Lipid Catabolism : Complex Lipid Degradation Lipid Catabolism Colloquium Organized and Edited by J. Harwood (School of Biosciences, Cardiff University) and P. J. Quinn (Division of Life Sciences, King's College London). 14th International Symposium on Plant Lipids held at Cardiff University, 23-28 July 2000.
Screening for lipase activity in the oil palm R. Sambanthamurthi*', N. Rajanaidu and S. Hasnah Parman Advanced Biotechnology and Breeding Centre, Malaysian Palm Oil Board, P.O. Box 10620, 50720 Kuala Lurnpur, Malaysia
Abstract
duction cost. Lipase (triacylglycerol acylhydrolase, EC 3 . 1 . 1 . 3 ) is the first enzyme involved in the degradation of stored triacylglycerols. T h e increase in NEFA levels in palm oil is attributed to the action of lipase. However, evidence as to the nature of the lipase has been contradictory. 00[l] and Tombs and Stubbs [2] reported the absence of endogenous lipase in the oil palm mesocarp. T h e latter workers attributed NEFA increases in the oil palm mesocarp to microbial activity. Hartley [3], however, suggested the presence of a very active endogenous lipase in the oil palm. This fact was firmly established by three reports [4-61. Sambanthamurthi et al. [5] also established that the lipase was activated at low temperature. This study aimed to investigate the variation in lipase activity among different clones with the view to selecting for lipase genotypes.
T h e oil palm mesocarp contains an endogenous lipase which is strongly activated at low temperature. Lipase activity is thus very conveniently assayed by prior exposure of the fruits to low temperature. More than 100 oil palm samples from the germplasm collection of the Palm Oil Research Institute of Malaysia (now known as the Malaysian Palm Oil Board) were screened for nonesterified fatty acid activity using both the lowtemperature activation assay and a radioactivity assay. T h e results showed good correlation between assay procedures. T h e different samples had a very wide range of lipase activity. Elaeis oleifera samples had significantly lower lipase activity compared with E . guineensis (var. tenera) samples. Even within E . guineensis (var. tenera), there was a wide range of activity. T h e results confirmed that lipase activity is genotype-dependent. Selection for lipase genotypes is thus possible and this will have obvious commercial value.
Materials and methods Plant material Fresh fruit bunches of oil palm (Elaeis guineensis Jacq. var. tenera, and E . oleifera) were obtained from the germplasm collection at the Palm Oil Research Institute of Malaysia (now known as the Malaysian Palm Oil Board). Fruits at 20 weeks after anthesis were used.
Introduction Palm oil is the world's second most important vegetable oil after soya bean. T h e level of nonesterified fatty acids (NEFAs) is a very important determinant of oil quality. Oxidation of NEFAs results in rancidity and impairment of oil quality. NEFAs can be removed during the refining process, but this adds significantly to the pro-
In vitro (radioactive) lipase assay T h e enzyme was assayed essentially as described by Sambanthamurthi et al. [6] using dry mesocarp powder.
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NEFA determination NEFAs were determined following overnight exposure of fruits to 5 "C as described by Sambanthamurthi et al. [ 5 ] .
Key word: Eloeis gurneensrs. Abbreviations used: NEFA, non-esterified fatty acid. 'To whom correspondence should be addressed (e-mail
[email protected]).
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method is more convenient and avoids the use of radioactive material.
Table I NEFA levels in different oil palm samples
Lipase activity in various oil palm samples from the germplasm collection T h e Malaysian Palm Oil Board has the world's largest oil palm germplasm collection. Fruits from various palms from this collection were screened for lipase activity. A sample of the results is shown in Table 1. T h e fruits analysed belonged to two species, E . guineensis and E . oleifera. E. guineensis fruits generally had higher lipase activities compared with those of E . oleifera. E . oleifera fruits also had much lower oil contents. T h e E . guineensis fruits had higher lipase activity, and there was also a wide variation in activity between fruits of E . guineensis. T h e fruits from the Cameroonian collection generally had lower lipase activities and hence lower NEFA contents compared with the fruits from Tanzania and Angola. Highest NEFA content was observed in fruits from the Angolan collection. Some of the clones from the Cameroonian and Tanzanian collections had very low NEFA levels, e.g. palms 0.219/P179, and 0.218/P1076. These results confirmed that lipase activity is genotype-dependent. It would be very useful to select for these lower lipase varieties. However, further investigation needs to be carried out to determine the oil contents of these genotypes to ensure that selection for low lipase does not result in decreased oil yield. There may also be commercial value in selecting for high lipase activity for specific purposes, e.g. for the production of NEFAs for oleochemical uses.
NEFA (%)
Palm
Species
Origin
0.2 I9/P I 676 0.2 I 9/P I 79 0.2 I9/P I757 0.2 I9/P I 680 0.2 I9/P833 0.2 I9/Pw I 867 0.2 I 8/P I064 0.2 I 8/P I076
E. guineensis E. guineensis E. guineensis E. guineensis E. guineensis E. guineensis E. guineensis E. guineensis
Cameroon Cameroon Cameroon Cameroon Cameroon Cameroon Cameroon Cameroon
26.6f I .O 3.2f0.5 I2.9f I .2 26.9f0.7 8.2f0.8 I2.6k I .O 29.0f0.7 3.4f0.3
0.256/P2202 0.256/P2 I 66 0.256/P2 I I 3 0.256/P2093 0.256/P2 I 15 0.256/P2094 0.256/P2 I I6 0.256/P2246
E. guineensis E. guineensis E. guineensis E. guineensis E. guineensis E. guineensis E. guineensis E. guineensis
Tanzania Tanzania Tanzania Tanzania Tanzania Tanzania Tanzania Tanzania
8.5f0.6 3 I .8f0.5 22.3f0.3 36. I f0.7 32.6f0.8 I3.9f0.3 40.4f0.4 I0.3k0.6
0.3 I3/P24 0.3 3/P I02 0.3 3/P25 0.3 3/P22 0.3 3/P99 0.3 2/P I060 0.3 2/P994
E. guineensis E. guineensis E. guineensis E. guineensis E. guineensis E. guineensis E. guineensis
Angola Angola Angola Angola Angola Angola Angola
0. I 7/P46 0. I77/P66 0. I77/P28 0. I88/P59 0. I88/P45 0. I88/P38 0.2 I I /P442 0.2 I I /P533
E. oleifera E. oleifera E. oleifera E. oleifera E. oleifera E. oleifera E. oleifera E. oleifera
Surinam Surinam Surinam Brazil Brazil Brazil Honduras Honduras
66.8f0.3 10.I I f0.5 I4.5f0.6 54.4f0.5 9.7f0.3 43.4f0.5 43.0f0.8 3.2f0.2 5.3fO. I 3.5f0.3 3. I f0.5 6.2fO. I 5.5f0.5 2.9f0. I 5.8f I .O
W e thank the Director-General of the Malaysian Palm Oil Board for permission to publish these results.
References
Results and discussion
I 00,K. C. ( I98 I ) Oleagineux 3 6 , 4 I 3 4 I6 2 Tombs, M. P. and Stubbs, J. B. ( I 982) J. Sci. Food Agric. 33, 892-897 3 Hartley. C. W. S. ( I 988) The Oil Palm, Longman, London 4 Hendenon. J. and Osbome, D. J. ( I 999) Phytochemistry 30, 1073- I078 5 Sambanthamurthi, R., Chong, C. L.. 00, K. C., Yeo, K. H. and Rajan, P. ( I 99 I) J. Exp. Bot. 42, I 199- I205 6 Sambanthamurthi, R., 00,K. C. and Parman, S. H. ( I 995) Plant Physiol. Biochem. 33, 353-359
Correlation between in vitro lipase assay and in vivo cold-induced lipase activity A random sample of oil palm fruits was assayed for lipase activity by both the in vitro radioactive assay and titration for NEFAs of the oil extracted from fruits exposed to 5 "C. A strong positive correlation was obtained between results from the two techniques. T h e low-temperature activation assay was subsequently used for screening a germplasm collection for lipase activity as this
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Received 23 June 2000
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