Kagoshima University Research Center for the Pacific Islands, Occasional Papers No.34, 169-175, 2001
Part 2, Section 2, Report 5. The Progress Report of the 1999 Survey of the Research Project“Social Homeostasis of Small Islands in an Island-zone”

169

BIOLOGICAL ACTIVITY OF FEIJOA PEEL EXTRACTS
Hideki NAKASHIMA

Abstract
Fractions of Feijoa peel extracts were studied for anti-tumor activity, 50% cell cytotoxicity(CC50), anti-human immunodeficiency virus activity and anti-bacterial activity. Two active fractions[A3] of acetone extract and [M2] of MeOH extract had potent inhibitory activity against Gram-positive
and Gram-negative bacteria as well as fungi tested. These results indicate the therapeutic value of
Feijoa peel extracts as potential anti-tumor and anti-microbial organism agents.
Key words: anti-bacterial, anti-HIV, anti-tumor, Feijoa extracts

Introduction
Feijoa sellowiana Berg (Myrtaceae) is mainly cultured in tropical and subtropical countries
such as southern Brazil, Uruguay, Paraguay and northern Argentina. Feijoa has a pleasant flavor;

A

B

C

D

Fig. 1. Pictures of Feijoa. A; Feijoa leaves, B; Feijpa fruit in the tree, C; Feijor flower, D; Feijoa fruit.

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Hideki NAKASHIMA

it is also eaten as stew, jam, jelly or juice. Feijoa contains many medicinally bioactive compounds (Figure 1).
The predominant aroma of Feijoa fruits is ascribed to methylbenzoate and ethylbenzoate(H ARDY & M ICHAEL 1970; H ERRMANN 1994). Other volatile components have also been identified (P INO 1997; BINDER & FLATH 1989; Di CESARE et al. 1998; STARODUBTSEVA &K HAREBAVA 1978; S TARODUBTSEVA & K HAREBAVA 1986). Feijoa is known to contain highamounts of vitamin P (P)-active polyphenols, such as catechin, leucoanthocyanins, flavonols,
proanthocyanidins, and naphthoquinones (BABA-ZADE 1972; KRIVENTSOV & K ARAKHANOVA
1972b; GERSHTEIN & G ABISONIYA 1972; V ANIDZE 1991). Leaves of Feijoa also containcatechins, such as (+)-catechin, (-)-epicatechin, (+)-gallocatechin and (-)-epigallocatechin(V ANIDZE et al. 1991). Further, tannins in Feijoa fruits and leaves have been identified (FOO &PORTER 1981). The flower part of Feijoa contains anthocyanin-3-glucoside of polyphenols(L OWRY 1976). Moreover, Feijoa fruits are rich in vitamin C (K RIVENTSOV & K ARAKHANOVA
1972a; M GALOBLISHVILI & LOLOSIDI 1971 ; BABA-ZADE 1972; GERSHTEIN & GABISONIYA
1972; SALVO et al. 1987). Vitamine C in concentrated Feijoa juice can be up to three times
higher than that in fresh juice (Di CESARE et al. 1998). Provitamin A, such as α-carotene,
β-carotene and β-cryptoxanthin, has also been determined (HOMNAVA et al. 1990). Both watersoluble and water-insoluble fibers are found in Feijoa (S ALVO et al. 1987; LINTAS &CAPPELLONI 1992). Amino acids such as tryptophan, lysine, methionine and asparagine with nutritional value are also found in Feijoa extracts (TSIKORIDZE et al. 1986; S ALVO et al. 1987).
Feijoa peel contains more than twice as much vitamin C and catechin's vitamin P (P)-active
polyphenols, such as leucoanthocyanins, flavonols and naphthoquinones (K RIVENTSOV &K ARAKHANOVA 1972a). Although the chemical constituents of Feijoa fruit have been reported,
pharmaceutical studies of constituents of the peel have barely been carried out. Feijoa fruit peel
is generally wasted. However, the peel is rich in biologically and nutritiously interesting compounds. With this consideration in mind, we have screened for new biologically active agentsvarious fruits and vegetables that are consumed by many people, including Feijoa. The purposeof this study is to investigate anti-tumor activity, 50% cell cytotoxicity (CC50 ), anti-humanimmunodeficiency virus (HIV) activity and anti-bacterial activity of fractions of Feijoa peel extracts.

Materials and Methods
Preparation of Feijoa peel extracts.
Feijoa peel (330 g) was cut into small pieces and successively extracted with hexane, acetone, MeOH and 70% MeOH at room temperature. The solvent was concentrated in vacuo, andthe hexane extract [H 0](0.44 g), acetone extract [A0](5.9 g), MeOH extract [M 0](16.7 g) and
70% MeOH extract [70M0](6.1 g) were obtained, respectively. First, the aliquot of hexane extract [H0](0.4 g) was subjected to silica gel column chromatography, which was then eluted witha hexane-acetone gradient. The hexane fraction [H 1](32 mg), hexane-aceton (9:1) fraction [H2](157 mg), hexane-acetone (4:1) fraction [H 3](47 mg) and [H 4](45 mg) were eluted stepwise.
Second, the acetone extract [A 0](5.5 g) was subjected to silica gel column chromatography,
which was then eluted with a benzene-EtOAc gradient. The benzene fraction [A1](92 mg),benzene-AcOEt (10:1) fraction [A2] (103 mg), [A3](145 mg), benzene-AcOEt (1:1) fraction[A4](170 mg), AcOEt fraction [A 5](582 mg), AcOEt-EtOH (5:1) fraction [A6](836 mg) and[A7](854 mg) were eluted stepwise. Third, the MeOH extract [E 0](16.7 g) was subjected toBovine Congenital Defects Caused by Arbovirus of Tropic Origin

171

Fig. 2. Fractional separation of Feijoa peel extracts. CC: column chromatography.

silica gel column chromatography, which was then eluted with a CH2 Cl2 -MeOH gradient. The
CH2 Cl2 fraction [M 1](24 mg), CH 2 Cl2 -MeOH (50:1) fraction [M 2](135 mg), CH 2C1 2-MeOH(9:1) fraction [M 3](25 mg), [M 4](761 mg), CH2 C1 2-MeOH (4:1) fraction [M 5](1317 mg), and[M 6](2100 mg) were eluted stepwise. Finally, the 70% MeOH extract [70M0](5 g) was subjected ODS column chromatography, which was then eluted with a H2 O-MeOH gradient. The
H 2O-MeOH (2:1) fraction [70 M 1 ] (2010 mg), [70M2](80 mg), H 2O-MeOH (1 : 1) fraction [70
M 3](18 mg), [70 M4] (43 mg), and MeOH fraction [70 M 5](1200 mg) were eluted stepwise(Figure 2).
Assay for anti-tumor and cytotoxic activity.
Human oral squamous cell carcinoma cells (HSC-2), human oral salivary tumor cells (HSG)and human oral gingival fibroblasts (HGF) (5-7 population doubling levels) were cultured in
Dulbecco’s modified Eagle’s minimum essential medium (DMEM) supplemented with 10%
heat-inactivated fetal bovine serum (FBS). These cells were incubated for 24 h with the indicatedconcentrations of test compounds, and the viable cell number was then determined by 3-(4,4dimethylthiazol-2-yl)-2,5-diphenyl-tetrazonium bromide (MTT) assay. In brief, cells were
washed with phosphate buffered saline (PBS), and incubated for 4 h with a fresh culture medium
containing 0.2 mg/ml MTT. After removing the medium, cells were lysed with 100 µl DMSO
and the absorbance at 540 nm of the cell lysate was measured with Labsystems MultiskanR(Biochromatic) with Star/DOT Matrix printer JL-10. The CC50 was determined from the doseresponse curve.

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Hideki NAKASHIMA

Anti-HIV assay
The suppression of HIV-induced cytopathic effects by Feijoa extracts was studied. Human
T cell leukemia virus 1 (HTLV-1)-bearing CD4 positive human T cell lines, MT-4 cells, were
infected with HIV-1IIIB at a multiplicity of infection (m.o.i.) of 0.01. HIV-infected or mockinfected MT-4 cells (1.5 x 10 5 /ml, 200 µl/well) were placed into 96-well microtiter plates in
RPMI 1640 medium supplemented with 10% heat-inactivted fetal calf serum (FCS) and incubated in the presence of varying concentrations of test compounds. After incubation for 5 days
at 37ºC in a CO 2 incubator, cell viability was quantified by a colorimetric assay (at 540 and 690nm), monitoring the ability of viable cells to reduce MTT to a blue formazan product(N AKASHIMA et al. 1992b). All data represent the mean values of triplicate measurements.
Anti-bacterial assay.
Escherichia coli LE 140 F’lac, Pseudomonas aeruginosa, Staphylococcus epidermidts,
Candida albioans, and Candida glabrata were isolated from clinical specimens obtained fromthe Department of Clinical Microbiology, Medical University of Szeged, Hungary. Helicobacter
pylori (ATCC43504) was purchased from the American Type Culture Collection (Rockville,
MD).
The anti-bacterial experiments were made by adding 10µl of original solutions as a droplet
on a minimal medium supplemented with 1% trypton and 0.5% yeast extract (MTE) broth andblood agar plates incubated with 10 5 cells of the tested strains. The plates were incubated at 37ºC
for 24 h, then the inhibitory zones were measured. As a control 10µl of DMSO was examinedon each strain. It was found that C. albicans, C. glabrata and P. aeruginosa were sensitive to
DMSO. The growth of E. coli and S. epidermidis was not suppressed by DMSO. Ampicillin discwas used as positive control in the studies of antibacterial effects.
The micro-dilution broth method was used to determine the minimum inhibitory concentration (MIC) against H. pylori growth. Mueller-Hilton broth containing 5% FBS was used as themedium, and was cultured in a jar conditioned with Campylo Pack (Dia Iatron) for 48 h. Briefly,
H. pylori strains were inoculated on a Brucella agar plate containing 10% horse serum, and cultured at 37ºC for 48 h. The collected bacterial colonies were diluted to a 10 7 colony forming unit
(UFC)/ml with 0.9% saline. The fractions were dissolved in DMSO, and then diluted with
Mueller-Hilton broth. To the solution of the fractions, a suspension solution of the bacteria wasadded to make 10 6 CFU/100ml/well. The mixture was incubated at 37ºC for 48 h. The MIC values of tested fractions were determined by observation of colony formation.

Results and Discussion
Anti-tumor and cytotoxic activity.
It is widely accepted that, in tumor cells, agents act as apoptosis-inducers and an importantreason for nonresponsiveness is the insufficiency of these drugs to trigger apoptosis (H ICKMAN
1992). The cytotoxicity of all fractions was evaluated in two human oral tumor cell lines (HSG2, HSG) and one human gingival fibroblasts (HGF), using the microculture tetrazolium (MTT)
assay. Crude hexane [H0], acetone [A0], MeOH [M0] and 70% MeOH [70M0] extracts showed
weak cytotoxic activity against these cells. However, after separation by column chromatography(Figure 2), cytotoxic activities became detectable in several fractions (Table 1). Most fractions
exhibited only low cytotoxicity (IC50 values >100 µg/ml). However, fraction [A3] displayed significant potency and was relatively cytotoxic to two tumor cell lines (HSC-2 and HSG) and the

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Bovine Congenital Defects Caused by Arbovirus of Tropic Origin

Table 1. Activitiy of anti-cancer, anti-HIV and anti-bacteria of Feijoa peel extracts
Cytitoxic activity (CC 50 µg/ml)
Fraction
HSC -2 HSG

HGF

Anti-HIV activity

Anti-bacterial activity (MIC 50, µg/ml)

SI
C C50
EC 50
SI
Staph.lococcus
Pseudomonas Candia Candi da Helicobacter
E.coli(HGF/HSC-2) (µg/ml) (µg/ml) (CC 50/EC50) epidermidis
aeruginosa
albi cans glabrata pylori

H0
H1
H2
H3
H4

381
386
395
＞ 500 ＞ 500 ＞ 500
313
357
370
＞ 500 ＞ 500 ＞ 500
269
356
409

1.0
＞＜ 1
1.2
＞＜ 1
1.5

100
＞ 200
162
＞ 200
117

＞ 200
＞ 200
＞ 200
＞ 200
＞ 200

＜1
＞＜ 1
＜1
＞＜ 1
＜1

+
-

+
+
+

A0
A1
A2
A3
A4
A5
A6
A7

444
462
452
160
302
305
132
123
165
81
79
80
274
318
382
335
383
403
＞ 500 ＞ 500 ＞ 500＞ 500 ＞ 500 ＞ 500

1.0
1.0
1.3
1.0
1.4
1.2
＞＜ 1
＞＜ 1

119
121
113
22
46
127
＞ 200
＞ 200

＞ 200
＞ 200
＞ 200
＞ 40
＞ 200
＞ 200
＞ 200
＞ 200

＜1
＜1
＜1
＜1
＜1
＜1
＞＜ 1
＞＜ 1

++
-a
-

+++
++
++
-

M0
M1
M2
M3
M4
M5
M6

＞ 500 ＞ 500 ＞ 500
378
421
441
119
101
152
338
268
277
458
235
＞ 500
＞ 500 ＞ 500 ＞ 500
458 ＞ 500 ＞ 500

＞＜ 1
1.2
1.3
0.8
＜ 0.5
＞＜ 1
＞ 1.1

＞ 200
＞ 200
120
78
＞ 200
＞ 200
＞ 200

＞ 200
＞ 200
＞ 200
＞ 200
＞ 200
＞ 200
＞ 200

＞＜ 1
＞＜ 1
＜1
＜1
＞＜ 1
＞＜ 1
＞＜ 1

-a
+
-a
-

++
+-

＞ 500 ＞ 500＞ 500 ＞ 500＞ 500 ＞ 500＞ 500 ＞ 500
280
419＞ 500 ＞ 500

＞ 1.3
＞ 1.3
＞ 1.3
＞2
2.0
＞ 1.0

＞ 200
＞ 200
99
83
102
＞ 200

＞ 200
＞ 200
15
＞ 200
＞ 200
＞ 200

＞＜ 1
＞＜ 1
7
＜1
＜1
＞＜ 1

-a
-a
-a
-

+
+
+
+
-

＞ 1000＞ 1000
268
2395

1.16
13.00
0.34
6.66

＞ 861
＞ 71
787
360

70M0
70M1
70M2
70M3
70M4
70M5
Curdlan sulfate
Dextran sulfate
AZT (µM)
ddC (µM)

380
396
394
250
208
488

a

ND b
＞ 100
＞ 100
＞ 100
＞ 100

+
-

+
+

+
+
+

++
++
++
+
++
+
-

+

+

ND b
＞ 100
＞ 100
＞ 100
＞ 100
＞ 100
＞ 100
＞ 100
ND b
＞ 100
＞ 100
＞ 100
＞ 100
＞ 100
＞ 100
ND b
＞ 100
＞ 100
＞ 100
＞ 100
＞ 100

b

++ very potent; + potent, + - slightly potent; - no effect. inhibited ha emolysis. Not de tected.
Metronidazole, Clarithromyc in, a nd Eruthromyc in were used as r eference anti-bacterial compounds for H.pylori, and their MIC50 values
were 74 µg/m l, 1.9 µg/ml, and 1,8 µg/m l, respectively.

healthy cell line (HGF). When water solubility was increased, the relative cytotoxic activity
against normal cells (SI=HGF/HSC-2) generally declined (Table 1).
Anti-HIV activity.
Table 1 demonstrated that most fractions did not significantly inhibit HIV-induced
cyopathic effects on MT-4 cells (selectivity index (SI)=1-7), compared to higher anti-HIV activities of curdlan sulfate (CRDS), dextran sulfate (DS), AZT, and ddC (SI=71-861). However, fraction [70M2] showed some anti-HIV activity. The present study demonstrates that fraction[70M2] with relatively high water-solubility has the highest anti-HlV activity (Table 1). This isconsistent with our previous finding that anti-HIV activity increases with water-solubility of thecompounds: in the order of lignin (NAKASHIMA et al. 1992a) > hydrolyzable tannins > condensed
tannins (NAKASHIMA et al. 1992b) > flavonoids (S AKAGAMI et al., unpublished data), and higher
molecular weight lignins and tannins (NAKASHIMA et al. 1992b) show higher anti-HIV activity

174

Hideki NAKASHIMA

in comparison with their lower molecular compounds.
Anti-bacterial activity.
Chemotherapy with antibiotics sometimes induces serious side effects, such as diarrhea,
nausea, abnormal taste, dyspepsia, abdominal pain/discomfort, headache, and angioedema.
Therefore, there is a strong demand for compositions having all of the beneficial anti-bacterial
properties with reduced side effects. Antibacterial agents of fruit or vegetable origin may be superior as compared with many antibiotics. All fractions were tested against a panel of microorganisms, including the Gram-positive bacteria, Staphylococcus epidermidis, and Gram-negative
bacteria, Escherichia coli. and Pseudomonas aeruginosa, and the fungi, Candida albicans andCandida glabrata (Table 1). The highest level of activity consistently arose in the acetone and
MeOH extract fractions. Two fractions [A3] and [M2] were found to exhibit potent anti-bacterial
activity against both Gram-positive and Gram-negative bacteria as well as Candida.
Anti-Helicobaoter pylori aotivity.
The current medical consensus suggests that H. pylori is the primary causative organism foracute gastritis (BLASER 1992). Since the drug will directly come in contact with the lining of thestomach, edible plants may be a good source of oral antiulcer agents. However, all Feijoa extract
fractions did not exhibit potent anti-H. pylori activity (MIC50>100 µg/ml), in contrast with threeeffective positive controls: metronidazole, clarithromycin and erythromycin (Table 1).
In conclusion, it is important to note that fractions [A3] and [M2] showed promising antibacterial activity. Although the activities were not strong, some fractions of Feijoa peel extracts
showed anti-tumor and anti-HIV activity. The results of this study apparently indicate therapeutic value for Feijoa peel extracts. Further work is required to identify and isolate the compounds
responsible for the observed biological activity.

Acknowledgements
The author is grateful for the collaboration of Dr. N. Motohashi (Meiji Pharmaceutical
Univ.), Dr. M. Kawase and Dr. Y. Shirataki (Faculty of Pharmaceutical Sciences, Josei Univ.),
and Dr. H. Sakagami (Meikai Univ. School of Dentistry).

References
BABA-ZADE, F. I. 1972. Accumulation of vitamin C in fruits of the feijoa. In: Tr Vses Semin
Biol Aktiv (Lech) Veshchestvam Plodov Yagod, 4th. (ed. F RANCHUK, E.P.), 183-186. Vses
Nauch-Issled Inst Sadovod, Michurinsk, USSR.
BINDER, R. G. & FLATH, R. A. 1989. Volatile components of pineapple guava. J Agric Food
Chem. 37: 734-736.
BLASER , M. J. 1992. Helicobaoter pylori: its role in disease. J. Clin. Infect. Dis. 15: 381-393.
Di CESARE, L. F., NANI, R., CITRO, D. & P ROIETTI, M. 1998. Concentration of aroma compounds and soluble substances in a feijoa juice by cold finger freezing-out. Ind Bevande 27:
126-130 (Chem. Abst. 128: 320839a).
FOO, L. Y. & PORTER, L. J. 1981. The structure of tannins of some edible fruits. J Sci Food
Agric. 32: 711 -716.
G ERSHTEIN, L. A. & GABISONIYA, L. D. 1972. Biologically active substances of the fruit of the
Feijoa. In: Tr Vses Semin Biol Aktiv (Lech) Veshchestvam Plodov Yagod, 4th. (ed.

Bovine Congenital Defects Caused by Arbovirus of Tropic Origin

175

FRANCHUK, E.P.), 187-189. Vses Nauch-Issled Inst Sadovod, Michurinsk, USSR.
H ARDY, P. J. & M ICHAEL, B. J. 1970. Volatile components of Feijoa fruits. Phytochemistry 9:1355-1357.
H ERRMANN, K. 1994. The aromatio substances of exotic fruits (part 2): Passion fruit, guava,
feijoa. Fluess Obst. 61: 398-401.
H ICKMAN J. A. 1992. Apoptosis induced by anticancer drugs. Canoer Metastasis 11: 121-139.
H OMNAVA, A., ROGERS, W. & EITENMILLER, R. R. 1990. Provitamin A activity of specialty fruit
marketed in the United States. J Food Compos Anal. 3: 119-133.
K RIVENTSOV. V. I. & K ARAKHANOVA, S. V. 1972a. Biologically active substances of several
subtropical fruit crops of the Crimea. In: Tr Vses Semin Biol Aktiv (Lech) Veshchestvam
Plodov Yagod, 4th. (ed. FRANCHUK, E.P.), 82-86. Vses Nauch-Issled Inst Sadovod,
Michurinsk, USSR.
K RIVENTSOV. V.I. & K ARAKHANOVA, S. V. 1972b. Changes in biochemically active
Substanoces in fruits of subtropical crops mediated by calcium iodide. In: Tr Vses Semin
Biol Aktiv (Lech) Veshchestvam Plodov Yagod, 4th. (ed. FRANCHUK, E.P.), 267-270. Vses
Nauch-Issled Inst Sadovod, Michurinsk, USSR.
LINTAS, C. & CAPPELLONI, M. 1992. Dietary fiber content of Italian fruit and nuts. J Food
Compos Anal. 5: 146-151.
LOWRY, J. 1976. Anthocyanins of the Melastomataceae, Myrtaceae and some allied families.
Phytochemistry 15: 513-516.
M GALOBLISHVILI, T. S. & LOLOSIDI, S. I. 1971. Biochemical characteristics of the fruits fromdifferent forms of feijoa. Subtrop Kul’t 6: 122-124.
N AKASHIMA, H., M URAKAMI, T., Y AMAMOTO, N., N AOE, T., KAWAZOE, Y., K ONNO, K. &SAKAGAMI, H. 1992a. Lignified Materials as Medicinal Resources. V. Anti-HIV (humanimmunodeficiency virus) activity of some synthetic lignins. Chem. Pharm. Bull. 40: 21022105.
N AKASHIMA, H., M URAKAMI, T., Y AMAMOTO, N., S AKAGAMI, H., T ANUMA, S., HATANO, T.,
YOSHIDA, T. & O KUDA, T. 1992b. Inhibition of human immunodeficiency viral replication
by tannins and related compounds. Antiviral Res. 18: 91-103.
PINO, J.A. 1997. Volatile constituents of tropical fruits. Ill. Feijoa, sapodilla and cashew apple.
Alimentaria (Madrid) 286: 41-45 (Chem. Abst. 127: 3304474w).
SALVO, F., T OSCANO, M. A. & D UGO, G. 1987. Chemical composition of Feijoa sellowiana
fruit. Riv Soc Ital Sci Aliment. 16: 471-474.
STARODUBTSEVA. V. P. & KHAREBAVA, L. G. 1978. Volatile complex of Feijoa sellowiana. Part
3. Subtrop Kul’t. 5: 70-72.
STARODUBTSEVA, V P. & KHAREBAVA, L. G. 1986. Volatile components of feijoa. Part 4.
Composition of the essential oil. Subtrop Kul’t. 5: 118-123.
TSIKORIDZE, N. G., TKESHELASHVILI, C. F. & KHURTSILAVA, D. I. 1986. Study of the free aminoacids of Feijoa fruit. Subtrop Kul’t. 2: 137-139.
V ANIDZE, M. R. 1991. Quantitative changes in foliar catechins, proanthocyanidins, andflavonols. Subtrop Kul’t. 3: 80-81.
V ANIDZE, M. R., S HALASHVILI, A. G., CHKHIKVISHVILI, l. D. & KVESITADZE, E. G. 1991.
Catechins of feijoa leaves. Subtrop Kul’t. 6: 87-90.