New Forests (2011) 41:89–94

DOI 10.1007/s11056-010-9212-1

Provenance variations in seed-related characters and oil

content of Calophyllum inophyllum L. in northern
Australia and Sri Lanka

Subhash Hathurusingha • Nanjappa Ashwath • David Midmore

Received: 26 August 2009 / Accepted: 22 June 2010 / Published online: 1 July 2010
Ó Springer Science+Business Media B.V. 2010

Abstract Seed morphometric characters and oil content were studied in multiple-use
plant, Calophyllum inophyllum L. of two countries, Australia (southern hemisphere) and
Sri Lanka (northern hemisphere). Seven provenances were selected which included three
from northern Australia and four from Sri Lanka. Twelve Candidate plus trees (CPTs) each
were selected from 2 to 3 different locations within each provenance based on the mor-
phometric and qualitative traits (GBH [ 100 cm). Seed collection in both hemispheres
was carried out from May to August 2008. Calophyllum inophyllum L. provenances
showed a distinct hemisphere variation in their seed-related characters and oil content. All
provenances differed significantly (P \ 0.05) with one another in seed length. Seeds from
Anuradhapura, Sri Lanka and seeds from Cardwell Australia recorded the highest (&57%)
and the lowest oil (&31%) content respectively. Strong correlations were found between
seed morphometric characters. Relatively weaker correlations were found between seed
morphometric characters and oil content. Variations in seed-related characters were largely
attributed by the provenance contribution which shows the significance of the effect of
genetic variability on above mentioned seed related characters.

Keywords Calophyllum inophyllum L.  Oil  Provenance  Seed  Variation

Introduction

Calophyllum inophyllum L. (Clusiaceae) commonly known as Alexandrian laurel, beauty

leaf or Domba (in Sri Lanka) is essentially a littoral tree of the tropics, occurring above the
high-tide mark along sea coasts of northern Australia and extending throughout Southeast
Asia and southern India. It is a medium-sized to large evergreen tree that grows up to
8–20 m in height with a broad spreading crown of irregular branches. The tree supports a
dense canopy of glossy, elliptical leaves, fragrant white flowers, and large round nuts
(Friday and Okano 2006).

S. Hathurusingha (&)  N. Ashwath  D. Midmore

Centre for Plant and Water Science, CQ University, Building 7, Rockhampton, QLD 4702, Australia
e-mail: s.hathurusingha@cqu.edu.au

123
90 New Forests (2011) 41:89–94

For centuries, Australian indigenous communities have recognised this plant as a vital
medicinal source in their day-to-day life. The tree has many economic uses. It has a high
commercial demand for seed oil from pharmaceutical companies (Agroforestry Data Base
2006). It is also valued as a high priced durable hardwood timber species (density 600–
900 kg/m3). Recently scientists have discovered anti-cancer and anti-HIV compounds from
Callophyllum inophyllum extracts (Patil et al. 1993; Taylor et al. 1994; Spino et al. 1998;
Itoigawa et al. 2001; Powar et al. 2003).
In spite of its high economic potential, scientific information relating to this species,
including seed-related traits is quite sparse. In Australia very little national attention has
been given to this species until now as it has been categorized as ‘species of least concern’
by Nature Conservation Act 2006. True economic potential of the species is yet to be
exploited.
Increasing world-wide demand for renewable energy promotes large scale commercial
plantations of potential biodiesel plants. Calophyllum inophyllum L. is an ideal source for
making biodiesel since it is non-edible, has high kernel oil (65%), fruits profusely (3,000–
10,000 seeds/tree/year), tolerates harsh environmental conditions (acidity, salinity, drought
and a range of temperature) and requires little maintenance (Azam et al. 2005; Sahoo et al.
2006; Hathurusingha and Ashwath 2007).
This species shows a wide range of geographical distribution (Friday and Okano 2006).
Geo-climatic variations may influence plastic changes in plant performance (Harper 1977).
There may be genotypic variations among provenances. Kaushik et al. (2007) and Rao
et al. (2008) have noticed such variations and strong heritability in important seed related
characters like oil content among Jatropa curcus provenances in India. Searching for
favourable variants may be useful in identifying suitable provenances and genotypes for
commercial cultivation. Quality of seeds is an important factor in plantation forestry.
Polymorphism has also been found to play a great role in seed germination, survival and
seedling growth (Pathak et al. 1980). Source variation tests are necessary to screen the
naturally available genetic variation to select the best planting material for higher pro-
ductivity (Bhat and Chauhan 2002). Above tests yield valuable information which may be
useful to commercial planters. The present investigation was envisaged to evaluate the
variations between provenances in seed parameters and oil content of Calophyllum ino-
phyllum L. in Sri Lanka (Northern Hemisphere) and in Australia (southern hemisphere).

Materials and methods

Seven provenances (Table 1) were selected which included three from northern Australia
(Yeppoon, Townsville and Cardwell) and four from Sri Lanka (Anuradhapura, Kurunegala,
Colombo and Matara). The selected provenances of each country are distinct in terms of
their climate, topography and soil conditions and are at least 100 km apart from one
another. Details on each site are presented in Table 1. Twelve Candidate plus trees (CPTs)
each were selected from 2 to 3 different locations within each provenance based on the
morphometric and qualitative traits (GBH [ 100 cm). Seed collection in both hemispheres
was carried out from May to August 2008.
Seeds of the same provenance (2–3 locations) were mixed together to form a repre-
sentative seed sample. Variability in seed parameters (length, diameter and exocarp
thickness) was investigated by measuring 30 randomly selected seed from each provenance.
Measuring was done with a digital Vernier calliper. For each provenance, kernel weight was
determined by individually weighing 30 kernels. Kernels were then oven dried for 6 days

123
New Forests (2011) 41:89–94 91

Table 1 Selected Australian and Sri Lankan provenances of Calophyllum inophyllum L.

Provenance Coordinates Altitude Mean annual Mean annual Soil types
from MSL rainfall (mm) temperature
(m) Max/Min (°C)

Sri Lankan Provenances

1. Matara 5°560 55 N, 3–60 2,150 29.3/24.1 Alluvial soils, Red-
(intermediate 80°320 34 E yellow podzolic
low) with soft and hard
laterite
2. Colombo (wet 6°540 N, 7–65 2,500 30.6/24.1 Red-yellow podzolic
low) 79°500 E with soft and hard
laterite
3. Kurunegala 7°450 N, 116 1,800 31.7/22.8 Low humic gley soils,
(intermediate 80°150 E red yellow podzoilc
low) soils with soft and
hard laterite and
regosols on red and
yellow sands
4. Anuradhapura 08°200 6000 N, 89 1,094 32.1/23.3 Reddish brown earths
(dry low) 80°220 6000 E & immature brown
loams
Australian Provenances
5. Townsville 19°130 S, 4 1,124 28.9/19.8 Clay loam or sodic
(wet tropics) 146°480 E clay soils,
calcareous dune
sands
6. Cardwell (wet 18°160 0 S, 5 2,125 28.7/18.9 Solonetz, Planosols,
tropics) 146°10 60 E calcareous dune
sands
7. Yeppoon 23°070 4200 S, 6 870.1 25.9/18.5 Rundle, Shallow
(central coast) 150°440 3400 E stony browns and
black massive
loams and sandy
clay loams

under 40°C, weighed, crushed and oil was extracted with diethyl ether (30 min-sonication at
35°C) using LABOROTA 4000 efficient rotary evaporator at 35°C. Only six provenances;
three from each country were selected for oil extraction and ten replications were used for
each provenance. Oil content was expressed as a percentage using following formula,
Weight of the oil fraction ðWoÞ  100
Oil content ð%Þ ¼ :
Weight of the dry Kernel used ðWkÞ

Statistical analysis

Data on each morphological trait investigated were subjected to analysis of variance.

Provenance variance (r2prov) and environment or error variance (r2e) were estimated, and
the proportion of r2prov to the total variance (r2T) calculated so as to compare the
magnitude of variation due to provenance (Ibrahim et al. 1997). After testing for normality
and homogeneity of error variances, differences among means were tested using the
General Linear Model of ANOVA (GLM) with GENSTAT ver.11.

123
92 New Forests (2011) 41:89–94

Results and discussion

Results of the study demonstrated significant provenance variations in seed-related traits of

C. inophyllum L. (Table 2). Hemisphere provenance variation was evident; Australian and
Sri Lankan provenances differed markedly. Several authors have reported similar findings
for other species such as; Cordia africana (Loha et al. 2006), Dalbergia sissoo (Gera et al.
2000), Faidherbia albida (Ibrahim et al. 1997), Strychnos cocculoides (Mkonda et al.
2003), and Tectona grandis (Jayasankar et al. 1999; Sivakumar et al. 2002).
Current study revealed a significant hemisphere variation in all seed-related characters
except kernel weight (Table 2). All provenances that were compared were found to be
significantly different (P \ 0.05) in their seed length. Variation in kernel weight was found
to be less pronounced compared to other seed- related characters. Kernel weights of
provenance 3 differed less significantly (P [ 0.05) with that of provenances 4, 5 and 7.
Seeds from Anuradhapura, Sri Lanka recorded highest oil content (&57%) while seeds
from Cardwell, Australia gave the lowest (&31%).
Loha et al. (2006) suggested that correlated quantitative traits are extremely important
in tree improvement as improving one character may cause simultaneous changes in other
character. Above authors have reported of a strong correlation between seed length and
seed weight. Our data (Table 3) show seed length to be strongly correlated with seed
diameter, kernel weight and exocarp thickness. Strong positive correlation was also seen
between seed diameter and exocarp thickness. However, morphological parameters were

Table 2 Provenance means for seed-related traits of C. inophyllum L.

Provenance Seed length Seed diameter Kernel weight Exocarp thickness Oil content
(mm) (mm) (g) (mm) (%)

Matara 25.61a 23.07a 3.11a 3.38b Na

Colombo 27.07b 25.06b 3.63b 3.32b 31.50a
Anuradhapura 29.42c 26.11bc 4.71c 2.93a 56.29d
Kurunegala 31.26d 26.38c 4.36c 3.34b 43.33bc
Townsville 38.72f 34.02e 5.16d 6.73d 42.97b
Cardwell 40.38g 33.79e 6.45e 5.27c 30.84a
Yeppoon 36.38e 31.53d 4.51c 5.13c 44.58c

Means that are not followed by the same letter are significantly different (at 0.05 level of probability) na not
available. All the data except kernel weights were transformed to log10 while conducting ANOVA and
means were retransformed to normal values for meaningful presentation

Table 3 Correlations between the oil content and seed morphometric traits traits of six C. inophyllum L.
provenances
Parameter Seed diameter Kernel weight Exocarp thickness Oil content

Seed length 0.98* 0.84* 0.86* -0.39

Seed diameter 0.78 0.93* -0.36
Kernel weight 0.55 0.43
Exocarp thickness -0.27

Values are Pearson product–moment correlation coefficient, r

* Correlation is significant at the 0.05 level

123
New Forests (2011) 41:89–94 93

Table 4 Estimates of provenance variables as determined from measurement of seed-related traits and oil
content of the C. inophyllum L.
Trait Overall mean Total variance (r2T) Contribution* (%)

Provenance Error
variance (r2prov) variance (r2e)

Seed length 33.77 0.176 0.0008 98.9

Seed diameter 29.42 0.132 0.0007 98.6
Exocarp thickness 4.80 0.563 0.0020 99.6
Kernel weight 4.45 28.396 0.7017 94.7
Oil content 39.86 0.298 0.0002 99.8
2 2
* Provenance contribution = (r prov)/(r T) 9 100
Yij = l (general mean) ? Pi (provenance effect) ? Bi (replicate effect) ? e9i j (random error)

not consistently correlated to oil content. These results are consistent with the finding of
Kaura et al. (1998) who observed similar correlations in Azadirachta indica (Neem).
In spite of their larger size, seeds from Australian provenances contained lower amount
of oil as compared to Sri Lankan provenances. However, seeds from Cardwell, Australia
and Colombo Sri Lanka recorded similar oil contents; 30.8 and 31.5% respectively.
Interestingly, both provenances share similar mean annual rainfalls ([2,000 mm). In
contrast, seeds from Anuradhapura, Sri Lanka and Yeppoon, Australia recorded the highest
oil contents, despite sharing the lowest mean annual rainfall in their respective countries.
This indicates that under favourable conditions (higher mean annual rainfall) C. inophyl-
lum L. seeds store a large amount of storage materials which are mostly carbohydrates
(Ibrahim et al. 1997) and in harsher conditions they produce slightly smaller oil-rich seeds
(Rose 1988). Seeds with very high oil content are sometimes dormant (Dweck and
Meadows 2002). It is an adaptation to an unfavourable environment which could reduces
the competition for available resources and improves the survival rate of germinated
seedlings.
Results (Table 4) shows that variations in seed related characters in Calophyllum ino-
phyllum were largely attributed by the provenance contribution which indicates the
influence of environmental and genetic variability of provenances on above mentioned
seed related characters (Ibrahim et al. 1997). The lowest provenance contribution was
found for kernel weight (94.7%). Ibrahim et al. (1997) have also reported similar findings
for Faidherbia albida. The variations in oil content in our study were highly influenced by
the provenance contribution (99.8%) which is in agreement with the findings of Kaushik
et al. (2007) and Rao et al. (2008).

Conclusions

Calophyllum inophyllum L. provenances have a distinct hemisphere variation in their seed

morphometric characters and oil content. Seed morphometric parameters demonstrate
strong correlations between themselves. Presence of strong provenance contribution con-
firms the significance of the combined effect of environment and genetic variability on
above mentioned seed related characters.

123
94 New Forests (2011) 41:89–94

References

Agroforestry Data Base (2006) World Agroforestry Centre PROSEA network, http://www.world
agroforestry.org/sea/Products/AFDbases/AF/asp/SpeciesInfo.asp?SpID=2. Accessed 17/08/2006
Azam MM, Waris A, Nahar NM (2005) Prospects and potential of fatty acid methyl esters of some non-
traditional seed oils for use as biodiesel in India. Biomass Bioenergy 29:293–302
Bhat GS, Chauhan PS (2002) Provenance variation in seed and seedling traits of Albizzia lebbek Benth. J
Tree Sci 21:52–57
Dweck AC, Meadows T (2002) Tamunu (Calophyllum inophyllum)-the African, Asian, Polynesian and
Pacific Panacea. Int J Cosmet Sci 24:1–8
Friday JB, Okano D (2006) Calophyllum inophyllum (kamani) species profiles for Pacific Island agrofor-
estry, Traditional Tree Initiative, Hawaii, http//.traditionaltree.org
Gera M, Gera N, Ginwal HS (2000) Seed trait variations in Dalbergia sissoo Roxb. Seed Sci Technol
28:467–475
Harper JL (1977) Population biology of plants. Academic Press, London
Hathurusingha HMSD, Ashwath N (2007) Beauty leaf, a tree with great economic potential. In: 12th
international forestry symposium, Kalutara, Sri Lanka
Ibrahim AM, Fagg CW, Harris SA (1997) Seed and seedling variation amongst provenances in Faidherbia
albida. For Ecol Manag 97:197–205
Itoigawa M, Ito C, Tan HT et al (2001) Cancer chemopreventive agents, 4-phenylcoumarins from Calo-
phyllum inophyllum. Cancer Lett 169:15–19
Jayasankar S, Babu LC, Sudhakar K, Unnithan VKG (1999) Provenance variation in seed and germination
characteristics of teak (Tectona grandis L.F.). Seed Sci Technol 27:131–139
Kaura SK, Gupta SK, Chowdhury JB (1998) Morphological and oil content variation in seeds of Aza-
dirachta indica A. Juss (Neem) from northern and western provenances of India. Department of
Genetics, CCS Haryana Agricultural University Hisar, India
Kaushik N, Kumar S, Kumar K, Beniwal RS, Roy S (2007) Genetic variability and association studies in
pod and seed traits of Pongamia pinnata (L.), Pierre in Haryana, India. Genet Resour Crop Evol
54:1827–1832
Loha A, Tigabu M, Teketay D, Lundkvist K, Fries A (2006) Provenancevariation in seed morphometric
traits, germination and seedling growth of Cordia africana Lam. New For 32:81–82
Mkonda A, Lungu S, Maghembe JA, Mafongoya PL (2003) Fruit- and seed-germinationcharacteristics of
Strychnos cocculoides an indigenous fruit tree from natural populations in Zambia. Agrofor Syst
58:25–31
Pathak SK, Gupta K, Debroy R (1980) Studies on seed polymorphism, germination and seedling growth of
Pongamia pinnata. Indian J For 2:64–67
Patil AD, Freyer AJ, Eggleston DS, Haltwanger RC, Bean MF, Tylor PB, Caranfa MJ, Breen AL, Bartus
HR, Johnson RK, Herzberg RP, Westley JW (1993) The inophyllums, novel inhibitors of HIV-1
reverse transcriptase isolated from the Malaysian tree, Calophyllum inophyllum Linn. J Med Chem
36:4131–4138
Powar KD, Joshi SP, Bhide SR, Thengane SR (2003) Pattern of anti-HIV dipyranocoumarin in callus
cultures of Calophyllum inophyllum Linn. J Biotechnol 130:346–353
Rao GR, Korwar GR, Shanker AK, Ramakrishna YA (2008) Genetic associations, variability and diversity
in seed characters, growth, reproductive phenology and yield in Jatropha curcas (L.) accessions. Trees
22:697–702
Rose IA (1988) Effects of moisture stress on the oil and protein components of soybean seeds. Aust J Agric
Res 39:163–170
Sahoo PK, Das LM, Babu MKG, Naik SN (2006) Biodiesel development from high acid value polanga seed
oil and performance evaluation in a CI engine. Fuel 86:448–454
Sivakumar V, Parthiban KT, Singh BG, Gnanambal VS, Anandalakshmi R, Geetha S (2002) Variability in
drupe characters and their relationship on seed germination in teak (Tectona grandis L.F.). Silvae
Genet 51:232–237
Spino C, Dodier M, Sotheeswaran S (1998) Anti-HIV coumarins from Calophyllum seed oil. Bioorg Med
Chem Lett 24:3475–3478
Taylor PB, Culp JS, Debouck C, Johnson RK, Patil AD, Woolf DJ, Brooks I, Herzberg RP (1994) Kinetic
and mutational analysis of human immunodeficiency virus type 1 reverse transcriptase inhibition by
inophyllums, a novel class of non-nucleoside inhibitors. J Biol Chem 269:6325–6331

123