Project Title: CHEMICAL CONSTITUENTS OF PITTOSPORUM RESINEFERUM

HEMSL

Researchers:
Agnes B. Alimboyoguen
Faculty Researcher
Research Center, Cvsu
alim.agnes.b@gmail.com

Evangeline M. Fidel
Research Assistant
Research Center, Cvsu

Budget: 200,000 pesos

Source of Funds: Cvsu Grant

Abstract

The purpose of this study was to determine the chemical constituents present in
Pittosporum resiniferum Hemsl. The results demonstrated that the air-dried dichloromethane
extract of the leaves of P. resiniferum contained a mixture of uvaol (1) and erythrodiol (2). The
structures of (1) and (2) were identified by comparison of their 1H-NMR with the literature data.

Keywords: Pittosporu resiniferum, Pittosporaceae, uvaol, erythrodiol

Introduction

Pittosporum resiniferum Hemsl, also known as Petroleum Nut or “hanga” in Tagalog is an

aromatic tree that belongs to the family Pittosporaceae. It is an endemic in the Philippines and can
be found only in high mountain ridges and forested areas in the Bicol and Cordillera region (Bakker
and Van Steenis, 1985). Traditionally, some parts of the tree are being used to cure different
diseases. The infusion of the fruit is used as remedy for intestinal and stomach pain while decoction
of leaves are employed for colds and cough. Studies also showed that the oleoresin from the plant
can cure leprosy and other skin diseases (Altschul, 1973) and muscular pains (Sescon et al., 2002).
Earlier reports also mentioned that the plant has used as a source of biofuel primarily because of
its high heptane, myrcene, dihydroterpene and α-pinene components (Cortez et al., 2016). GC/MS
analysis of the oil of ripe P. resiniferus fruit yielded n-heptane (5%), n-nonane (7%), three isomeric
monoterpenes (85%) and six minor sesquiterpenes (6%). Two major monoterpenes were isolated
by preparative GC and identified as pinene (38%) and myrcene (40%), while catalytic
hydrogenation of the oil gave pinane and2,6- dimethyloctane (Nemethy and Calvin, 2016).

Previous study also showed that the leaves of P. pentandrum afforded eudesm-11-en-4-α-
O-β-D-3-senecioyloxy-6-deoxyglucopyranoside, eudesm-11-en-4-α-O-β-D-3-tigloyloxy-6-deoxy
glucopyranoside, and botulin (Ragasa et al., 1997). Here, the study presents the isolation of a
mixture of uvaol (1) and erythrodiol (2). This research is part of the studies on the chemical
constituents of trees belonging to the genus Pittosporum and also, the first report on the isolation
of 1 and 2 from the plant.

Figure 1. Chemical Constituents of Pittosporum resiniferum Hemsl:

Uvaol (1) and Erythrodiol (2).

Methodology

Sample Collection and Identification of Pittosporum resiniferum Hemsl.

The leaves of Pittosporum resiniferum Hemsl. were collected from the Deparment of
Environment and Natural Resources Compound, La Trinidad, Benguet, Philippines in May 2015.
It was authenticated at the Jose Vera Santos Herbarium, Institute of Biology, University of the
Philippines, Diliman, Quezon City.

Isolation and Purification

The air-dried leaves (0.9 kg) of P. resiniferum were cut into small pieces, ground in a
blender, soaked in CH2Cl2 for 3 days and then filtered. The solvent was evaporated under vacuum
to afford a crude extract (30.5g) which was chromatographed using increasing proportions of
acetone in CH2Cl2 at 10% increment by volume as eluents. The 40% and 50% acetone in CH2Cl2
fractions were combined and rechromatographed (7 ×) using 15% EtOAc in petroleum ether to
afford a mixture of 1 and 2 (3 mg) after washing with petroleum ether.

Fractions were monitored by TLC which was performed with plastic-backed plate coated
with silica gel F254. The spots were visualized by spraying with vanillin-H2SO4 followed by
warming.
 The isolation and purification scheme of the crude extract is shown below.

Crude Extract
30.5 g

Chromatographced
Me2CO in CH2Cl2 (10% increment)

40% 50%
Fraction Fraction
combined

Rechromatographed in CH2Cl2

Fractions
D-H

Rechromatographed
(7x) in 15% EtOAc
in pet. ether

Compound PR7
(3 mg)

Scheme 1. Isolation and Purification of Compound PR7 Pittosporum resineferum

 Structure Elucidation

The TLC pure isolate was sent to the University of Newcastle-Australia, Callaghan,
Australia for NMR analyses. NMR spectra was recorded in CDCl3 on a JEOL JNM ECP-400
spectrometer (Tokyo, Japan) at 400 MHz for 1H.

Results and Discussions

Identification of Compound PR7 in Pittosporum resineferum

Physical Properties
The mixture PR7 isolated from CH2Cl2 extract of the air-dried leaves of P. resiniferum was
obtained as white solid. It produced a blue-violet spot TLC spot when warmed with vanillin-
sulfuric acid visualizing agent. It has an Rf value of 0.48 when developed with 20% ethyl acetate
in petroleum ether.

Functionalization of Protons by 1H-NMR Spectroscopy

As shown in Figures 2 - 4, the 1H-NMR indicated the presence of a mixture of two related
compounds as deduced from integrals and disparity of some resonance signals. In the deshielded
region, the doubling resonance of olefinic protons at δ 5.17 and δ 5.12 and 2 oxygenated protons
at δ 3.52 and δ 3.18 were observed. While in the shielded portion of the spectra displayed nine
methyl singlets at δ 1.23, 1.14, 1.08, 0.98, 0.97, 0.93, 0.92, 0.91 and δ 0.77 in which three of these
singlets gave repeating resonances at δ 0.98, 0.91 and δ 0.77 and two methyl doublets at δ 0.86
and δ 0.78.
Putting the spectra of PR7 and comparing the obtained signals with the spectra and
resonances of uvaol and erythrodiol in the literature disclosed total match among them as shown
in Table 1.
 Table 1. Comparison of the 400 MHz 1H-NMR Spectral Data of PR7
with Uvaol (1) and Erythrodiol (2) in CDCl3

1 1
Type of Proton H Shift, δ H Shift, δ
PR7 Uvaol (1)
Methyl singlet 1.08 1.09
Methyl singlet 0.98 0.99
Methyl singlet 0.93 0.94
Methyl singlet 0.91 0.90
Methyl singlet 0.77 0.72
Methyl doublet 0.86 0.93
Methyl doublet 0.78 0.80
Olefinic (t) 5.17 5.17
Oxygenated (t) 3.52 3.54
Oxygenated 3.18 3.20
1
H Shift, δ
Erythrodiol (2)
Methyl singlet 1.23 1.22
Methyl singlet 1.14 1.13
Methyl singlet 0.98 0.96
Methyl singlet 0.92 0.92
Methyl singlet 0.93 0.89
Methyl singlet 0.91 0.91
Methyl singlet 0.77 0.76
Olefinic (t) 5.12 517
Oxygenated (t) 3.52 3.54
Oxygenated 3.18 3.20

Previous studies have shown that uvaol (1) and erythrodiol (2) which belongs to ursane
and oleanane classes, respectively, exhibited antibacterial, antifungal, anti-inflammatory and anti-
malarial (Fanuck et al., 2016).
Figure 2. The Complete 400 Mhz 1H-NMR Spectrum of PR7 in CDCl3
Figure 3. The Expanded 400 Mhz 1H-NMR Spectrum of PR7 in CDCl3
Figure 4. The Expanded 400 Mhz 1H-NMR Spectrum of AB43 in CDCl3
Conclusion and Recommendation

The air-dried leaves of dichloromethane extract of Pittosporum resiniferum yielded

mixture of bioactive compounds, uvaol (1) and erythrodiol (2). Since the plant and its constituents
have wide range of medicinal applications, it is suggested that further investigation on the other
components and their biological activities be conducted.

References

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Downloaded from udr.slu.edu.ph:8080/jspui/bitstream/123456789/1213/1/6_Cortez.docx

Esther K. Nemethy and Melvin Calvin.(1982). Terpenes from Pittosporaceae. Downloaded from
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