REVIEW OF RELATED LITERATURE

Phenolic compounds

Phenolic compounds are ubiquitous in plants. This compound is composed of an Commented [P1]: They are

aromatic ring with one or more hydroxyl groups. The antioxidant activity of phenolic

compounds depends on the structure, in particular the number and positions of the

hydroxyl groups and the nature of substitutions on the aromatic rings. Fruits, vegetables

and beverages are the major sources of phenolic compounds in the human diet. The food Commented [P2]: delete

and agricultural products processing industries generate substantial quantities of

phenolics-rich by-products, which could be valuable natural sources of antioxidants.

Some of these by-products have been the subject of investigations and have proven to be

effective sources of phenolic antioxidants. When tested in edible oils, and in fish, meat

and poultry products, phenolic-rich extracts have shown antioxidant activities comparable

to that of synthetic antioxidants (Balasundram et al., 2006).

These compounds were derived from the pentose phosphate, shikimate, and

phenylpropanoid pathways in plants (Randhir, Lin, & Shetty, 2004). Phenolic compounds

are essential for their physiological and morphological roles in plant. These compounds

are responsible for the reproduction and growth of the plant. Phenolics’ physiological

properties were: anti-allergenic, anti-artherogenic, anti-inflammatory, anti-microbial, Commented [P3]: Some of the physiological properties of
phenolics are ...

antioxidant, anti-thrombotic, cardioprotective and vasodilatory effects (Balasundrum et

al., 2006).
Biological Activity

The biological activity of compounds of plant extracts or pure substances isolated

from living organisms is identified through different bioassays. Small scale in vitro

biological test systems are one of the key features in the medication discovery process

that provide results with enough replicates for the activity of large quantity of samples.

This mentioned process is called the bioactivity screening with assays based on cultured

cells, isolated enzymes or cloned receptors (Houghton, 2000).

Usage of cultured cells in a model system results to measurement of different

effects. In cytotoxicity, the amount of the plant extract needed to kill cells was

determined or the defensive impact of the extracts against cytotoxic agent was measured.

Test for measurement of cell viability such as the mitochondrial activity based test, MTT Commented [P4]: Tests

(Mossman, 1983) or sulphorhodamine B test that determines the total protein (Skehan et Commented [P5]: ,such as the mitochondrial activity-based
test, MTT, or sulphorhodamine B test that determines the total
protein,
al., 1990) were used in determining anticancer agents.

The defensive impact of the substances were investigated through test of Commented [P6]: was

antioxidant activity. In this test, cells were exposed to oxidizing solutions such as

hydrogen peroxide. The survival rates of the extract without the oxidizing agent is Commented [P7]: are

compared to that of the extract with the oxidizing agent (Yamasaki et al., 1994).

Another activity that can be determined is the enzyme activity. Enzyme activity

measured in a compound can either be the activity of the enzyme inside the cell or the

enzyme activity that leaked into to medium from the cell. In a study of cholinee sterase,

an enzyme found in the brain, a bioassay which measure the inhibition of the activity of
this enzyme was used to find agents which are useful for disorders such as Alzheimer’s

disease (Perry et al., 1996).

To discover different bioactive compounds, a variety of screening models are used.

Some of the screening models used are animal models, serum pharmacology models, Commented [P8]: them

tissue models, cell models, receptor/ enzyme models, biochromatographic models, and

genomic models. A more detailed type of screening models were shown in Figure 1 Commented [P9]: is

(Wang 2011).

Commented [P10]: masyadong maliit

Figure 1. The tree view of screening models (Wang,2011).

Since some animals can mimic a number of the physiological and clinical features

of humans with disease, animal models is the most classic. Screening which uses this Commented [P11]: are

model benefits from the research of efficacy, side-effect and toxicity of medicines in

whole [27]. Screening using serum pharmacology model is a straightforward approach

and helpful in proving true positive compounds. In serum pharmacology, through blood

concentrations and urinary excretion rates after consumption of the extract mixtures the
action of the bioactive compounds were characterized [28]. Compared to animal models, Commented [P12]: is

tissue models requires less amount of sample, reduced labor intensity, improved target Commented [P13]: require

activity at various pathological tissues and organs, and lowered interference. In tissue

models, tissue and organs were isolated and used to analyze the activity of the Commented [P14]: are

compounds in vitro (Wang 2011). Another in vitro model is the cell model. Cell culture

systems mimic the physiopathological state of various diseases and provide useful

information on the good predictability of bioactive components in vivo, as well as on

permeability and toxicity profiles [29].
One of the main targets of drug action is the receptor or enzyme. Because of its
Commented [P15]: ). Another in vitro model is that based on
cell culture systems, which mimic the physiopathological state of
various diseases and provide useful information on the good
predictability of bioactive components in vivo, as well as on
permeability and toxicity profiles

high sensitivity and specificity. Receptor/enzyme models are broadly used for screening

of susceptible drug from natural products. Another screening model are the Commented [P16]: Ha? Hahahaha
Commented [P17]: is
biochromatographic models. In biochromatography, the interactions between the Commented [P18]: model

bioactive components and immobilized proteins, enzymes, receptors, cell membranes and

biomimetic membranes coupled with conventional chromatography were utilized. Some

classification of the biochromatographic models are molecular chromatography, cell Commented [P19]: classifications

membrane chromatography, immobilized liposome chromatography, immobilized

artificial membrane chromatography and microdialysis chromatography (Wang 2011).

Genomic screening from natural products provide new opportunities for the treatment of Commented [P20]: provides

significant and largely unmet human diseases [50-53]. In this screening, the target genes

of a disease is explored (Wang, 2011). Commented [P21]: are

Caimito (Chrysophyllum cainito)

Chrysophyllum cainito is under Chrysophyllum genus which used as folk medicine

for the pronounced bioactivity present in the bark, leaves and fruit of the tree (Meira et Commented [P22]: Chrysophyllum cainito . which is under the
Chrysophyllum genus, is used as folk medicine for the pronounced
bioactivity present in the bark, leaves and fruit of the tree
al., 2014). This plant is highly cultivated in Southeast Asia, Caribbean, West tropical

Africa, Zanzibar, Brazil and the warmer parts of India (Shailajan&Gurjar, 2016) out.pdf. Commented [P23]: ?

C. Cainito is a tropical tree of family Sapotaceae with fleshy fruit with soft endocarp

(Luo et al., 2002) Oranusi2015. This has different names such as caimito, caimito, star Commented [P24]: ?
Commented [P25]: Difference? haha
apple, golden leaf tree and milk fruit (National Research Council, 2008).

The tree is hermaphrotic in nature with round, purple-skinned fruit that is often

green around the calyx with a star pattern in the pulp. Sometimes there is a greenish –

white or yellow variety of the fruit (National Research Council, 2008). The skin is rich in

latex; though the skin and the rind are not edible. The seed is flat, hard and light brown

in color (Einbond et al., 2004). The fruits are delicious as a fresh dessert fruit; it is sweet

and best served chilled.

The fruits, bark, and leaves of C. caimito were usually used as an antitussive, Commented [P26]: are

astringent, antioxidant agent. Traditionally, C. caimito were used as treatment for Commented [P27]: is

laryngitis with inflammations, pneumonia, and diabetes mellitus. In addition, this was Commented [P28]: it
Commented [P29]: is
also used to treat diarrhea, fever, and venereal disease (Ningsih et al., 2016).

Bioctivities of Chrysophyllumcainito

Caimito is used for the treatment for some inflammatory disease (Meiraet al.,

2014). From the study of Luo et al. (2002), nine polyphenolic antioxidants were isolated
from the ethyl acetate extract of Caimito fruit. The antioxidants were (+)-catechin, (-)-

epicatechin, (+)-gallocatechin, (-)-epigallocatechin, quercetin, quercitrin, isoquercitrin,

myricitrin, and gallic acid with emphasis on the antioxidant property of quercetin. C.

caimito leaves also contain triterpene antioxidants such as β-amyrin acetate and gentistic

acid (Ningsih, 2016). In 2010, Bastos et al. have determined the anti-helminthic activity

of the ethanolic extract of this plant.

Meira et al. (2014) evaluated the anti-hypersensitive and anti-inflammatory activity

of the crude methanolic extract, fractions, and two isolated triterpenes gathered from the

Caimito leaves. Mice were used for the in vitro experiments of this study. Inflammatory

pain were induced intraperitoneally with the crude methanolic extracts and isolated Commented [P30]: was

triterpenes, identified as 3β-Lup-20(29)-en-3-yl acetate (1) and Lup-20(29)-en-3β-O-

hexanoate (2), before injecting the λ-carrageenan. Using electronic anethesiometer, the

hypersensitivity were assessed. Based from the result the chloroform fraction greater Commented [P31]: was
Commented [P32]: results,
affect and interfere with the carrageenan induced hypersensitivity compared to crude Commented [P33]: fraction has a greater effect and
interference on
methanolic extract and ethyl acetate fraction. Anti-inflammatory activity of the crude

methanolic extract and chloroform fraction were also evaluated using the paw-oedema Commented [P34]: was

induced by carrageenan to mice. Results shows that only the crude methanolic extracts Commented [P35]: show

reduced the paw oedema. Data obtained from this experiment shows that the crude

extracts, fraction, pure compounds isolated from Caimito leaves have anti-hypersensitive

properties against inflammatory pain in mice (Meira et al., 2014).

Moo-Huchin et al. (2014) identified the antioxidant compounds and their

antioxidant activity and phenolic content in the peel of Caimito (Chrysophyllum caimito

L.). Maximum extraction of total soluble phenols and flavonoids was accomplished using
80% methanol. DPPH (2,2´-diphenyl-1-picrylhydrazyl) assay and ABTS (2,2´-Azinobis-

3-ethylbenzotiazoline-6-sulfonic acid) assay showed that freeze-dried peel of caimito

contents were vitamin C, anthocyanins, phenolic compounds, flavonoids and carotenoid

which exhibit good antioxidant activity.

Antioxidant activity of caimito leaves extract were also determined via DPPH Commented [P36]: was

assay, total phenolic content, and total flavonoid content. According to Ningsih et al.

(2016), solvent selection was an important factor in obtaining leaf extract with high level

of total phenol content, total flavonoid content, and antioxidant activity.

Lanzones (Lansium domesticum)

Lansium domesticum, commonly known as lanzones, belongs to the family

Meliaceae. This single-trunked tree stands as high as 30 m. This plant is local to

Southeast Asia particularly in the Philippines. Fruit of this plant is edible while the peels

were used as arrow poison and to repel mosquitos and the seeds were used as febrifuge Commented [P37]: as mosquito repellent, and...

and vermifuge. The bark of this tree was used as treatment for dysentery and malaria

while the extracts of the leaves can be used as anti-inflammation for the eye (Marfori et Commented [P38]: anti-inflammatory medication for the eye

al., 2014).

Compounds that were found in the fruit peels were onoceroidslansic acid and

lansiosides A, B and C. Meanwhile, the seeds contain dukunolides A-E, which are

responsible for its bitter principles. Additionally, they also contain domesticulides A-E, Commented [P39]: ?

which are identifiend as antimalarian tetranortriterpenoids. The twigs contain

onoceranoid-type triterpenoids which generates moderate activity versus the Gram-

positive bacteria. Lansiolic acid and lansionic acid were also reported to be isolated in

this plant (Tanaka et al., 2002).

Bioactivities of Lanzones

In the study of the crude extract of fruit peel from L. domesticum, the extract have

found to have inhibitory activity against Staphylococcus aureus. According to Marfori et Commented [P40]: was found

al. (2014), new triterpenoid which is lansioside D is the major antimicrobial compound in Commented [P41]: the new triterpenoid lansioside D

the methanol extract of the fruit peel. Lansioside D is reportedly exhibit the same Commented [P42]: was recorded to

biological activites as Lansiosides A-C, which are responsible for controlling the

hormones for baldness, acne and prostate hypertrophy.

Seed extract of L. domesticum can be used as antibacterial drug against

Staphylococcus aureus and Escherichia coli infections in gastrointestinal infections,

urinary tract infections and diarrhea (Alfonso et al., 2017). Borges et al. (2012) and

Borges et al. (2014) cited that some phenolic acids (gallic and ferulic acids) and GHP

(allylisothiocyanate and 2-phenylethilisotiocyanate) reduced the potential of adhesion to

plasmon surface of some pathogenic bacteria, including E. coli and S. aureus. In other

work performed by Luis et al. (2014), gallic acid was also able to influence the adhesion Commented [P43]: In another work performed by

properties of methicillin-resistant S. aureus (MRSA) to PS. Similar results were obtained

by Lemos et al. (2014.)

Pili (Canariumovatum)

Canariumovatum, commonly known as Pili, is a native plant from the Philippines.

This tropical tree is a member of the family Burserceae with 600 species. Bulk of this tree
grow specifically in the provinces of Albay, Sorsogon, and Camarines Sur (General & Commented [P44]: grows

Guerrero, 2017). The pili fruit is drupe, 4 to 7 cm long and 2.3 to 3.8 cm in diameter and

weighs 15.7 to 45.7 g (10.1007%252Fs11746-000-0156-8.pdf) (Kakuda et al., 2000)..

Seeds of this tree is known to contain two white cotyledons with 8% moisture content, Commented [P45]: are

14.2% protein and 68.5% fat (Gallegos et al., 2013). This evergreen plant grow for a bout Commented [P46]: grows
Commented [P47]: about
20 to 35 meters.

Pili nut is the main product of this tree, though the other parts were also used Commented [P48]: specify since different paragraph

economically. The pili pulp is edible and can be consumed as appetizer or dessert. The oil

obtain from this tree is comparable to coconut oil and can be used for cooking and for

manufacturing soaps, perfumes and other cosmetic products (Gallegos et al., 2013).

Bioactivities of Pili

Kikuchi et al. (2012) isolated 18 known terpenoids from the resin of methanol

extract of C. Ovatum - four were sesquiterpenes and 14 were triterpenes. In evaluation for

melanogenes in B16 melanoma cells, three sesquiterpenes show inhibition with no

cytotoxicity to the cells namely, cryptomeridiol, 4-epicryptomeridiol , and cadin-

1(14)ene-7a,11-diol.

Antioxidant activity of Pili decreases upon digestion as shown in the ABTS and

DPPH assay of pili (Arenas et al., 2016). Based from the results of the study of Arenas et

al. (2016), the non-digested Pili showed higher antioxidant activity than the digested Pili.

This observed decrease is due to the decomposition of the phenolic compound in the

gastric environment.
Biochemical assays

Antioxidant assay

DPPH or 2,2´-diphenyl-1-picrylhydrazyl assay was used in determining the

antioxidant activity of caimito peel in an experiment done by Moo-Huchin et al. (2014).

This assay was done by first preparing the DPPH radical by mixing with methanol. The

DPPH radical were mixed with an amount of antioxidants extracts or standard in a test

tube. Absorbance of the solution were determined using UV-Vis spectrophotometer. At 1

min intervals the decrease in absorbance were measured at 515 nm.

The DPPH radical scaventint activity is affected by the presence of hydroxyl

groups in the phenolic and flavonoid compounds. Reduction of DPPH radicals into the

reduced form DPPH-H determined the ability of phenolic and flavonoid compounds to

donate hydrogen atom or electron to the unpaired DPPH radicals. When DPPH reacted

with radical scavenger, the purple color changed into pale purple or light yellow

(Nurhanan et al, 2012). The free radical scavenging activity was evaluated by absorption

reduction of the stable radical DPPH at 515 nm (Ningsihet al.,. 2016).

ABTS or 2,2´-Azinobis-3-ethylbenzotiazoline-6-sulfonic acid is another assay used

to determine the antioxidant activity of caimito peel. This assay used solution of ABTS,

HPLC-grade water and potassium persulfate which was incubated in dark at room

temperature for 16 h. The activated radical produced were mixed with ethanol then

subjected to UV-Vis spectrophotometer to obtain absorbance. Diluted ABTS solution

was then mixed with antioxidant extracts or standard and subjected to spectrophotometer

to obtain antioxidant capacity.

 Antibacterial assay

In the study of Alfredo et al. (2017), Staphylococcus aureus and Escherichia coli

were stored in nutrient agar slants with 20% glycerol at -20 C. Mueller-Hinton broth

placed in a test tube were inoculated with S. aureus and E. coli and incubated for 4 to 6

hours with ambient room temperature. Small amount of the bacterial inoculum were

added to the sterile Mueller-Hinton broth. The solution was then inoculated with

bacterial culture and incubated for 34 hours at 26-28 C. Erythromycin (ERY) for E. coli

and tetracycline (TET) for S. aureus served as positive control and sterile distilled water

served as negative control.

Anticancer assay

Manosroi et al. (2012) tested the cytotoxicity of cancer lines by SRB assay. The

fruit extracts showed cytotoxic effect against all cancer cell lines. Results shows that

triterpenoids, secondary metabolite constituents in L. Domesticum, is may be responsible

for the cytotoxic effects of the extracts. In addition, triterpenoids were the major

constituents which are responsible for the recorded high apoptotic effect. Triterpenoids

were inhibitor of tumor initiation and promotion according to Oguro et al. (1998) and

induce apoptosis (Choi et al., 2000).