than 0.06% of the total fatty acids).Saturated fatty acids constituted about 22.93% of the total fatty acids of the pulp oil while in the seed oil was about 32.45%.All saturated fatty acids from C 6 to C 24 except for C 21 and C 23 were identified in the seed oil.The C 16 fatty acid (20.85% of total fatty acids) was also the principal saturated fatty acid of the seed oil followed by fatty acids C 15 , C 17 , C 24 , C 18 and C 22 which were present in concentrations of 2.33, 1.72, 1.68, 1.18 and 1.11% of total fatty acids, respectively.Other saturated fatty acids were present in concentrations ranging from 0.16% (C 13 ) to 0.80% (C 6 ) while C 10 , C 11 , and C 20 fatty acids were present in trace quantities.
Avocado seed oil contained high concentrations of C 18:2 (38.89%) and C 18:3 (6.57%) fatty acids.The concentration of these fatty acids in the pulp oil was relatively low (9.14% and 0.46%, respectively).A favourable C 18:2 /C 18:3 fatty acid ratio of the avocado seed oil could possibly be responsible for reducing the triglycerides and HDL in blood plasma as was observed by Werman et al. (1991) in a nutritional study with rats.In our study, the unsaturated to saturated fatty acids ratios were 3.36 and 2.07 and polyunsaturated to saturated fatty acids were 0.42 and 1.44, respectively for the pulp and seed oils.However, for the pulp oil of the same cultivar, Slater et al. (1975) reported higher values for these ratios (7.4 and 0.73, respectively).In relation to oleic-Iinoleic acid ratio, a parameter responsible for the shelf life of the oils, the avocado pulp oil (7.4) had a much higher ratio than the seed oil (0.447).
To detect adulteration of avocado oil with avocado seed oil, Werman et al. (1996) have reported a thin layer chromatographic method.Alternatively based on these findings, we also suggest that the concentrations of C 18:2 and C 18:3 fatty acids and their ratio (about 6 in seed oil and 18 in the pulp oil) could possibly be used for determining the authenticity of avocado oils of Fuerte cultivar.