Review Article

Is propolis safe as an alternative medicine?

Maria Graça Miguel, Maria Dulce Antunes1

Faculty of Sciences and ABSTRACT Technology, Department Propolis is a resinous substance produced by honeybees as defense against intruders. It has relevant therapeutic of Chemistry and properties that have been used since ancient times. Nowadays, propolis is of increasing importance as a Pharmacy, 1Faculty of Sciences and Technology, therapeutic, alone or included in many medicines and homeopathic products or in cosmetics. Propolis is Department of produced worldwide and honeybees use the flora surrounding their beehives for its production. Therefore Biological Sciences and its chemical composition may change according to the flora. The phenolic and volatile fractions of propolis Bioengineering, University have been revised in the present study, as well as some of the biological properties attributed to this natural of Algarve, IBB, Center product. An alert is given about the need to standardize this product, with quality control. This has already for Plant Biotechnology, been initiated by some authors, mainly in the propolis from the poplar-type. Only this product can constitute Building 8, Campus de a good complementary and alternative medicine under internationally acceptable quality control. Gambelas, 8005-139 Faro, Portugal

Address for correspondence: Dr. Graça Miguel, E-mail: [email protected]

Received : 06‑03‑11 Review completed : 10‑04‑11 Accepted : 20‑05‑11 KEY WORDS: Biological properties, phenols, propolis, volatiles

ropolis or bee glue (CAS number 9009-62-5) is a generic studies have revealed that propolis in honey bee colonies may P name for a resinous substance obtained from beehives, play a more subtle role in colony level immunity than direct used traditionally as an antimicrobial. It is a heterogeneous defense against parasites and pathogens.[2] mixture of many substances collected, transformed, and used by bees to seal holes in their honeycombs, smooth out the Bees make use of the mechanical properties of propolis and internal walls, and protect the entrance against intruders.[1] In of its biological action. This action against microorganisms fact, propolis means ‘defense of the city’. The word propolis has also been used by human beings since ancient times.[4] is derived from the Greek words ‘pro’ (meaning ‘in front of”), Nevertheless, other biological properties have been attributed to and ‘polis’ (meaning ‘the city’). In this manner, propolis might this natural product: hepatoprotective, antitumor, antioxidative, serve as a means for colonies to better maintain homeostasis and anti-inflammatory.[1,5-8] In fact, propolis, along with other of the nest environment through the reduction of microbial honeybee products such as honey, royal jelly, and pollen, have growth on hive walls, prevention of uncontrolled airflow into relevant therapeutic properties, being used since 300 years B.C. the nest, waterproofing of walls against external moisture, and in folk medicine worldwide.[9] Research regarding the chemical protection against invaders.[2] Therefore, propolis could simply composition of propolis of diverse origins, their pharmacological mean defense of the hive.[3] However, more recently, some properties, and relation between its composition and biological activities has increased in the last few years.[10] This product Access this article online has, therefore, gained popularity as an alternative medicine for [11] Quick Response Code: health amelioration and disease prevention. Some examples Website: include its utilization for increasing the body’s natural resistance www.jpbsonline.org to infections and lowering blood pressure and cholesterol levels. It has been also used in mouthwash products and toothpastes DOI: to prevent caries and treat gingivitis and stomatitis,[12] in cough 10.4103/0975-7406.90101 syrups, oral pills, lozenges, ointments, lotions, and vitamins.[13] Holistic therapists often use propolis for the relief of some

How to cite this article: Miguel MG, Antunes MD. Is propolis safe as an alternative medicine?. J Pharm Bioall Sci 2011;3:479-95.

Journal of Pharmacy and Bioallied Sciences October-December 2011 Vol 3 Issue 4 479   Miguel and Antunes: Propolis and health inflammations, viral diseases, fungal infections, ulcers, and is Populus spp. of section Aigeiros, most often P. nigra. (2) superficial burns along with acupuncture, ayurveda, and Birch propolis from Russia, for which the plant source is Betula homeopathy.[13] verrucosa Ehrh.; (3) Green propolis from Brazil, mainly from Baccharis spp., predominantly B. dracunculifolia DC.; (4) Red Among the chemical substances found in propolis are propolis from Cuba and Venezuela, from Clusia spp.; (5) ‘Pacific’ waxes, resins, balsams, aromatic and ethereal oils, pollen, propolis from Okinawa and Taiwan, for which the plant source and other organic matter.[14] Typically propolis is comprised is unidentified yet; and (6) ‘Canarian’ propolis from Canary of resin (50%), which is composed of flavonoids and Islands, for which the plant source is also unknown [Table 1]. related phenolic acids, generally called as the polyphenolic fraction, waxes (30%), essential oil (10%), pollen (5%), The main biologically active substances in these types of and other organic compounds (5%).[15,16] However, this propolis are different. Flavones, flavanones, phenolic acids, and chemical composition may change according to the their esters predominate in the poplar propolis [Figure 1], while plant source, which is related to the regional vegetation flavones and flavonols dominate in the birch propolis [Figure 2]. and to the season in which it is collected by the bees.[17] In green and red propolis, prenylated p-coumaric acids, Therefore, the standardization of this product is difficult diterpenic acids [Figure 3], and polyprenylated benzophenones on account of the inherent difficulties associated with the [Figure 4] dominate. C-prenylflavanones [Figure 5] and analysis of complex mixtures from different vegetal sources.[18] furofuran lignans [Figure 6] predominate in the ‘Pacific’ and ‘Canarian’ propolis, respectively.[20] However, particularly due to The present study is focused on the chemical variability of the the richness of the flora in some countries, different types of phenols and volatiles of propolis and its repercussion in the propolis have been reported for the same region. biological activities.

Flavones and flavonols Chemical Variability of the Phenols OH O OH O Propolis cannot be used as a crude material. It must be purified OH by extraction with adequate solvents, to remove the unwanted OH O OH O material, preserving the active components, for example, the Chrysin Galangin polyphenolic fractions. Several solvents have been used such as water, ethanol, methanol, hexane, acetone, and chloroform, with Flavanones and dihydroflavonols ethanol being the most common, particularly at concentration [12,19] of 70%. OH O OH O OH O

OH OCOCH3 The compounds present in the propolis resin have three OH O OH O OH O origins: substances actively secreted by plants and substances Pinocembrin Pinobanksin Pinobanksin-3- O-acetate exuded from wounds in plants (lipophilic materials on leaves and leaf buds, resins, mucilage, gums, lattices, among Caffeate esters others) collected by bees, secreted substances from bee O O OH OH metabolism, and materials which are introduced during O O [10,20] propolis elaboration. As such compounds have a plant OH OH origin, the composition of the plant source determines the chemical composition of propolis. 3-Methyl-2-butenyl caffeate 2-Methyl-2-butenyl caffeate

The chemical composition of propolis depends on the local OH OH [4,10] O O floral at the site of collection, being therefore, highly variable. OH OH In spite of this diversity and according to the most studied O O propolis types, Bankova[20] cited at least six main chemical types Phenylethyl caffeate Benzyl caffeate of propolis: (1) Poplar propolis found in Europe, North America, Figure 1: Some chemical compounds detected in poplar propolis and the non-tropical regions of Asia, for which the plant source from Europe

Table 1: Types of propolis, their origins and chemical compositions (Adapted from[20]) Zone of the world Type of compounds Plant source Figures Europe Flavones and flavonols Populus nigra L. 1 Flavanones, and dihydroflavonols Russia Flavones and flavonols Betula verrucosa Ehrh 2 Brazil (‘Green propolis’) Prenylated p-coumaric acids Baccharis dracunculifolia DC 3 Cuba and Venezuela (‘Red propolis’) Diterpen acids Clusia spp. 4 Okinawa and Taiwan (‘Pacific’ propolis) C-prenylflavanones Unknown 5 Canary Islands (‘Canarian propolis’) Furofuran lignans Unknown 6

 480 Journal of Pharmacy and Bioallied Sciences October-December 2011 Vol 3 Issue 4 Miguel and Antunes: Propolis and health 

Phenolic compounds Flavones

OCH3 OH

O OH O OH O OH O COOH COOH O

2,2-Dimethyl-8-prenylchromene 4-Hydroxy-3,5-diprenyl cinnamic acid (artepillin C) 3-Prenyl cinnamic acid allyl ester OH O OH O OCH3 O Acacetin Apigenin

OH O O O O -Methylated flavonol and Flavonol O H3CO OH OH OH O O OCH3 OH KaempferidePropolisbenzofuranA

OH O H3CO O

Terpenoid compounds

OCH3 OH

CH2OH OH O OH O H COH ErmaninRhamnocitrin COOH

OH OCH3 HOOC Isocupressic acid (labdane diterpenoid) 13-Symphyoreticulic acid (clerodane diterpenoid) Farnesol (sesquiterpenoid) OH O

H3CO OH OH O OH O

Kaempferide [ ] n O

Esters of long chain fatty acids (3-hydroxystearic acid n=11, procrim a; 3-hydroxystearic acid n=13, procrim b) and Triterpenóide lupeol, a pentacyclic triterpenoid Figure 3: Some chemical compounds detected in green propolis from Brazil

CH OCOCH 2 3 C-Prenylated flavanones OH OH OH OH

OH OH O OH O Acetoxi-betulenol

OH O OH O

Figure 2: Some chemical compounds detected in birch propolis from Isonymphaeol-B or propolin FNymphaeol-A or propol in C Russia

OH OH OH OH

Polyprenylated benzophenone derivatives OH O OH O

OH O OH O Nimphaeol -B or propol in DNymphaeol-C

O O OO OO OH OH OH OH OH

OH O OH O O O HOO OOH OH

OH O OH O PropoloneA Nemorosone (isomers) Propol in APropol in B OH

O O

OH HOO O OH

OH OH OH O O O

OH OH Guttiferone EXanthochymol OH O Propolin E

Figure 4: Some chemical compounds detected in red propolis from Figure 5: Some C-prenylated flavanones detected in propolis from Cuba and Venezuela Okinawa and Taiwan (‘Pacific’ propolis)

Journal of Pharmacy and Bioallied Sciences October-December 2011 Vol 3 Issue 4 481   Miguel and Antunes: Propolis and health

Lignans AcO

AcO OH O OCH3 OCH 3 3-Acetoxymethyl-5[( E)-2-formylethen-1-yl]2-(4-hydroxy-3-methoxyphenyl-7-methoxi-2,3-dihydrobenzofuran

OCH3 O O O O O O

O H CO O 3 O

O H3CO

OCH3 Sesamine Sesartenin

Figure 6: Some lignans detected in propolis from Canary Islands

Examples of diterpenes OH CH3COO CH2OH

CHO COOH 13 -Manool Communal (1)Copal ol (1)Communicacid(1,2) epi OH OH OH OH OH OR OR

O O CHO COOH CH2OH COOH 1 R OCH 3 1 R OCH3 13-epi-Torulosal Pimaricacid(1) 13-epi-Torulosol 13- epi-Cupressic acid (1)

OH CH OH CH2OH CH2OH 2 O O O 2 R OH 2 R OH COOH CHO CH2OH Isogatholal (1)Agathadiol(1) Totarolon(1) Isocupressicacid(1,2) Figure 8: Terpenyl esters in Maltese propolis: 1. 2-acetoxy-6-p-

COOH CH2OAc methoxybenzoyl jaeschkeanadiol; 2. 2- acetoxy-6-p-hydroxybenzoyl jaeschkeanadiol; 3. ferutinin (ferutinol p-hydroxybenzoate); 4. teferin (ferutinol vanillate) COOH COOH CH2OOCH2CH2COOH

Junicedric acid Acetylisocupressicacid(2) 18-Succinyloxyabieta-8,11,13-triene also be found in Croatia and Malta [Table 2 and Figure 7].[21-24] O The botanical origin of diterpenes in the propolis samples from

COOH Greece is yet unidentified, but on the basis of the diterpenic 14,15-dinor-13-oxo-8(17)-labden-19-oic acid profile, the source plant should be the conifer species of the Figure 7: Some diterpenes detected in propolis of some regions of Cupressaceae family, in which the flora of the region is very Greece, Malta (1) and Sicilia (2) rich.[21] In addition, in Malta, there are other propolis samples also, wherein another compound group is observed: mono- and Propolis from Europe, China, and North America sesquiterpenyl esters of substituted benzoic acids [Table 2 and Figure 8],[24] which are attributed to a genus Ferula plant found In Europe, China, and North America, propolis is generally in this region. Mono- and sesquiterpene esters of benzoic acids considered to be of the poplar-type; nevertheless, several authors have also been found in propolis samples from Iran, along have demonstrated that other types can be found, owing to the with others in which flavonoids and caffeate ester compounds flora specificity of each region. In Europe, the Mediterranean predominate.[25,26] Such results may be due to the simultaneous propolis from diverse places in Greece contains mainly diterpenes presence of Populus spp. and Ferula spp. and almost no phenolics.[21] Such a profile has been described for propolis from Sicily and northwestern Greece,[22,23] and can In another study, Kalogeropoulos et al.,[27] also reported that

 482 Journal of Pharmacy and Bioallied Sciences October-December 2011 Vol 3 Issue 4 Miguel and Antunes: Propolis and health  the Greek (mainland Greece and Greek islands) and east Adana (Central Anatolia), in which the poplar type compounds Cypriot propolis samples shared characteristics that were were present in lower amounts, the main ones being cinnamyl different from typical European propolis, because those cinnamate and diterpenic acids (dihydroabietic, abietic, and samples presented anthraquinones (mainly emodin and isopimaric); a sample from Eastern Anatolia that presented chrysophanol) [Table 2 and Figure 9] and terpenes and/or significant amounts of hydroxyl fatty acids (hydroxypalmitic, flavonoids in significant amounts, and had low abundance hydroxystearic) and triterpenoid alcohols besides the poplar of phenolic acids and their esters. phenolics in lower amounts; and finally, also from Eastern Anatolia, a sample with low concentrations of flavonoids and In Cretan propolis, five new terpenes were isolated: the diterpenes 14,15-di(nor)-13-oxo-8(17)-labden-19-oic acid and Table 2: Some components present in the Mediterranean a mixture of labda-8(17),13E-dien-19-carboxy-15-yl oleate propolis and palmitate, as well as the triterpenes, 3,4-seco-cycloart-12- hydroxy-4(28),24-dien-3-oic acid and cycloart-3,7-dihydroxy- Country or Island Components Figure References 24-en-28-oic acid [Table 2 and Figure 10].[28] These cycloartane Greece mainland Diterpens 7 [21] [21] triterpenes were isolated only from Brazilian propolis (tropical and crete 13-epi-Manool 7 Communal 7 [21] type), Mangifera indica (Anacardiaceae) being the plant source 14,15-dinor-13-oxo-8(17-labden- 7 [21] [29] [Table 3 and Figure 11]. 19-oic acid Copalol 7 [21] The main sources of phenolic compounds found in Turkish Communic acid 7 [21] propolis were the poplar bud exudates. Nevertheless some 13-epi-Torulosal 7 [21] [21] authors were able to classify the samples collected in different Pimaric acid 7 13-epi-Torulosol 7 [21] regions of Turkey in four main groups: the typical poplar 13-epi-Cupressic acid 7 [21] samples from Central and Western Anatolia; a sample from Isogatholal 7 [21] Agathadiol 7 [21] Totarolon 7 [21] OH O OH OH O OH Isocupressic acid 7 [21] Junicedric acid 7 [21] Acetylisocupressic acid 7 [21] [21] OH 18-Succiniloxyabieta-8,11,13- 7 triene O O 14,15-dinor-13-oxo-8(17)- 7 [21] labden-19-oic acid Chrysophanol Emodin Malta Communal 7 [24] [24] Antraquinones found in some propolis from East Cypriot, Copalol 7 Figure 9: [24] mainland Greece, and Greek Islands Pimaric acid 7 Cycloartane triterpenes 13-epi-Cupressic acid 7 [24] Isogatholal 7 [24] Agathadiol 7 [24] Totarolon 7 [24] Isocupressic acid 7 [24] Sicilia Communic acid 7 [22] [22] H Isocupressic acid 7 Acetylisocupressic acid 7 [22] Malta Sesquiterpenyl esters of substituted benzoic acids OH OH 2-Acetoxy-6-p-methoxybenzoyl 8 [24] COOH jaeschkeanadiol 2-Acetoxy-6-p-hydroxybenzoyl 8 [24] Cycloart -3 ,7-dihydroxy-24-en-28-oic acid jaeschkeanadiol Ferutinin (ferutinol 8 [24] p-hydroxybenzoate) [24] OH Teferin (ferutinol vanillate) 8 Greece mainland Antraquinones and crete H Chrysophanol 9 [27] HOOC Emodin 9 [27] Cyprus Antraquinones Chrysophanol 9 [27] Emodin 9 [27] Crete Triterpenes 3,4-seco-Cycloart-12-hydroxy- 10 [28] 3,4-seco-cycloart-12-hydroxy4(28),24-dien-3-oicacid 4(28),24-dien-3-oic acid Cycloart-3,7-dihydroxy-24-en- 10 [28] 28-oic acid Figure 10: Cycloartane triterpenoids found in some Cretan propolis

Journal of Pharmacy and Bioallied Sciences October-December 2011 Vol 3 Issue 4 483   Miguel and Antunes: Propolis and health

Table 3: Some components present in green and red propolis Cycloartane triterpenoids from Brazil COOH COOH COOH Country Components Figure References H H H

[29] H OH Brazil Green propolis 3 O OH H H H H Phenolic comounds IsomangiferolicacidMangiferolicacidMangiferonicacid [29] 2,2-Dimethyl-8-prenylchromone 3 O O O 4-Hydroxy-3,5-diprenyl cinnamic acid 3 [29] CH2OH COOH COOH (artepillin C) H H H 3-Prenylcinnamic acid ally ester 3 [29] H OH Kaempferide 3 [29] O OH H H H H Propolis benzofuran A 3 [29] 24-Methylen-cycloartan-3,26 diol Ambolic acid Ambonic acid Terpenoid components Isocupressic acid 3 [29] Figure 11: Some cycloartane triterpenes isolated only from Brazilian 13-Symphyoreticulic acid 3 [29] propolis (tropical propolis) Farnesol 3 [29] Esters of long chain fatty acids and 3 [29] OH lupeol (procrim a and procrim b) OH O [29] OH O H3CO O O Isomangiferolic acid 3 OCH3 Mangiferolic acid 3 [29] OH OH OCH O OCH O OCH O OCH O Mangiferonic acid 3 [29] 3 3 3 3 Chrysin-5-methyl ether Kaempferol-5-methyl ether Hesperitin-5,7-dimethyl ether 5-Methoxy-3-hydroxy-flavanone [29] 24-Methylen-cycloartan-3β,26 diol 3 OH Ambolic acid 3 [29] [29] Ambonic acid 3 OH O Red propolis OCOC4H9 OO Flavonoids OCH 3O [54] (2S)-7-Hydroxyflavanone 17 Pinobanksin-5-methyl-ether-3- O-pentanoate [54] (2S)-Liquiritigenin 17 OH p-Coumaric ester derivative ester (2S)-7-Hydroxy-6-methoxyflavanone 17 [54] (2S)-Naringenin 17 [54] OH (2S)-Dihydrobaicalein 17 [54] [54] (2S)-Dihydrooroxylin A 17 OO (2R,3S)-3,7-Dihydroxyflavanone 17 [54] Garbanzol 17 [54] (2R,3S)-3,7-Dihydroxy-6- 17 [54] methoxyflavanone OH Alnusin 17 [54] Figure 12: Some compounds found in propolis from Portugal Daidzein 17 [54] Formononetin 17 [54] OH COOH COOH H CO COOH Calycosin 17 [54] 3 H3CO COOH [54] OH OH OH Xenognosin B 17 H3CO [54] Biochanin B 17 Caffeicacid p-C oumaric acid Ferulicacid3,4-Dimethoxycinnamicacid [54] O Pratensein 17 O COOH OH OH 2’-Hydroxybiochanin A 17 [54] O O (3S)-Vestitone 17 [54] OH OH (3S)-Violanone 17 [54] Cinnamyl caffeate Caffeicacid phenethylester Cynnamylideneaceticacid (3S)-Ferrerin 17 [54] (3R)-4’-Methoxy-2’,3,7- 17 [54] OH O OH O OH O OH O

trihydroxyisoflavanone OH OH OCOCH 3 OH O OH O (3S)-Vestitol 17 [54] OCH3O OH O (3S)-Isovestitol 17 [54] 5-Pinobanksin-5-methyl etherPinobanksinPinocembrin Pinobanksin-3-acetate (3S)-7-O-Methylvestitol 17 [54] OH O OH O (3S)-Mucronulatol 17 [54] H3CO O Pterocarpans OH (6aS,11aS)-Medicarpin 17 [54] O O O (6aS,11aS)-3,10-Dihydroxy-9- 17 [54] ChrysinGalanginTectochrysin methoxypterocarpan Figure 13: Structures of some constituents of propolis from China (6aR,11aR)-3-Hydroxy-8,9- 17 [54] dimethoxypterocarpan (6aS,11aS)-6a-Ethoxymedicarpin 17 [54] very high levels of p-coumaric and ferulic acids, as well as a series (6aR,11aR)-3,4-Dihydroxy-9- 17 [54] of phenolic glycerides.[30] methoxypterocarpan [54] (6aR,11aR)-4-Methoxymedicarpin 17 In recent times, some authors found in the propolis samples Neoflavonoid (7S)-Dalbergiphenol 17 [54] from North Portugal, phenolic acids and their esterified and / Lignans 17 [54] or methylated derivatives, as well as dihydroflavonols, flavones, (+)-Pinoresinol dimethyl ether 17 [54] flavanones and flavonols, either as free from or with methylated/ [54] (+)-Pinoresinol 17 esterified forms, similar to those of the European-type. (+)-Syringaresinol 17 [54] Nevertheless, new compounds could also be found: methylated

 484 Journal of Pharmacy and Bioallied Sciences October-December 2011 Vol 3 Issue 4 Miguel and Antunes: Propolis and health 

OH O

OH OH

OH

OCH3

8-[(E)-4-phenylprop-2-en-1-one](2R,3S)-2-(3,5-dihydroxyphenyl)-3,4-dihydro-2H-2-benzopyran-5-methoxyl-3,7-diol

OH O

OH OH

OH

OCH3

8-[(E)-4-phenylprop-2-en-1-one](2S,3R)-2-(3,5-dihydroxyphenyl)-3,4-dihydro-2H-2-benzopyran-5-methoxyl-3,7-diol

OCH3 O

OH OH

OH

OCH3

Racemate:

8[(E)-4-phenylprop-2-en-1-one]-(2R,3S)-2(3-methoxyl-4-hydroxyphenyl-3,4-dihydro-2H-2-benzpyran-5-methoxyl-3,7-diol 8[(E)-4-phenylprop-2-en-1-one]-(2S,3R)-2(3-methoxyl-4-hydroxyphenyl-3,4-dihydro-2H-2-benzpyran-5-methoxyl-3,7-diol OH OH

H3CO

OH OH Rhamnetin Figure 14: New compounds found in propolis from China (cytotoxic)

OH Table 4: Some new components of propolis from North of Portugal H CO O OH 3 OOO OOO Country Components Figure References

O OH Portugal Flavonoids and derivatives [31] O OH O Chrysin-5-methyl ether 12 O Kaempferol-5-methyl ether 12 [31] [31] 4'-Methoxy-bavachromanol LaserpitinIsolaserpitin Hesperitin-5,7-dimethyl ether 12 Pinobanksin-5-methyl ether-3-O-pentanoate 12 [31] Figure 15: Some new compounds isolated from propolis of Korea 5-Methoxy-3-hydroxy-flavanone 12 [31] 12 [31] p-coumaric ester derivative dimmer and/or esterified or hydroxylated derivatives of common poplar p-coumaric ester derivative dimmer 12 [31] flavonoids as well as a p-coumaric ester derivative dimmer [Table 4 and Figure 12].[31] Jeju, a southern island of Korea with a subtropical climate, a new With regard to China, the propolis of this region is also of chalcone 4’-methoxy-bavachromanol as well as new khellactone the poplar-type [Table 5 and Figure 13].[20,32-35] However in derivatives (laserpitin and isolaserpitin) were found [Table 6 the Chinese propolis some authors also have described some and Figure 15].[38] new compounds that had not yet been referred [Table 5 and Figure 14].[33,35] Continuing in the Asian Continent, propolis In the same continent, and for the first time, open-chain samples from Myanmar have cycloartane-type triterpenoids neoflavonoids were identified in propolis samples from Nepal, and prenylated flavanones, practically dissimilar from those of as also new chalcone, flavanone, flavan-3-ol, isoflavonoids, and its neighboring China.[36] flavanonol compounds [Table 7 and Figure 16].[39-41]

In Korea, studies performed by some authors allowed to classify Bud exudates of poplar trees are the main source of propolis the propolis from several places of this country into poplar-types, in the temperate zones and chemical data show a clear such as those of Europe and China.[37] Also, for the first time in preference for the Populus species belonging to the section

Journal of Pharmacy and Bioallied Sciences October-December 2011 Vol 3 Issue 4 485   Miguel and Antunes: Propolis and health

Open-chain neoflavonoids Table 5: Some components present in Chinese propolis

OCH3 H3CO O H CO O 3 H3CO O O H3CO OH Country Components Figure References

OH OH OH OH OH China Phenolic acids and derivatives H H H H H H HH Caffeic acid 13 [20,32] p-Coumaric acid 13 [20,32] Ferulic acid 13 [20,32] OH O O [20,32] H3CO O HOO H OH 3,4-Dimethoxycinnamic acid 13 [20,32] OH OH Cinnamylideneacetic acid 13 H H OH H H H Caffeic acid phenethyl ester 13 [20,32] Cynnamil caffeate 13 [20,32] Flavonoids H CO O [20,32] 3 H3CO O Pinobanksin-5-methyl ether 13 [20,32] O O Pinobanksin 13 Chrysin 13 [20,32] [20,32] OH OH Pinocembrin 13 OH Galangin 13 [20,32] (S)-3'-Hydroxy-4-methoxydalberg ione (S)-3',4'-dihydroxy-4-methoxydalbergione Pinobanksin-3-acetate 13 [20,32] Flavanone Tectochrysin 13 [20,32] OH 8-[(E)-4-phenylprop-2-en-1-one](2R,3S)- 14 [33,35] H CO O 3 OH 2-(3,5-dihydroxyphenyl)-3,4-dihydro-2H-2-

OH benzopyran-5-methoxyl-3,7-diol O Racemate of 8-[(E)-4-phenylprop- 14 [33,35] (2R)-3'4',6-trihydroxy-7-methoxy flavanone 2-en-1-one](2R,3S)-2-(3methoxyil- Isoflavonoids 4-hydroxyphenyl-3,4-dihydro-2H-2-

OH O OH O OH O benzopyran-5-methoxyl-3,7-diol [33,35] OH OH OH Rhamnetin 14 OH H3CO O O O OCH 3 OH OCH3 OCH 3 Figure 16: Some new compounds belonging to a diverse group of Table 6: Some components present in Korean propolis compounds isolated from propolis of Nepal Country Components Figure References Korea Chalcone [38] Aigeiros, as is seen in Europe and North America.[42] In fact, 4’-Methoxy-bavachromanol 15 Coumarin in North America, mainly in the propolis from Ontario Laserpitin 15 [38] (Canada), most studies demonstrate that propolis also Isolaserpitin 15 [38] originates from the Populus section Aigeiros. Nevertheless, the compounds found in the propolis from Canada in which Table 7: Some components present in propolis from Nepal Aigeiros poplars are absent (Boreal forest and Pacific Coastal forest regions), are different: in Victoria, the components Country Components Figure References found in propolis are dihydrochalcones, considered to be Nepal Open chain neoflavonoids (S) -3’-Hydroxy-4-methoxydalbergione 16 [39,40,41] characteristic of bud exudates of poplars from Populus (S) -3’,4’-dihydroxy-4-methoxydalbergione 16 [39,40,41] section Tacamahaca and in the Richmond region, are found [39,40,41] large amounts of p-coumaric and cinnamic acids, typical of Flavonoids 16 [39,40,41] [39,40,41] poplars of section Leuce.[43] Isoflavonoids 16

However, in South America, concretely the Uruguayan propolis Propolis from Brazil components were similar to those from Europe and China, suggesting that there is a similar plant origin in the Uruguayan The Brazilian propolis represents 10 – 15% of the worldwide [44] propolis also. production, Brazil being the third world producer, behind Russia and China.[46] Among the types produced in Brazil, Propolis from Russia green propolis (from greenish-yellow to deep green) prevails, gaining preference in the world propolis market.[47] Prenylated Significant amounts of phenolic glycerides such as dicoumaroyl derivatives of p-coumaric, artepillin C (4-hydroxy-3,5-diprenyl acetyl glycerol, diferuloyl acetyl glycerol, feruloyl coumaroyl cinnamic acid), dupranin (4-hydroxy-3-prenyl cinnamic acid), acetyl glycerol, and caffeoyl coumaroyl acetyl glycerol have (E)-3-prenyl-4-(dihydroxicinnamoyloxy)-cinnamic acid, and been isolated from the propolis obtained in Northern Russia.[42] diterpenic acids were detected in green propolis as well as in These compounds are attributed to Populus tremula, in which the buds of Baccharis dracunculifolia, an Asteraceae family from the exudates are rich in such components. Birch propolis (Betula southeast and western-central Brazil.[11,18,47,48] verrucosa Ehrh) predominates in Russia and its main biologically active substances are flavones and flavonols, other than those Nevertheless, other types of Brazilian propolis have been reported for European propolis.[20,42,45] reported. For example, a new type of red propolis, in which

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Flavanones isoflavonoids[49,50] instead of prenylated p-coumaric acids R 4 [51-53] OH O predominated has been reported. The red propolis is

R3 R1 mainly collected in north east Brazil where Baccharis spp are R2 O practically absent, in this case, Dalbergia ecastophyllum (L.) R R R R Name 1 2 3 4 Taub. (Leguminosae) is the main source of isoflavonoids.[49,50] H 2 HHH(2S)-7-Hydroxyflavanone

H 2 HHOH (2S)-Liquiritigenin Flavonoids, flavonol, isoflavones, isoflavanones, isoflavans, H 2 HOCH3 HH (2S)-7-Hydroxy-6-methoxyflavanone chalcones, auronol, pterocarpans, 2-arylbenzofuran, H 2 OH HOH(2S)-Naringenin neoflavonoids, and lignans in the red propolis from Brazil H 2 OH OH H(2S)-Dihydrobaicalein

H 2 OH OCH3 H(2S)-Dihydrooroxylin A have also been referred by Awale et al. to possess cytotoxic [54] β OH HHH(2R,3S)-3,7-Dihydroxyflavanone activities [Table 3 and Figure 17]. β OH HHOH Garbanzol

β OH HOCH3 H(2R,3R)-3,7-dihydroxy-6-methoxyflavanone β OH OH OCH3 HAlnustinol Red propolis from Cuba and Venezuela Flavonol

OH O Red propolis from Cuba was reported to be particularly rich

H3CO OH in polyprenylated benzophenones (propolone A, nemorosone, O

Isoflavones guttiferone, E, xanthochymol, propolones B-D, garcinielliptone [55-58] OH O I, and hyperibone), due to the presence of Clusia spp. R4 R5 Nevertheless, the chemical composition of other propolis

R2 O R6 samples from the same country did not present prenylated R2 R 4 R5 R6 Name

HHHOHDaidzein benzophenones. In the samples from Pinar del Rio province, [59] HHHOCH3 Formononetin isoflavonoids predominated. Of late, some authors classified HHOH OCH3 Calycosin the Cuban propolis into three types: (1) Brown Cuban propolis HOHH OCH3 Xenognosin B mainly comprised of polyisoprenylated benzophenone derivatives; OH HHOCH3 BiochaninA

OH HOHOCH3 Pratensein (2) Red Cuban propolis, characterized by the presence of 1 OH OH HOCH3 2'-HydroxybiochaninA isoflavonoids; and (3) Yellow Cuban propolis, in which H- and 13C-NMR spectral data suggested the presence of aliphatic Isoflavanones OH O R compounds (terpenoids and sterols) as the main constituents, R1 4 R [60] 5 but these compounds were not identified. Only recently, R2 O OCH 3 and by GC-MS, the yellow Cuban propolis, collected from R R R R Name 1 2 4 5 different regions of Cuba, was classified into two main types: β -H HOHH (3S)-Vestitone

β -H HOCH 3 OH (3S)-Violanone type A, rich in triterpenic alcohols [Table 8 and Figure 18] with β -H OH OH H(3S)-Ferreirin polymethoxylated flavonoids [Table 8 and Figure 19] as minor a -OHOHOHH (3R)-4'-Methoxy-2',3,7-trihydroxyisoflavanone compounds, and type B, containing acetyl triterpenes as the main Isoflavans constituents.[61] According to these authors the botanical sources R3 O R4 R5 of the two types of yellow Cuban propolis were not yet identified.

R6 R R R R Name 3 4 5 6 Some authors suggested that Clusia minor and C. major OH OH HOCH3 (3S)-Vestitol (Guttiferae) were the source of the main phenolics present in OH OCH3 HOH(3S)-Isovestitol

OCH 3 OH HOCH3 (3S)-7-O-Methylvestitol the propolis from Venezuela. Their resins exuded polyprenylated

OH OCH3 OH OCH3 (3S)-Mucronulatol benzophenones as found in propolis. Only few samples presented flavonoids and in each case they were found to R1 Pterocarpans OH O be lipophilic methylated 6-oxygenated flavones (eupatorin, R2 [62] [63] R 5 O R3 hispidulin, among others). Later on, Trusheva et al. found

OCH R4 3 two new polyisoprenylated benzophenones: 18-ethyloxy-17- R R R R R Name 1 2 3 4 5 hydroxy-17,18-dihydroscrobiculatone A and 18-ethyloxy- H-β HH H-β H(6aS,11aS)-medicarpin

H-β HH OH β -H (6aS,11aS)-3,10-dihydroxy-9-methoxypterocarpan 17-hydroxy-17,18-dihydroscrobiculatone B, together with a H-a HOCH3 H-H(6aR,11a R)-3-hydroxy-8,9-dimethoxypterocarpan the known scrobiculatones A and B, in the propolis samples H-a OEtH H-a H(6aS,11aS)-6a-ethoxymedicarpin from Venezuela [Table 9 and Figures 4 and 20]. These OH a -H HHa -H (6aR,11a R)-3,4-dihydroxy-9-methoxypterocarpan a a OCH3 -H HH -H (6aR,11a R)-4-methoxymedicarpin polyisoprenylated benzophenones were only found in the floral resin of C. scrobiculata. According to the authors, this plant Neoflavonoid Lignans H3CO material was used by honeybees as a source for propolis, in the H CO OCH 3 3 O R 1 R tropical rain forest of the Truhillo state in Venezuela. OH HH2 R2 O R1

OCH (7S)- Dalbergiphenol 3 In the red-type Mexican propolis, samples of flavanones, R R Name 1 2 isoflavans, and pterocarpans were found to be in agreement with OCH3 H(+)-Pinoresinol dimethyl ether

OH H(+) Pinoresinol the chemical profiles of the Cuban and Brazilian red propolis,

OH OCH3 (+)Syringaresinol but three new isoflavonoids were also found along with the Figure 17: Some compounds present in red propolis from Brazil presence of compounds with a 1,3-diaryl-propane and 1,3-diaryl

Journal of Pharmacy and Bioallied Sciences October-December 2011 Vol 3 Issue 4 487   Miguel and Antunes: Propolis and health

Triterpenes alcohols Polymethoxylated flavones

O

H H H

H H O OH OH O H H H Compound A-ring B-ring

Lanosterol β-Amyrin-β Am yrone I -OCH3 (2)-OCH3, OH

III -OCH3 (2), OH -OCH 3, OH H IV -OCH3 (2), OH -OCH 3 (4) H H

H H H V -OCH3 (2)-OCH 3 (2), OH OH OH O H H H VIII -OCH3 (2), OH -OCH 3 (2), OH α -Amyrin-α Am yroneGermanicol Polymethoxylated flavanones

H H O

OH OH H O Germanicol Cycloartenol Compound A-ring B-ring Figure 18: Some triterpenic alcohols and derivatives found in yellow II -OCH (2), OH -OCH , (3) propolis from Cuba 3 3

VI -OCH3 (2), OH -OCH3, OH

Polyisoprenylated benzophenones VI I -OCH3 (2), OH -OCH3 (4) Figure 19: Some polymethoxylated flavonoids found in yellow propolis O O O O from Cuba

OO O O

H H

OCH 3

ScrobiculatonesA Scro biculatonesB

OH OCH 3 OH OCH CH OCH2CH3 2 3 OH OH OH OH (3',4'-dihydroxy-2'-methoxyphenyl)-3-(phenyl)propane ( Z)-1-(2'-methoxy-4',5'-dihydroxyphenyl)-2(3-phenyl)propene O O O O

OO O O O OH O OH O

H H H3CO OH OCH 3 O OH O 3-Hydroxy-5,6-dimethoxyf lavan7-H ydro xyflavanone Pinocembrin 18-Ethyloxy-17-h ydroxy-17,18-dihydroscrobiculatone A 18-Ethyloxy-17-hydroxy-17,18-dihydroscrobiculatoneB

OH O OH O OH O Figure 20: Polyisoprenylated benzophenones found in propolis from OCH 3 OH OH Venezuela OH OH

OCH3 OCH3 OCH 3 Mucronulatol Arizonicanol AVestitol propene carbon skeleton, which was related to the presence of OH OH [64] H CO O Dalbergia genus [Table 10 and Figure 21]. H3CO O 3

O ‘Pacific’ propolis from Okinawa and Taiwan O OCH 3 OH OCH3 Melilotocarpan AMelilotocarp an D In the ‘Pacific’ propolis from Okinawa and Taiwan, the structures Figure 21: Some components present in red-type Mexican propolis of two prenylflavanones were elucidated by the NMR spectral technique in the Taiwanese propolis. They were reported for to the European and Chinese ones (European-type). However, the first time and called propolins.[65] Further studies allowed [70,71] the finding of other similar structures within the propolin group in some places in Japan, some variability was reported. in the Taiwanese propolis, whose concentrations were closely These authors considered such diversity was due to the great dependent on the locations and collection seasons [Table 11 diversity of climate and vegetation in the islands of Japan, which and Figure 5].[66,67] These propolins (prenylated flavonoids) had extended from north (Hokkaido) to south (Okinawa). This Macaranga tanarius as the plant source.[68] place was located in the subtropical zone and the vegetation was quite different from the other areas.[70] Studies conducted Fujimoto et al.,[69] analyzing several propolis samples from all by some authors[72] revealed that not all propolis from the over the world, considered that samples from Japan were similar Okayama Prefecture (central Japan) considered as ‘poplar type’

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Table 8: Some components present in yellow propolis from Table 10: Some components present in red propolis from Cuba Mexico Country Components Figure References Country Components Figure References Cuba Triterpenoids Mexico 1,3-diaryl-proa(e)ne Lanosterol 18 [61] 3’,4’-Dihydroxy-2’-methoxyphenyl)-3- 21 [64] b-Amyrone 18 [61] (phenyl)propane b-Amyrin 18 [61] (Z)-1-(2’-Methoxy-4’,5’-dihydroxyphenyl) 21 [64] Germanicol 18 [61] -2(3-phenyl)propene a-Amyrone 18 [61] Flavonoids a-Amyrin 18 [61] 3-Hydroxy-5,6-Dimethoxyflavan 21 [64] Lupeol 18 [61] 7-Hydroxyflavanone 21 [64] Cycloartenol Pinocembrin 21 [64] Lanosterol acetate Isoflavonoids b-Amyrin acetate Mucronulatol 21 [64] Polymethoxylated flavones 18 [61] Vestitol 21 [64] Polymethoxylated flavanones 18 [61] Arizonicanol A 21 [64] Pterocarpans Melilotocarpan A 21 [64] Melilotocarpan D 21 [64] Table 9: Some components present in red propolis from Cuba and Venezuela Country Components Figure References Table 11: Some components present in propolis from Japan Cuba and Polyisoprenylated benzophenones (Okinawa) and Taiwan Venezuela Propolone A 4 [20] Country Components Figure References Nemorosone A isomers 4 [20] Japan (Okinawa) C-prenylated flavanones Guttiferone E 4 [20] and Taiwan Propolin A 5 [66,67] Xanthochymol 4 [20] [66,67] Polyisoprenylated benzophenones 20 [63] Propolin B 5 [66,67] Venezuela Scrobiculatone A 20 [63] Propolin E 5 [66,67] Scrobiculatone B 20 [63] Isonymphaeol B or propolin F 5 [66,67] 18-Ethoxy-17-hydroxy-17,18- 20 [63] Nymphaeol A or Propolin C 5 [66,67] dihydro- scrobiculatone A Nymphaeol A 5 18-Ethoxy-17-hydroxy-17,18- 20 [63] dihydro- scrobiculatone A Table 12: Some components present in propolis from Canarian Islands were effectively of this type. Propolis from Takebe-cho had Country Components Figure References constituents not present in the propolis of other places of the Spain (Canarian Lignans same Prefecture. Such results were attributed to a plant grown Islands) 3-Acetoxymethyl-5[(E)-2- 6 [74] mainly in this region (Rhus javanica var. chinensis). formylethen-1-yl]2-(4-hydroxy- 3-methoxyphenyl-7-methoxy-2,3- dihydrobenzofuran In the Southern Hemisphere, the chemical composition of Sesamin 6 [74] [73] propolis from different regions in Java were also evaluated. Seartenin 6 [74] The authors found some variability of constituents in the samples of propolis from three different places, in spite of that, all of them contained phenolic acids. The samples from Batany the samples of this region detected that they were significantly showed the presence of aromatic acids groups, terpenes, and different from the remaining ones studied so far in the world. 3,4-dimethylthioquinoline and 3-quinolinecarboxamine, two new Lignans [Table 12 and Figure 6], mainly of the furofuran type, compounds. In the Lawang sample, high amounts of aromatic and carbohydrates (pentoses, hexoses, and dissacharide) were acids were found and three new compounds were also reported found to be the main components.[74] According to the authors, (4-oxo-2-thioxo-3-thiazolidinepropionic acid, glucofuranuronic two plant species could be responsible for that composition: acid, and patchoulene). In the Sukabumi propolis sample, only one of them would produce resinous exudates rich in lignans of very low amounts of aromatic acids were found and silanol was the furofuran type and a second plant source from which most reported for the first time. The presence of this compound in the propolis samples was explained by the presence of rubber plants sugars were used by honeybees for creating propolis. in that area and the extract latex product from the Hevea rubber plants was polydimathylsiloxane elastomer.[73] Volatile Compounds

‘Canarian’ Propolis from Canary Islands Very few studies on the volatile fraction of worldwide propolis were found. This was not surprising, as essential oils constituted Only one study was found about the chemical composition of generally 10% of the samples against 50% of the resinous the propolis from Canary Islands. The authors who studied substances.

Journal of Pharmacy and Bioallied Sciences October-December 2011 Vol 3 Issue 4 489   Miguel and Antunes: Propolis and health

Volatile compounds are in fact found in low concentrations in b-caryophyllene, and selina-3,7(11-diene) constituted the propolis, but their aroma and biological activity make them most important components of the essential oil of propolis.[82] significant for the characterization of propolis.[4] Mono- and Green propolis collected in southeastern Brazil, was mainly sesquiterpenoids constitute the most important group of composed of nerolidol, a-pinene, 1-phenyl-ethanone, linalool, compounds found in propolis samples from diverse regions of trans-caryophyllene, d-cadinene, Spathulenol, and globulol.[83] Europe. Prenylated acetophenones can also be found in some Generally all the compounds were also present in the essential [4] samples from Brazil. Similar to reports on the extracts, the oil of Baccharis dracunculifolia. volatile chemical composition of propolis is also dependent on the flora. Some authors have reported that the volatile The red propolis from Maceio city, Alagoas State, had trans- composition of propolis is also dependent on the bee species, anethole, methyl eugenol, trans-methylisoeugenol, elemicin, because Apis mellifera and Melipona beechei in the same region and trans-isoelemicin. These components were partially of Yucatán (Mexico) have produced propolis with different responsible for the unusual anis-like odor of this red propolis.[84] [75,76] volatile compositions. The same has occurred in Turkey Red propolis from Goiana, Pernambuco State, had major volatile where phenols and terpenes present in the propolis samples components like trans-anethole, a-copaene, and methyl cis- depend on the race of honeybees.[77] isoeugenol. The authors observed also that some compounds could be present or absent according to the collection season, There are variations in the chemical composition of the volatile although trans-anethole was always the major one.[85] Also, in fraction of propolis from the temperate and tropical zones. October they found, d-cadinol, b-gurjunene, isocaryophyllene, Although propolis from the temperate climate zones is generally classified into two types: eudesmol and benzyl benzoate, in the and d-cadinene, which were absent in the remaining collection tropical zones there is great chemical variability.[76] However, seasons, whereas, in June, 1,8-cineole and a-selinene were the propolis collected at five different locations in Greece revealed a main components found, in contrast to the remaining collection predominance of a-pinene, except in samples from one location, seasons. in which junipene existed. The presence of junipene along with trans-b-terpineol, manool or manoyl oxide in these samples was Nerolidol, spathuelenol, aliphatic hydrocarbons, and reported for the first time as a propolis constituent.[78] However, alkylbenzenes and alkylnaphtalenes constituted the major in Europe, -pinene does not predominate in the volatile fraction components or the group of components present in samples of propolis. For example, the major volatile components of of propolis from Piaui State and Parana State (Brazil).[86] Dalmatian (Croatia) propolis were benzyl alcohol, benzoic Kusumoto et al.,[87] found in the Brazilian essential oils of acid, and benzyl benzoate (49%), along with terpenes (30%). propolis from the virgin forests of Alecrim, in the Southern In the Canary Islands, the main components appeared to be part of Minas Gerais State, the following components: terpenoids, mainly sesquiterpene hydrocarbons and alcohols. 2,2-dimethyl-8-prenyl-6-vinylchromene and 2,6-diprenyl-4- Spathulenol was the major sesquiterpene in the Canary Island vinylphenol (two new compounds), along with 2,2-dimethyl- samples, and benzoyl benzoate was also found.[74] 6-vinylchromene, acetophenone, 2-prenyl-4-vinylphenol, 3,4-dimethoxy-styrene, 3,4-dimethoxy-allylbenzene, 4-hydroxy- With regard to terpenoids obtained by extraction with 70% 3,5-diprenylbenzaldehyde, and (-)-spathulenol. ethanol, it was found that propolis from Anatolia (Turkey) had a-cedrol, b-eudesmol, a-eudesmol, -bisabolol, b-caryophyllene, These findings show the variability of the chemical composition d-cadinene, caryophyllene oxide, b-selinene, a-cadinol, of the volatile fraction of propolis. Another example is that in aromadendrene 2, and germacrene A. These compounds were which propolis from five different places of Brazil is shown to be not present in all samples, but depended on the region from qualitatively diverse: 1,8-cineole, exo-fenchol, terpin-4-ol, and [79] where they were collected. fenchone (Teresina); α-pinene, caryophyllene oxide, α-pinene, and α-copaene (Pio IX); (E)-caryophyllene, α-copaene, The volatile components of 23 propolis samples from 17 α-pinene, caryophyllene oxide, and α-cadinene (Campo Maior); provinces of China were analyzed by dynamic headspace (E)-caryophyllene, α-gurjunene, and α-selinene (Pedro II); and sampling, with gas chromatography and mass spectrometry.[80] (E)-caryophyllene, α-gurjunene, α-cadinene, and α-copaene The major components were a-cedrene, g-eudesmol, 3-methyl-3- (Lagoa de Sao Francisco). All these places belong to the Piaui buten-l-ol, g-terpineol, acetic acid, benzyldehyde, benzyl alcohol, State.[76] More examples can be found, the most abundant cedrol, butanoic acid, ethyl ester, g-cadinene, phenylethyl alcohol, and styrene. Acetic acid was already found in higher components of propolis from Rio de Janeiro collected in July content in other Chinese propolis.[81] Luo et al.[80] suggested were: b-caryophyllene, acetophenone, linalool, g-elemene, [88] that the volatile chemical composition detected in the propolis g-cadinene, and g-muurolene. samples could be related to different climates and plant origins. Biological Properties of Propolis Several studies concerning the essential oils isolated from Brazilian propolis were found, and they demonstrated the Despite the great chemical variability of propolis, there are diversity of the chemical composition. The essential oil hundreds of studies dealing with the biological activity of isolated from the green propolis of Minas Gerais was for the propolis, sometimes from different origins, but with similar first time studied and the authors reported that (E)-nerolidol, properties.[89-134]

 490 Journal of Pharmacy and Bioallied Sciences October-December 2011 Vol 3 Issue 4 Miguel and Antunes: Propolis and health 

The distinct chemistry of propolis from diverse origins the phenolic fraction were different. In fact, in the samples from sometimes does not mean dissimilar properties.[20] Propolis, Akita, caffeic acid and phenethyl caffeate were predominant. being a defense system of bees against infections, they have to In Okinava, the antioxidant activity was due to the prenylated find components that possess such properties. Bees can find flavonoids. Kumazawa et al.[138] showed that the position of the them in the flora that surrounds their beehives. In Europe, such geranyl or prenyl groups on the flavonoid skeleton played an compounds include flavanones, flavones, phenolic acids, and important role on the antioxidant activity. Also Chen et al.[66] their esters from poplars, whereas in Brazil the anti-infectious reported that some propolins had better activity than the others. compounds are prenylated p-coumaric acids and diterpenes from B. dracunculifolia or prenylflavanones from ‘Pacific’ Both propolis from Europe and Brazil, with diverse chemical propolis.[20,115] compositions, possessed anti-inflammatory activities. In both cases the mechanism was due to the inhibition of NO Some authors[135] compared the antibacterial activity of three production.[108,139] The open-chained neoflavonoids from Nepal, groups of propolis: European, Brazilian, and Central American. also had the capacity for inhibiting NO production, mainly Propolis from Brazil and Europe possessed similar activities those having a ketone carbonyl at C-1 in ring A, instead of those despite the dissimilarity in the composition, and the propolis having a methoxyl substituent.[39] from Central America had lower antibacterial activity. In the same study, no significant correlation was found between The caffeic acid phenethyl ester and its related polyphenolic geographic origin and potential cytotoxicity. acid esters elicited an important effect on erythrocyte membrane lipid peroxidation, cellular strand breakage, and However, it is also true that different components can give protein fragmentation. According to these results, the authors [101] diverse antimicrobial properties. Seidel et al. comparing the considered that caffeic acid phenethyl ester could be used as a antibacterial activity of propolis from different geographical origins, template for designing novel drugs to combat diseases induced by including countries from tropical, subtropical, and temperate oxidative stress components, such as, diverse types of cancer.[140] zones, found that propolis extracts were mostly active against Propolis from Brazil, China, Peru, and The Netherlands also Gram positive bacteria. They also observed that propolis from the possessed strong cytotoxicity toward murine colon 26-L5 wet-tropical, rainforest-type climate had the highest antibacterial carcinoma and human HT-1080 fibrosarcoma cells, although [101] activity. The methodology used was the same for all analyses those from The Netherlands and China possessed the strongest (broth microdilution assay). Other studies demonstrated that cytotoxicity toward murine colon 26-L5 carcinoma cells, which propolis from Brazil, Peru, The Netherlands, and China, generally may because these samples had the highest presence of phenolic possessed hepatoprotective activity and scavenging activity compounds.[136] against DPPH free radicals, whereas, those from China and The Netherlands had the strongest cytotoxic activity.[136] Despite the chemical difference among the three types of propolis studied by Messerli et al.,[132] the caffeic acid phenethyl In vitro antioxidant activity (b-carotene bleaching test and ester-based propolis in Europe, Far East, and New Zealand, the DPPH free radical scavenging assays) of the ethanol extracts artepillin C-based Brazilian green propolis, and the Brazilian of propolis from diverse regions (Argentina, Austria, Brazil, red propolis, they found that all of them possessed anticancer Bulgaria, Chile, China, Hungary, New Zealand, South Africa, activity. They blocked selective oncogenic PAK1 signaling and Thailand, Ukraine, Uruguay, the USA, and Uzbekistan) was suppressed the growth of neurofibromatosis tumors in mice. evaluated by Kumazawa et al.[137] and compared with the chemical composition. This was performed by HPLC analysis with photo-diode array (PDA) and mass spectrometry (MS). Conclusions The most active propolis was that obtained from Argentina, Australia, China, Hungary, and New Zealand, which also Propolis is a heterogeneous product constituted by several possessed the highest total polyphenol and flavonoid contents. groups of compounds. Moreover, the chemical composition The activity correlated well with the levels of kaempferol depends strongly on the phytogeographic characteristics of and phenethyl caffeate present in these samples of propolis. the collection site, as honey bees can only use the plant species Practically the same compounds were present in the propolis existing in their habitats. Their chemical variability can give rise obtained from various places in China (caffeic acid, ferulic acid, to diverse types of biological activities or diverse structures may and caffeic acid phenethyl ester), presenting in all of them a present similar properties. Therefore, to make a standardization strong antioxidant activity.[35] Similar results were observed in and quality control of this product is very difficult, particularly if the propolis from different locations of Korea, which means, we take into account the quantification of the active substances. samples with high amounts of caffeic acid, kaempferol, and Popova et al.[67] have proposed to specify multiple standards phenethyl caffeate possessed the best antioxidant activity, in for different propolis types according to their plant source and contrast to a sample from a more distant place, in which the corresponding chemical profile. Popova et al.[141] has already levels of those components were lower.[37] Some authors,[70] made a standardization for the poplar-type propolis from reported a relationship between high phenolic content and Europe, Asia, and Americas. More recently, Popova et al.[67] have antioxidant activity in the propolis from different zones of validated a spectrophotometric method for the quantification of Japan. The samples with the best activities were those from prenylated flavanones in the ‘Pacific’ propolis from Taiwan. In Akita (Minamiakita) and Okinawa. However, the constituents of addition, it is necessary to connect a particular chemical propolis

Journal of Pharmacy and Bioallied Sciences October-December 2011 Vol 3 Issue 4 491   Miguel and Antunes: Propolis and health type to a specific type of biological activity for formulating propolis. Experientia 1978;34:157-8. recommendations for the practitioners. Only by following this 15. Juliano C, Pala CL, Cossu M. Preparation and characterisation of polymeric films containing propolis. J Drug Deliv Sci Technol scheme will it be possible for people to choose and make more 2007;17:177-80. efficient use of the beneficial properties of propolis, in respect 16. Popova M, Chen CN, Chen PY, Huang CY, Bankova V. A validated to complementary and alternative medicine.[142] spectrophotometric method for quantification of prenylated flavanones in pacific propolis from Taiwan. Phytochem Anal 2010;21:186-91. In spite of propolis being commonly used in cosmetic 17. de Sousa JP, Bueno PC, Gregório LE, da Silva Filho AA, Furtado NA, and medicinal preparations owing to its antiseptic, anti- de Sousa ML, et al. A reliable quantitative method for the analysis inflammatory, and anesthesic properties, it is not completely of phenolic compounds in Brazilian propolis by reverse phase high innocuous because 1.2 to 6.6 patients who were patch-tested performance liquid chromatography. J Sep Sci 2007;30:2656-65. 18. Chang R, Piló-Veloso D, Morais SA, Nascimento EA. Analysis of a for dermatitis were sensitive to propolis. The main allergens Brazilian green propolis from Baccharis dracunculifolia by HPLC-APCI- were 3-methyl-2-butenyl caffeate and phenylethyl caffeate, that MS and GC-MS. Braz J Pharmacogn 2008;18:549-56. is, components present in the poplar-type propolis.[143] Clinical 19. Miguel MG, Nunes S, Dandlen SA, Cavaco AM, Antunes MD. Phenols and antioxidant activity of hydro-alcoholic extracts of propolis from allergy in humans is presented as contact dermatitis or oral Algarve, South of Portugal. Food Chem Toxicol 2010;48:3418-23. mucositis, beekeepers being the most affected. Nevertheless 20. Bankova V. Chemical diversity of propolis and the problem of there has been a recent rise in this incidence among biocosmetic standardization. J Ethnopharmacol 2005;100:114-7. users, on account of the increasing popularity of natural products 21. Popova MP, Graikou K, Chinou I, Bankova VS. GC-MS profiling of [144] diterpene compounds in Mediterranean propolis from Greece. J such as propolis. According to these authors, patients with Agric Food Chem 2010;58:3167-76. an allergy to propolis may be at risk of cross-sensitization with 22. Trusheva B, Popova M, Bankova V, Tsvetkova I, Naydensky H, Sabatini balsam of Peru, a common allergen found in flavoring agents, AG. 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A. Antibacterial activity of essential oils from Brazilian propolis. activity of ethanol extracts of Chinese and Brazilian propolis in four Fitoterapia 1999;70:190-93. human colon carcinoma cell lines. Oncol Rep 2009;22:349-54. 87. Kusumoto T, Miyamoto THiguchi R, Doi S, Sugimoto H, Yamada H. 111. Kunimasa K, Ahn MR, Kobayashi T, Eguchi R, Kumazawa S, Fujimori Isolation and structures of two new compounds from the essential Y, et al. Brazilian Propolis Suppresses Angiogenesis by inducing oil of Brazilian propolis. Chem Pharm Bull (Tokyo) 2001;49:1207-9. Apoptosis in Tube-forming Endothelial Cells through Inactivation of 88. Oliveira AP, França HS, Kuster RM, Teixeira LA, Rocha LM. Chemical Survival Signal ERK1/2. Evid Based Complement Alternat Med 2011; composition and antibacterial activity of Brazilian propolis essential DOI: 10.1093/ecam/nep024 oil. J Venom Anim Incl Trop Dis 2009;16:121-30. 112. Kang L-J, Lee HB, Bae H-J, Lee S-G. Antidiabetic effect of propolis: 89. 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133. Szliszka E, Czuba ZP, Domino M, Mazur B, Zydowicz G, Krol W. Kadota S. Caffeic acid phenetyl ester (CAPE) analogues: potent Ethanolic extract of propolis (EEP) enhances the apoptosis-inducing nitric oxide inhibitors from the Netherlands propolis. Biol Pharm potential of TRAIL in cancer cells. Molecules 2009;14:738-54. Bull 2003;26:487-91. 134. Demestre M, Messerli SM, Celli N, Shahhossini M, Kluwe L, Mautner 140. Wang T, Chen L, Wu W, Long Y, Wang R. Potential cytoprotection: V, et al. CAPE (caffeic acid phenetyl ester)-based propolis extract (Bio antioxidant defense by caffeic acid phenethyl ester against free 30) suppresses the growth of human neurofibromatosis (NF) tumor radical-induced damage of lipids, DNA, and proteins. Can J Physiol xenografts in mice. Phytother Res 2009;23:226-30. Pharmacol 2008;86:279-87. 135. Popova M, Bankova V, Naydensky C, Tsvetkova I, Kujumgiev A. 141. Popova MP, Bankova VS, Bogdanov S, Tsvetkova I, Naydenski C, Comparative study of the biological activity of propolis from Marcazzan GL, et al. Chemical characteristics of poplar type propolis different geographic origin: A statistical approach. Maced Pharm of different geographic origin. Apidologie 2007;38:306-11. Bull 2004;50:9-14. 142. Bankova V. Recent trends and important developments in propolis 136. Banskota AH, Tezuka Y, Adnyana IK, Midorikawa K, Matsushige research. eCAM 2005;2:29-32. K, Message D, et al. Cytotoxic, hepatoprotective and free radical 143. Walgrave SE, Warshaw EM, Glesne LA. Allergic contact dermatitis scavenging effects of propolis from Brazil, Peru, the Netherlands from propolis. Dermatitis 2005;16:209-15. and China. J Ethnopharmacol 2000;72:239-46. 144. Lieberman HD, Fogelman JP, Ramsay DL, Cohen DE. Allergic contact 137. Kumazawa S, Hamasaka T, Nakayama T. Antioxidant activity of dermatitis to propolis in a violin maker. J Am Acad Dermatol 2002;46(2 propolis of various geographic origins. Food Chem 2004;84:329-39. Suppl):S30-1. 138. Kumazawa S, Ueda R, Hamasaka T, Fukumoto S, Fujimoto T, Nakayama T. Antioxidant prenylated flavonoids from propolis collected in Okinawa, Japan. J Agric Food Chem 2007;55:7722-5. Source of Support: Nil, Conflict of Interest: None declared. 139. Nagaoka T, Banskota AH, Tezuka Y, Midorikawa K, Matsushige K,

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