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2011, Science Hu, ISSN: 2153-649X, doi:10.5251/ajsir.2011.

High accumulation of lipids during off-tree ripening and senescence in Jatrophacurcas Linn Luanti accession kernels

Jupikely James Silip, 2Armansyah H. Tambunan, 2Erliza Hambali, 2Sutrisno, and 2 Memen Surahman

School of Sustainable Agriculture, Universiti Malaysia Sabah, Locked Beg No.2073, 88999 Kota Kinabalu, Sabah, Malaysia, 2 Bogor Agricultural University, Bogor 16680, Indonesia.
ABSTRACT This study was conducted to firstly determine changes in extracted oil yield, soluble solid concentration and free fatty acids over time of harvested physiologically mature green Jatrophacurcas Linn Luanti accession fruits. Secondly, to understand identified fruit bunch harvesting indicators by examining the ripening percentage during storage based on the identified characters of fruits bunches at harvest. The results showed a significantly high oil yield extracted from off-tree fruits ripened and senesced kernels compared to kernels from on-tree ripened and senesced fruits. Extracted oil yield from off-tree ripened and senesced fruits were about 59 and 63% compared to only about 49 and 48% for on-tree ripened and senesced fruits respectively. Fruit bunches with less than 10% of their fruits yellow in color is recommended as the best harvesting indicator. The results of this study demonstrate promising harvesting and postharvest handling practices to increase commercialization viability of this non-edible biodiesel feedstock. Keywords: Jatropha, oil content, off-tree ripening, off-tree senescence

INTRODUCTION Indeterminate reproductive characteristics has been identified as the major factor leading to poor harvesting of this crop and as a result, today this nonedible biodiesel feedstock is labeled as blunder crop and also considered a none viable industrial crop (Silip et al., 2010a; Achten et al., 2008; Biswas et al., 2006 and Heller, 1996). Breeding programs are considered long term solutions to develop Jatropha varieties with determinate reproductive characters and it is thought that a near-term solution to this problem could be through good harvesting and postharvest handling practices. Understanding the physicochemical changes during the life of the fruit will provide basic data to justify recommendations for new harvesting and postharvest handling practices. Recommendations on when to harvest Jatropha fruits is still a difficult issue. Most of the available information relates that harvesting time is best when the fruits produce high oil content (Heller, 1996; Hambali, 2007; Santoso 2008 and Annarao et al., 2008). Since harvesting of individual fruits is noted to be laborious and time consuming, harvesting fruit bunches was recommended and contrary to recommendations on fruit bunches characteristics which has been reported (Hambali et al., 2007). A good recommendation on harvesting of fruit bunches

will increase harvest quantity and directly reduce harvesting visits but there will be loses in harvest quantity due to rejection of unripe fruits. Our earlier study indicated that unripe or physiologically mature green Jatropha fruits could be ripened off the tree and will have similar or much higher oil content compared with those that ripen on the tree (Silip et al., 2009 and Silip et al., 2010a). No information was adduced on the changes of oil content during the life of Jatropah fruits off the tree. Therefore the main objective of this study was to determine these changes. MATERIALS AND METHODS Plant Materials. The source of Jatropha fruits for this study was from local Luanti accession, harvested from a Jatropha pilot project conducted by the Institute of Agro-Biotechnology Malaysia at Luanti Baru village, Keningau, Sabah, Malaysia. Fruits of five different maturity stages were collected for oil extraction yield analysis. The stages included immature, mature, yellow ripe and black wet senescence. The selected healthy, uniform sized fruits were harvested manually from the plot for the various maturity stages and damaged seeds were discarded. Immediately after harvest, the samples were brought to the School of Sustainable Agriculture

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Laboratory, Universiti Malaysia Sabah for on-tree sample or fresh sample analysis. The harvested physiologically mature fruits were left to ripen and senesce at room temperature (27+2oC) for three days before analysis as off-tree samples. Plant materials for fruit bunches indicator was collected randomly from the plot based on their characters. Oil Extraction. The soxhlet technique was used for chemical extraction with hexane solvent (boiling point of 40 60 oC). The extracted lipid was obtained by filtrating the solvent using a rotary evaporator apparatus at 40 oC followed by heating in an oven at 105 oC for three hours to evaporate any remaining solvent and water. Samples for oil extraction were prepared according to Silip et al. (2010b). Measurement of Extracted Oil Yield. The weight of oil extracted from 10 g of kernel powders was measured to determine the lipid content. Results were expressed as the percentage of oil in the dry matter of kernel powder. Measurement of Soluble Solid Concentration. The soluble solid concentration of coats and kernels was determined using a hand refractometer (Model N1, Atago) based on the modified method of Dadzie and Orchard (1997). A total of 30 g jatropha tissue in 90 ml distilled water was blended using a kitchen blender for 2 min and filtered through a filtering paper. A drop of the filtrate was placed on the prism glass of the refractometer. The refractometer was pointed towards a light source and readings of SSC (%) were recorded. The recorded value was multiplied by a factor of three (because the initial tissue sample was diluted three times with distilled water) and the reading was corrected to a standard temperature of 20 oC according to Bourne (1982). Measurement of Free Fatty Acid. Acid value of kernel oil was determined according to American Organization of Chemical Scientist Official Method Cd 3a-63 and the percentage of free fatty acid was calculated using oleic acid as a factor. Measurement of Ripening Percentage. Jatropha of maturity stage of 1 to 5 was first established using the modified guava colour index by Silip (2003) where stage 1 = green, stage 2 = more green than yellow, stage 3 = more yellow than green, stage 4 = yellow or fully ripe and stage 5 = senescence wet black. The percentage of ripening uniformity was assessed by calculating the percentage of similar maturity stage divided with the total number of fruits observed multiplied by hundred.

Experimental Design and Statistical Analysis. The experimental design of this study was a completely randomized design with a fixed identified variable and was replicated accordingly with measurable variables. The data was analyzed using one way ANOVA. The differences between means were calculated from the standard error of measurements. RESULTS AND DISCUSSION Changes on the extracted oil yield. The results showed that the percentage of extracted oil yield (d.b) in the dried kernels from off-tree ripened and senesced fruits increased significantly from about 52% in mature green to about 59 and 63% in the ripe and senesced fruits (Figure 1). The extracted oil yield in the kernels from on-tree ripened and senesced fruits did not change with extracted oil yield (d.b) of only about 50%. These results show that significant accumulation of lipid occurred during ripening off the tree compared to ripening on the tree. The increase in extracted oil in this study indicated direct triple advantages of harvesting physiologically mature green fruits. This harvesting and postharvest handling practice will increase harvestable fruits, reduce harvesting visits and increase extractable oil.

Fig 1. Extracted oil yield (%)d.b by chemical extraction technique from on and off-tree fruits dried kernels at different maturity stages. Vertical bars indicate the standard error of measurements (n=18).

Accumulation of lipid in oilseed during maturation is a normal biochemical occurrence. Multiple


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interconnected pathways leading to TGA biosynthesis have been described in maturing oilseeds (Bates et al., 2009). The only enzymatic reaction committed to TGA biosynthesis was 1,2-sndiacylglycerol acyltransferase (DGAT) (Cao and Hung 1986) and ripening and senescence off the tree could create favourable conditions for natural occurring DGATs or mutagenized versions of enzymes. Changes in Soluble Solid Concentration. Sugars and amino acids are known to be important precursors of fatty acid and TAG synthesis (Baud and Lepiniec 2010). In our study, it was found that the sugar in the kernels of off-tree ripened and senesced fruits reduce during ripening but increase during senescence (Figure 2). However, the sugar in the kernels of on-tree ripened and senesced fruits showed a decrease from mature stage to ripening and senescence. The results of this study indicate that sugar was converted to lipids during ripening as shown by the decrease from mature to ripe stage. Significant decrease in sugar from mature to ripe kernels from off-tree ripe fruits was expected because the fruit was no longer attached to the parent tree and as such nutrient loss on the fruit was no longer replaced from their mother plant.

However, increase in sugar content after ripening during senescence was contrary to the fundamental theory of oil synthesis because the extracted oil from this maturity stage in this study was high. This result indicates that the seed could be at the final stage of oil synthesis and massive conversion of biochemical requirements for seed germination has just started. According to Kornberg and Beevers, (1957), production of sugar as fuel for seed growth and development was primarily from oxidation of reserved lipid in castor bean.

Fig 3. Changes in the percent free fatty acid (FFA) of fruits during the life on the tree. Vertical bars indicate standard error of measurement (n=10).

High soluble solid concentration in the seeds from fruits ripened off the tree could also indicate that the seeds are ready to germinate. However, the lipid oxidation to sugar in this study might not yet reach a point where oil extracted yield is affected. It is important to note that we observed the occurrence of seed germination due to prolonged fruit senescence off the tree. The wet condition resulting from senesced fruits coats might have provided favorable conditions for seed germination.
Fig 2.Changes on soluble solid concentration (%) during ripening and senescence on and off the tree of Jatropha kernels. Vertical bars indicate the standard error of measurements (n=30).

Significant changes in the sugar in kernels of off-tree fruits during ripening could be the reason for the high extracted quantity of oil in this study and this result corroborates the fundamental theory of oil synthesis.

Changes in the Free Fatty Acid Content. Free fatty acids (FFA) content in biodiesel feedstock is impotent because its content will determine transesterification processes (Goodrum, 2002). High FFA is reported to reduce yield of biodiesel products due to soap formation. However, an alternative process such as a two-step process was investigated for feedstocks having high FFA content (Veljkovic et al., 2006). In this study we found that the percentage of FFA was


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less than 5%. However, these were significant differences in FFA content among the maturity types which implied shortening oil storage shelf life. Fruit Bunch Harvesting Indicator. The potential benefit of harvesting the so called physiologically mature green Jatropha fruit in this study is dependent on the availability of fruit bunch harvesting indicators. The results from our study showed that fruit bunches

with at least 10% of their fruits in the yellow stage could be the minimum bunch harvesting indicator (Table 1). Fruit bunches more than 10% yellow are better because after two days in storage all fruits in the group start to turn yellow or ripe. From the results of this study, it is recommended not to harvest fruit bunches which are 100% green as not all of the fruits will ripen off the tree after five days of storage.

Table 1. Changes on ripening percentage of harvested fruits for five different fruit bunch characters (100% MG, 10% yellow, 25% yellow, 50% yellow and 70% yellow fruits) during storage. Characters of Harvested Fruit Bunches Fruits 100% 10% 25% 50% 70% Days of Storage Group MG* Yellow Yellow Yellow Yellow Green 100.0 50.6 41.0 21.1 8.5 MGY** 0.0 33.5 29.2 35.2 34.0 MYG*** 0.0 13.3 16.8 23.8 33.0 0 Yellow 0.0 2.5 13.0 19.9 23.9 Black 0.0 0.0 0.0 0.0 0.0 Green 63.3 8.2 1.2 1.2 1.6 MGY 28.3 23.4 14.9 14.8 5.3 MYG 6.6 36.1 46.6 32.8 12.2 1 Yellow 1.8 32.3 32.3 40.6 68.6 Black 0.0 0.0 4.3 9.0 1.6 Green 44.0 1.3 0.0 0.8 0.5 MGY 4.8 12.0 1.9 0.0 0.5 MYG 16.9 15.8 13.7 5.5 1.6 2 Yellow 31.9 74.7 70.2 77.0 63.8 Black 0.0 2.5 14.9 20.7 27.7 Green 41.6 0.0 0.0 0.0 0.0 MGY 0.6 1.3 0.0 0.0 0.0 MYG 6.6 5.7 2.5 1.2 1.1 3 Yellow 45.2 91.8 74.5 66.4 39.4 Black 3.0 6.3 26.1 32.4 58.5 Green 31.3 0.0 0.0 0.0 0.0 MGY 3.6 0.0 0.0 0.0 0.0 MYG 2.4 1.3 0.0 0.8 0.5 4 Yellow 19.3 37.3 16.1 14.8 5.3 Black 36.7 62.0 77.6 88.3 94.1 Green 20.5 0.0 0.0 0.0 0.0 MGY 13.3 0.0 0.0 0.0 0.0 MYG 3.6 0.6 0.0 0.0 0.0 5 Yellow 9.6 12.0 2.5 2.3 2.1 Black 50.0 90.5 97.5 97.7 98.4 *MG (mature green), **MGY (more green than yellow), ***MYG (more yellow than green)

Ripening failure in harvested Jatropha fruits from fruit bunches which are of full size and 100% green could indicate potential loss in implementing recommended harvesting of the so called physiologically mature green fruits. The reasons of ripening failure could be due to the presence of immature fruits in the harvested bunches. Immature fruits were expected to

be unripe as its development and maturation has not yet completed. CONCLUSION AND RECOMMENDATION The results of this study highlight a direct benefit of harvesting physiologically mature green fruits and the harvesting indicators of fruit bunch. The harvested fruits showed significantly high extracted oil yield


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when allowed to ripen and or senesce off the tree. It is recommended to harvest fruit bunches which could also increase harvestable fruits and indirectly reduce harvesting visits and cost. ACKNOWLEDGMENT The authors acknowledge the Institute of AgroBiotechnology Malaysia, Ministry of Science, Technology and Innovation for the financial support. REFERENCES
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