Bees have been in existence for 125 million years and their
evolutionary success has allowed them to become perennial
species that can exploit virtually all habitats on Earth. This
success is largely because of the chemistry and application of
the specific products that bees manufacture: honey, beeswax,
venom, propolis, pollen and royal jelly. As the most important
‘chemical weapon’ of bees against pathogenic microorganisms,
propolis has been used as a remedy by humans since ancient
times. It is still one of the most frequently used remedies in the
Balkan states (1), applied for treatment of wounds and burns,
sore throat, stomach ulcer, etc.
For this reason, propolis has become the subject of intense
pharmacological and chemical studies for the last 30 years.
As a result, much useful knowledge has been gathered.
However, it is important to note that in the last decade, the
paradigm concerning propolis chemistry radically changed. In
the 1960s, propolis was thought to be of very complex, but
more or less constant chemistry, like beeswax or bee venom
(2,3). In the following years, analysis of numerous samples from
different geographic regions led to the disclosure that the
chemical composition of bee glue is highly variable. This
circumstance was soon understood by seasoned chemists, such
as Popravko (4) and Ghisalberti (5). Nevertheless, most of the
scientists studying the biological properties of propolis contin-
ued to assume that the term ‘propolis’ was as determinative with
respect to chemical composition as the botanical name for a
medicinal plant. Numerous studies, carried out with the com-
bined efforts of phytochemists and pharmacologists, led in
recent years to the idea that different propolis samples could be
completely different in their chemistry and biological activity.
The Problem of Chemical Variability of
To understand what causes the differences in chemical com-
position, it is necessary to keep in mind the plant origin of
propolis. For propolis production, bees use materials resulting
from a variety of botanical processes in different parts of plants.
These are substances actively secreted by plants as well as sub-
stances exuded from wounds in plants: lipophilic materials on
leaves and leaf buds, gums, resins, latices, etc. (6). The plant
origin of propolis determines its chemical diversity. Bee glue’s
chemical composition depends on the specificity of the local
flora at the site of collection and thus on the geographic and
© The Author (2005). Published by Oxford University Press. All rights reserved.
Recent trends and important developments in propolis research
Institute of Organic Chemistry with Centre of Phytochemistry, Bulgarian Academy of Sciences, 1113 Sofia,
The newest developments in propolis pharmacological research are summarized. The problem regarding
biological studies, caused by the chemical variability of propolis, is discussed. The most important
trends and developments in recent propolis research are outlined: biological studies performed with
chemically characterized samples, bioassay-guided studies of active principles and comparative biological
studies of propolis of different origin and chemical composition. These types of studies are extremely
valuable with respect to propolis standardization and practical applications in therapy. They will allow
scientists to connect a particular chemical propolis type to a specific type of biological activity and
formulate recommendations for practitioners.
Keywords: propolis – plant origin – bioactive compounds – composition/activity relationship
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climatic characteristics of this site. This fact results in the
striking diversity of propolis chemical composition, especially
of propolis originating from tropical regions.
Nowadays, it is well documented that in the temperate zone
all over the world, the main source of bee glue is the resinous
exudate of the buds of poplar trees, mainly the black poplar
Populus nigra (7). For this reason, European propolis contains
the typical ‘poplar bud’ phenolics: flavonoid aglycones
(flavones and flavanones), phenolic acids and their esters (8).
Poplar trees are common only in the temperate zone; they can-
not grow in tropical and subtropical regions. For this reason, in
these habitats, bees have to find other plant sources of propo-
lis to replace their beloved poplar. As a result, propolis from
tropical regions has a different chemical composition from that
of poplar type propolis. In the last decade, Brazilian propolis
attracted both commercial and scientific interest. The main
source of Brazilian bee glue turned out to be the leaf resin of
Baccharis dracunculifolia (9,10). Among the main compound
classes found in Brazilian propolis are prenylated derivatives
of p-coumaric acid and of acetophenone. Diterpenes, lignans
and flavonoids (different from those in ‘poplar type’ propolis)
have also been found (9). However, in Brazil, several types of
propolis were registered in recent studies (11,12), that come
from plant sources different from B.dracunculifolia and
containing compounds other than those mentioned above.
Recently the chemistry of Cuban propolis caught the attention
of scientists. Its main components are polyisoprenylated ben-
zophenones, and this makes Cuban propolis different from
both European and Brazilian bee glue. The plant source of this
propolis type was detected to be the floral resin of Clusia
rosea, from whence came the prenylated benzophenones (13).
There is no doubt that in other ecosystems, propolis plant
sources and the chemical composition of propolis will con-
tinue to surprise scientists.
The distinct chemistry of propolis from different origins
leads to the expectation that the biological properties of differ-
ent propolis types will be dissimilar. However, in most cases.
this is not true! Actually, propolis is the defense of bees against
infections, and the antibacterial and antifungal activity of all
samples is not surprising. The similarity in many of the other
types of activity is less obvious but it is a fact. Of course, the
responsible compounds are different, as shown in Table 1: mainly
favanones, flavones, phenolic acid and their esters in poplar
type (European) propolis, prenylated p-coumaric acis and
diterpenes in Baccharis type (Brazilian) propolis; prenylated
benzophenones in Cuban red propolis, etc.
The only exception seems to be the allergenic property of
European (poplar type) propolis. This problem needs detailed
investigations. Until now, no studies have been performed to
find out if other propolis types have allergenic properties. It is
very tempting to search for propolis that causes no contact
allergy or causes it much less often.
The fact that different chemistry leads to the same type of
activity and in some cases even to activity of the same order of
magnitude is amazing. Nonetheless, it is important to have
detailed and reliable comparative data on every type of biolog-
ical activity, combined with chemical data, in order to decide if
some specific areas of application of a particular propolis type
can be formulated as preferable. The biological tests have to
be performed with chemically well characterized and, if possible,
chemically standardized propolis. Such detailed comparative
investigations are a challenge to propolis researchers. The most
important recent developments in propolis research are those
which are aimed at meeting this particular challenge.
Important Trends and Developments in
Recent Propolis Research
Biological Studies Performed with Chemically
More and more publications are appearing which combine
antimicrobial and other biological studies with chemical
analyses of the tested propolis samples. The most often used
techniques for chemical analyses are gas chromatography–
mass spectrometry (GC–MS) (18–24) and high-performance
liquid chromatography (HPLC) (25–27). In a recent work,
qualitative chemical characterization of the samples tested for
antibacterial activity was combined with quantification of
the major groups of biologically active substances of the
corresponding samples (28). The use of chemically character-
ized propolis samples for biological experiments is the only
30 Recent trends in propolis research
Table 1. Compounds responsible for the biological activity of different propolis types
Propolis type Antibacterial Antiinflammatory Antitumor Hepatoprotective Antioxidant Allergenic
activity activity activity activity activity action
European Flavanones, flavones, Flavanones, flavones, Caffeic acid Caffeic acid, ferulic Flavonoids, phenolic 3,3-Dimethylallyl
(poplar type) phenolic acids and phenolic acids and phenethyl ester (16) acids acid, caffeic acid and their esters (15) caffeate (14)
their esters (14) their esters (15) phenethyl ester (15)
Brazilian Prenylated p-coumaric Unidentified (15) Prenylated p-coumaric Prenylated p-coumaric Prenylated p-coumaric Not tested
(Baccharis acis, labdane acids, clerodane acis, flavonoids, acis, flavonoids (15)
type) diterpenes (15) diterpenes, lignans, caffeoyl
benzofuranes (15) quinic acids (15)
Cuban Prenylated Not tested Prenylated Unidentified (15) Prenylated Not tested
benzophenones (17) benzophenones (13) benzophenones (13)
Taiwanese Not tested Not tested Prenylated Not tested Prenylated Not tested
flavanones (42) flavanones (42)
meaningful way to study the biological and pharmacological
activities of bee glue at the beginning of the third millennium.
Bioassay-guided Studies of Active Principles
Studies based on bioassay-guided chemical analysis represent
a very promising trend in propolis research. Using this
approach, Chen et al. isolated two new cytotoxic prenylflavones
from Taiwanese propolis (29). Both compounds demonstrated
cytotoxic properties on three cancer cell lines and also were
potential radical scavengers – radicals of 1,1-diphenyl-2-picryl-
hydrazyl (DPPH). Banskota et al. (30) isolated the active
components from Netherlands propolis with antiproliferative
activity in cancer cell lines: caffeic acid phenethyl ester
(CAPE) and several analogs, including two new glyceryl esters
of substituted cinnamic acids. The same compounds were
found by Nagaoka et al. (31) to be responsible for the nitric
oxide-inhibiting activity of Netherlands propolis. Usia et al.
(32) isolated from Chinese propolis a number of compounds
with antiproliferative activity. Most of them were known
‘poplar propolis’ constituents, but among them were
two new flavonoids: 2-methylbutyrouylpinopbanskin and
6-cinnamylchrysin. From Greek propolis, the new flavanone
derivative 7-prenylpinocembrin has been isolated, together
with totarol and 7-prenylistrobopinin, as important antibacterial
Banskota et al. (34) studied Brazilian propolis in order to
identify the substances with hepatoprotective activity and
those active against Helicobacter pylori. They found that these
activities were due mainly to phenolic components, but diter-
penic acids also contributed to hepatoprotective activity.
Several anti-HIV compounds, derivatives of moronic acid, and
a new triterpenoid called melliferon were isolated from
Brazilian bee glue (35). The major component of Cuban red
propolis, the prenylated benzophenone nemorosone, was
found to possess cytotoxic activity against several tumor cell
lines and to have radical scavenging action (13).
Comparative Biological Studies of Propolis of Different
Origin and Chemical Composition
Perhaps the most interesting trend in recent propolis research
is the comparative study of biological properties of propolis
from different geographic locations and different chemical
composition. The number of this type of studies is as yet
limited. Kujumgiev et al. (36) compared the antimicrobial
(antibacterial, antifungal and antiviral) activity and chemical
composition of propolis from diverse geographic origins.
The results presented unambiguous proof that in spite of the
great differences in the chemical composition of propolis
from different geographic locations, all samples exhibit signif-
icant antibacterial and antifungal (and most of them antiviral)
activity. This study clearly demonstrated that in different
samples, different combinations of substances are essential for
the biological activity of bee glue. Trying to develop this
comparative approach, Popova et al. (37) searched for a statis-
tically significant correlation between biological activity and
geographic origin of propolis samples. Analysis of variance
(ANOVA) was used to compare the antibacterial action of
three groups of bee glue: European, Brazilian and Central
American. The results showed that propolis from Europe and
Brazil had similar activity despite the drastic differences in
chemical composition. Their antibacterial activity was signifi-
cantly higher than that of Central American propolis. The
ANOVA was also applied to compare the toxicity of the same
three propolis groups with Artemia salina (Crustaceae). In this
case, there was no significant correlation between geographic
origin and potential cytotoxicity. This demonstrates that the
search for new promising cytotoxic compounds in new pro-
polis sources is reasonable.
The cytotoxic, hepatoprotective and free radical scavenging
activity of propolis from Brazil, Peru, The Netherlands and
China was compared by Banksota et al. (38). They found that
propolis from The Netherlands and China possessed the
strongest cytotoxic activity; while almost all samples pos-
sessed significant hepatoprotective activity. The scavenging
activity against DPPH free radicals of all samples was similar;
only the Peruvian sample showed weaker scavenging activity.
Salomao et al. (39) compared the microbicidal activity of
Brazilian and Bulgarian propolis and analyzed their chemical
composition by High Temperature – High Resolution Gas
Chromotography – Mass Spectrometry (HT-HRGC-MS), and
found that although they were of totally distinct compositions,
they were active against Trypanozoma cruzi and some patho-
The work of Kumazawa et al. (40) is an excellent example of
this approach. The authors compared the antioxidant activity of
propolis of various geographic origins (Argentina, Austria,
Brazil, Bulgaria, Chile, China, Hungary, New Zealand,
South Africa, Thailand, Ukraine, Uruguay, the USA and
Uzbekistan) and combined these data with chemical analyses.
Major constituents of the samples tested were identified by
HPLC with photo-diode array and mass spectrometric detec-
tion. Seventeen phenolic compounds in 16 kinds of propolis
were identified and quantified by HPLC. Propolis with strong
antioxidant activity contained antioxidative compounds such
as kaempferol and phenethyl caffeate. In the same way, antiox-
idant activities and chemical constituents of propolis from
different regions of Japan were compared by the same research
group (41). They concluded that strong antioxidant activity
correlated with a high concentration of caffeic acid and
phenethyl caffeate. In addition, propolis from Okinawa was
found to have antioxidants not seen in propolis from other
Following a similar model, Chen et al. (42) compared the
radical scavenging activity, cytotoxic effects and apoptosis
induction in human melanoma cells of Taiwanese propolis
from different locations. Propolins (C-prenylated flavanones)
in the samples were detected by HPLC, and the total phenolic
content was determined by spectrophotometry. The high
concentration of propolins was found to be essential for the
apoptosis induction in human melanoma cells and for the
eCAM 2005;2(1) 31
Such comparative studies are extremely valuable with respect
to propolis standardization and practical applications in therapy.
It is our hope that in the near future their number is going to
grow significantly. They will allow scientists to connect a par-
ticular chemical propolis type to a specific type of biological
activity and formulate recommendations for the practitioners.
This could help the general public to make more efficient use
of the beneficial properties of propolis with respect to CAM.
1. Wollenweber E, Hausen BM, Greenaway W. Phenolic constituents and
sensitizing properties of propolis, poplar balsam and balsam of Peru.
Bull Groupe Polyphenols 1990;15:112–20.
2. Lindenfelser LA. Antimicrobial activity of propolis. Am Bee J 1967;107:
3. Kivalkina BP. Propolis: its antimicrobial and healing properties. PhD
Dissertation, Kazan University, 1964 (in Russian).
4. Popravko SA. Chemical composition of propolis, its origin and standard-
ization. In: A Remarkable Hive Product: PROPOLIS. Harnaj, V. (ed)
Bucharest: Apimondia Publishing House, 1978, 15–8.
5. Ghisalberti EL. Propolis: a review. Bee World 1978;60:59–84.
6. Crane E. Beekeeping: Science, Practice and World Recourses.
Heinemann, London, 1988.
7. Bankova VB, De Castro SL, Marcucci MC. Propolis: recent advances in
chemistry and plant origin. Apidologie 2000;31:3–15.
8. Bankova V, Popova M, Bogdanov S, Sabatini AG. Chemical composition
of European propolis: expected and unexpected results. Z Naturforsch
9. Marcucci MC, Bankova VS. Chemical composition, plant origin and
biological activity of Brazilian propolis. Curr Top Phytochem 1999;2:
10. Kumazawa Sh, Yoned M, Shibata I, Kanaeda J, Hamasaka T, Nakayama Ts.
Direct evidence for the plant origin of Brazilian propolis by the observa-
tion of honeybee behavior and phytochemical analysis. Chem Pharm Bull
11. Park YK, Alencar SM, Aguiar CL. Botanical origin and chemical compo-
sition of Brazilian propolis. J Agric Food Chem 2002;50:2502–6.
12. Sawaya ACHF, Tomazela DM, Cunha IBS et al. Electrospray ioniza-
tion mass spectrometry fingerprinting of propolis. Analyst 2004;129:
13. Cuesta Rubio O, Frontana-Uriba BA, Ramirez-Apan T, Cardenas J.
Polyisoprenylated benzophenones in Cuban propolis; biological activity
of nemorosone. Z Naturforsch 2002;57c:372–8.
14. Burdock GA. Review of the biological properties and toxicity of bee
propolis (propolis). Food Chem Toxicol 1998;36:347–63.
15. Banskota AH, Tezuka Y, Kadota Sh. Recent progress in pharmacological
research of propolis. Phytother Res 2000;15:561–71.
16. Grunberger D, Banerjee R, Eisinger K et al. Preferential cytotoxicity on
tumor cells by caffeic acid phenethyl ester isolated from propolis.
17. Hernandez NMR, Cuesta Rubio O, Aviles A, Avellanede DLS. Actividad
antimicrobiana de nemorosona aislada de Clusia rosea. Rev Cubana Farm
2001;35 (Suppl Especial):197–9.
18. Velikova M, Bankova V, Sorkun K, Houcine S, Tsvetkova I, Kujumgiev A.
Propolis from the Mediterranean region: chemical composition and
antimicrobial activity. Z Naturforsch 2000;55c:790–3.
19. Keskin N, Hazir S, Baser HC, Kurkcuoglu M. Antibacterial activity and
chemical composition of Turkish propolis. Z Naturforsch 2001;56c:
20. Hegazi AG, Abd El Hady F. Egyptian propolis: 1—Antimicrobial activity
and chemical composition of Upper Egypt propolis. Z Naturforsch 2001;
21. Hegazi AG, Abd El Hady F. Egyptian propolis: 3—Antioxidant, antimi-
crobial activities and chemical composition of propolis from reclaimed
lands. Z Naturforsch 2002;57c:395–402.
22. Abd El Hady F, Hegazi AG. Egyptian propolis: 2—Chemical composi-
tion, antiviral and antimicrobial activity of East Nile Delta propolis.
Z Naturforsch 2002;57c:386–91.
23. Yildirim Z, Hacievlyagil S, Onur Kutlu N et al. Effect of water extract of
Turkish propolis on tuberculosis infection in guinea-pigs. Pharmacol Res
24. Erdem BG, Olmez S. Inhibitory effect of Bursa propolis on dental caries
formation in rats inoculated with Streptococcus sobrinus. Turk J Zool
25. De Laurentis N, Cafarchia C, Lai O, Losacco V, Milillo MA. Chemical
composition and biological investigation of Apulia region propolis. Riv
Ital EPPOS 2002;34:29–41.
26. Santos FA, Bastos EMA, Uzeda M et al. Antibacterial activity of Brazilian
propolis and fractions against oral anaerobic bacteria. J Ethnopharmacol
27. Da Silva Cunha IB, Salomao K, Shimizu M et al. Antitrypanosomal activity
of Brazilian propolis from Apis mellifera. Chem Pharm Bull 2004;52: 602–4.
28. Popova M, Silici S, Kaftanoglu O, Bankova V. Antibacterial activity of
Turkish propolis and its qualitative and quantitative chemical com-
position. Phytomedicine 2004. In Press, corrected proof online, 5th
29. Chen Ch, Wu, Chi, Shy H, Lin J. Cytotoxic prenylflavones from
Taiwanese propolis. J Nat Prod 2003;66:503–6.
30. Banskota AH, Nagaoka T, Sumioka LY et al. Antiproliferative activity of
the Netherlands propolis and its active principles in cancer cell lines.
J Ethnopharm 2002;80:67–73.
31. Nagaoka T, Banskota AH, Tezuka Ya, Midorikawa K, Matsushige K,
Kadota Sh. Caffeic acid phenethyl ester (CAPE): potent nitric oxide
inhibitor from the Netherlands propolis. Biol Pharm Bull 2003;26:487–91.
32. Usia T, Banskota AH, Tezuka Ya, Midorikawa K, Matsushige K, Kadota Sh.
Constituents of Chinese propolis and their antiproliferative activity. J Nat
33. Melliou E, Chinou I. Chemical analysis and antimicrobial activity of
Greek propolis. Planta Med 2004;70:515–9.
34. Banskota AH. Tezuka Y, Adnyana IK, Midorikawa K, Matsushige K,
Kadota S. Hepatoprotective and anti-Helicobacter pylori activities of
constituents from Brazilian propolis. Phytomedicine 2001;8:16–23.
35. Ito J, Chang F, Wang. H., Park YK, Ikegaki M, Kilgore N, Lee K. Anti-
AIDS agents. 48. Anti-HIV activity of moronic acid derivatives and the
new melliferone related triterpenoid isolated from Brazilian propolis.
J Nat Prod 2001;64:1278–81.
36. Kujumgiev A, Tsvetkova I, Serkedjieva Yu, Bankova V, Christov R, Popov S.
Antibacterial, antifungal and antiviral activity of propolis from different
geographic origins. J Ethnopharmacol 1999;64:235–40.
37. Popova M, Bankova V, Naydensky Ch, Tsvetkova I, Kujumgiev A.
Comparative study of the biological activity of propolis from different
geographic origin: a statistical approach. Macedonian Pharm Bull 2004;
38. Banksota AH, Tezuka Y, Adnyana IK et al. Cytotoxic, hepatoprotective
and free radical scavenging effects of propolis from Brazil, Peru, the
Netherlands and China. J Ethnopharmacol 2000;72:239–46.
39. Salomao K, Borba CM, Campos LC, Machado DG, Aquino Neto FR,
de Castro SL. Chemical composition and microbicidal activity of extracts
from Brazilian and Bulgarian propolis. Lett Appl Microbiol 2004;38:
40. Kumazawa Sh, Hamasaka T, Nakayama Ts. Antioxidant activity of
propolis of various geographic origins. Food Chem 2004;84:329–39.
41. Hamasaka T, Kumazawa Sh, Fujimoto T, Nakayama Ts. Antioxidant
activity and constituents of propolis collected in various areas of Japan.
Food Sci Technol Res 2004;10:86–92.
42. Chen Ch, Weng M., Wu Ch, Lin J. Comparison of radical scavenging
activity, cytotoxic effects and apoptosis induction in human melanoma
cells by Taiwanese propolis from different sources. eCAM 2004;1:175–85.
Received September 30, 2004; revised December 8, 2004;
accepted December 24, 2004
32 Recent trends in propolis research