Floral Markers in Honey of Various Botanical
and Geographic Origins: A Review
Vilma Kaškonienė and Petras R. Venskutonis
Abstract: In view of the expanding global market, authentication and characterization of botanical and geographic
origins of honey has become a more important task than ever. Many studies have been performed with the aim of
evaluating the possibilities to characterize honey samples of various origins by using specific chemical marker compounds.
These have been identified and quantified for numerous honey samples. This article is aimed at summarizing the studies
carried out during the last 2 decades. An attempt is made to find useful chemical markers for unifloral honey, based on the
analysis of the compositional data of honey volatile compounds, phenolic acids, flavonoids, carbohydrates, amino acids,
and some other constituents. This review demonstrates that currently it is rather difficult to find reliable chemical markers
for the discrimination of honey collected from different floral sources because the chemical composition of honey also
depends on several other factors, such as geographic origin, collection season, mode of storage, bee species, and even
interactions between chemical compounds and enzymes in the honey. Therefore, some publications from the reviewed
period have reported different floral markers for honey of the same floral origin. In addition, the results of chemical
analyses of honey constituents may also depend on sample preparation and analysis techniques. Consequently, a more
reliable characterization of honey requires the determination of more than a single class of compounds, preferably in
combination with modern data management of the results, for example, principal component analysis or cluster analysis.
Introduction
With the globalization of the honey market, which currently
involves approximately 150 countries, the identification of honey
origin as well as the proof of its authenticity has become an important issue. Therefore, the search for reliable chemical markers
indicating floral and/or its geographic origin has been the focus
of many studies in the last 15 y. Honey composition, the nutritional contribution of its components, as well as its physiological
and nutritional effects were comprehensively reviewed 6 y ago by
Bogdanov and others (2004), the analytical methods for determination of volatile compounds from the honey were also reviewed
by Cuevas-Glory and others (2007), while the composition of
volatile compounds and sugars in honey of different floral origin
were reviewed only in Portuguese (Moreira and De Maria 2001;
De Maria and Moreira 2003). The review dealing with geographical and botanical origin of honey was published by Anklam in
1998; however, it was focused mainly on analytical methods used
to determine the origin of honey.
It is difficult to find a unanimous opinion about the floral markers of honey collected from different floral origins; indicators that
MS 20100312 Submitted 3/23/2010, Accepted 6/25/2010. Author Kaškonienė is
with Dept. of Biochemistry and Biotechnologies, Vytautas Magnus Univ., Vileikos 8,
LT-44404, Kaunas, Lithuania. Author Venskutonis is with Dept. of Food Technology,
Kaunas Univ. of Technology, Radvilėnu rd, 19, LT-50254, Kaunas, Lithuania. Direct
inquiries to author Venskutonis (E-mail: [email protected]).
might distinguish one honey from another, depend on many factors. Therefore, unambiguous and unique indicators for honey
collected from the same plant source have not been identified
until now; very often honey from the same botanical origin analyzed in various studies is characterized by different compounds.
For example, Cabras and others (1999) found that strawberry-tree
(Arbutus unedo L.) honey from Italy can be differentiated from
others by homogentisic acid, Cherchi and others (1994) reported
gluconic acid as a typical component for the same sort of honey,
while most recently Tuberoso and others (2010), based on their
studies of honey from Sardinia, concluded that in addition to homogentisic acid, abscisic acid isomer levels, and the presence of
unedone could represent further useful markers for fingerprinting
the botanical origin of strawberry-tree honey. In this case, the
different results may be due to the different extraction methods:
Cabras and others (1999) as well as Tuberoso and others (2010)
used solvent extraction, while Cherchi and others (1994) applied
solid phase extraction (SPE). Isolation method may also have an
effect on the results of analysis of honey aroma compounds, as
it was shown by Alissandrakis and others (2005a); they reported
that differences in aroma composition may be due to sorbent selectivity (for example, in the solid phase microextraction [SPME]
method) or solubility in the organic solvent used (for example, in
ultrasound-assisted extraction).
Honey is a very complex product, because its properties and
composition depend not only on the nectar-providing plant
species, but also on other factors such as bee species, geographic
area, season, mode of storage, and even harvest technology and
620 Comprehensive Reviews in Food Science and Food Safety r Vol. 9, 2010
c 2010 Institute of Food Technologists®
doi 10.1111/j.1541-4337.2010.00130.x
Floral markers of honey . . .
FLORAL MARKERS OF HONEY
Volatile compounds
Phenolic compounds
Phytochemicals
Pollen
Carbohydrates
Nitrogen containing
compounds
Other minor compounds
Terpenes
Flavonoids
Amino acids
Microelements
Norisoprenoids
Non-flavonoid
phenolic compounds
Non-amino acids
Miscellaneous
Other volatiles
Figure 1–Possible floral markers of honey.
conditions. It is known that accumulation of phytochemicals depends on climatic conditions (sunlight, moisture), soil characteristics, and other factors; therefore, it is reasonable to believe that
differences between honeys from country to country appear due
to the different compositions of pollen or nectar, which have the
greatest influence on the chemical composition of honey. For
instance, Karousou and others (2005) showed that essential oil
composition was very dependent on the geographic location even
for the same plant species, which suggests that even the same floral
origin honey composition may be quite different.
The fact that unifloral honeys are never actually unifloral should
also be carefully considered. The nectar from various flowers contributes to the production of every honey sample; therefore, the
researchers should be very careful proposing the compounds found
in low concentrations as floral markers. Moreover, this problem
becomes even more remarkable when conclusions are made from
a small number of samples. In addition, many compounds of biological and botanical origin found in honey are not stable (particularly volatile substances), and their structures may transform
in the course of honey maturation and storage (Kaškonienė and
others 2008). Several groups of phytochemical markers of floral
origin may now be differentiated according to recently published
data (Figure 1). The sections in this review are based on this classification in order to provide the state-of-the-art results attempting
to define typical honey compounds most closely associated with
its floral origin. In addition, pollen analysis (melissopalynological
method) as a technique that has been used in many laboratories as
a nonchemical method of botanical characterization of honey will
be briefly discussed in a separate section below.
Pollen Analysis for Characterization of Honey
Botanical Origin
Traditionally, the botanical origin of honey has been determined
in many laboratories by pollen analysis, a method that is called
melissopalynology. The melissopalynological method, which was
elaborated and proposed by the Intl. Commission for Bee Botany
(ICBB) in 1970 (Louveaux and others 1970) and later revised and
updated in 1978 (Louveaux and others 1978), is frequently used
c 2010 Institute of Food Technologists®
until the present time. Moreover, in EU Council Directive (2002)
related to honey, it is indicated that the product names may be
supplemented by information referring to floral or vegetable origin, if the product comes wholly or mainly from the indicated
source and possesses the organoleptic, physico-chemical, and microscopic characteristics of the source. However, this method is
time-consuming, requires specialized knowledge and expertise,
and involves a laborious counting procedure, with interpretation
of results and an identification of botanical origin being rather
difficult. Melissopalynological determination of botanical origin
of honey is based on the relative frequency of the pollen from the
nectar-secreting plants. Particular difficulties in this method are
associated with the need of good experience and knowledge of
pollen morphology and the availability of a comprehensive collection of pollen grains. Different opinions may occur regarding the
use of pollen present in honey for the indication of its botanical
origin. For instance, Molan (1998) reported such disadvantages of
this method as possible alterations of pollen content in honey by
the action of bees or contamination by the actions of apiarists; in
addition, the source of some honeys, for example, collected from
cotton plants, castor-oil plants, and rubber trees, cannot be identified by melissopalynology. According to this author, melissopalynology is valid only for the determination of geographic origin
of honey, while it is less valid for determining the botanical origin
of honey. Alissandrakis and others (2007a) reported that Greek
citrus honey is also difficult to identify with pollen analysis. Most
recently, Stephens and others (2010) observed that melissopalynological analysis was not able to differentiate manuka (Leptospermum
scoparium) and kanuka (Kunzea ericoides) honeys from New Zealand
due to the similarity of pollen grains.
In some cases, most likely due to the above-mentioned limitations of the melissopalynological method, studies on the chemical
composition of honey were performed without pollen analysis;
in such cases, botanical origin of honey was based on the claims
of local beekeepers, when determination of honey origin is performed by sensory analysis or by considering the predominant
flowers surrounding the hive. The latter approach may be quite
precise, for example, allowing collecting honey with 72% to 75%
Vol. 9, 2010 r Comprehensive Reviews in Food Science and Food Safety 621
Floral markers of honey . . .
of a predominant pollen (Jerković and others 2009a). Alissandrakis
and others (2003) recommend to obtain a unifloral honey sample
by placing bee colonies containing unbuilt combs in the middle
of a large area of the plants from which the honey is collected.
Some researchers point to unsuitability of melissopalynology for
rapid routine analysis as the main disadvantage of this method
(Arvanitoyannis and others 2005; Terrab and others 2005). Even
though pollen analysis has some disadvantages, it is the only way to
detect contribution of nectar from other floral origin. In spite of
the limitation and disadvantages of melissopalynology, Dimou and
others (2006) proved that microscopic analysis is suitable for the
discrimination of fir (Abies cephalonica) and pine (Pinus sp.) honeys
honeydew elements.
Therefore, studies of honey and a search for faster methods of
its characterization, which would be suitable for routine analysis,
particularly involving many samples in a short work period, are
now carried out with the help of novel instrumental techniques,
such as atomic absorption spectroscopy (AAS) (Hernández and
others 2005), high-performance liquid chromatography (HPLC)
(Andrade and others 1997a), gas chromatography with mass spectrometry (GC-MS) (Alissandrakis and others 2007a; Aliferis and
others 2010), electro-spray mass spectrometry (ES-MS) (Beretta
and others 2007), inductively coupled plasma optical emission
spectrometry (ICP-OES) (Terrab and others 2005), thin-layer
chromatography (TLC) (Rezić and others 2005), highperformance anion-exchange chromatography with pulsed amperometric detection (HPAED-PAD) (Nozal and others 2005),
and nuclear magnetic resonance (NMR) (Tuberoso and others
2010), Fourier transform–Raman (FT-Raman) (Paradkar and Irudayaraj 2002), Fourier transform infrared (FTIR) (Etzold and
Lichtenberg-Kraag 2008), and near-infrared (NIR) spectroscopies
(Woodcock and others 2007).
Volatile Compounds as Floral Markers of Honey
Aroma is one of the distinguishable characteristics of honeys
collected from different flowers. Therefore, many studies have
been carried out in order to specify volatile compounds that are
most closely associated with a particular type of honey and consequently would be helpful for a fast and reliable identification
of its botanical source. In general, the aroma of honey is formed
by volatile compounds that may come from the nectar or honeydew collected by bees; consequently, it may largely depend on
the plant of honey origin and on the state of honey maturity.
In addition, flavor constituents may be formed by the honeybee, as well as during thermal processing and/or storage of honey
(Bonvehı́ and Coll 2003; Soria and others 2003). And although
volatile compounds do not comprise a specific category of compounds that are related to their chemical structure overwhelming
majority of studies on honey volatiles were dealing with all constituents, which are present in honey headspace or may be distilled
from the product. Therefore, these compounds are reviewed in
1 section.
More than 600 organic compounds have been identified in the
extracts of volatiles or headspace isolates of honey samples originating from different floral types (Jerković and others 2009b).
Taking into account that aroma composition of some types of
honey has not yet been studied and that the sensitivity of analysis is continuously improving, it is very likely that the number
of identified compounds will further increase; the method of extraction also plays an important role. Usually, unifloral honeys
possess highly individual aroma profiles as compared with polyfloral honeys, due to the presence of characteristic volatile organic
components deriving from the original nectar sources. A number
of authors have reported the same volatile compounds and/or their
metabolites in honey, in its precursor nectar, and in the flowers
from which it is collected (Rowland and others 1995; Alissandrakis and others 2003; Moreira and De Maria 2005). Volatile
honey compounds belong to 7 major groups: aldehydes, ketones,
acids, alcohols, esters, hydrocarbons, and cyclic compounds. Although some aldehydes and alcohols reflect product quality and
may also be a consequence of microbiological activity, heat exposure, and honey aging, some other compounds, such as linear
aldehydes are considered as characteristic honey compounds associated with certain floral origin. Typical volatile components can
be identified for honeys from some definite floral sources; such
compounds are specified as floral markers of the corresponding
honey. Some volatile compounds have been used for the geographical discrimination of honey samples. Radovic and others
(2001) have classified honeys according to the geographic origin.
It was suggested that English honey samples can be distinguished
by the presence of 1-penten-3-ol; honey from Denmark by the absence of 3-methylbutanal. 2,2,6-Trimethylcyclohexanone, ethyl2-hydroxypropanoate, 3-hexenyl-formate, and a few not as yet
identified compounds with characteristic fragment ions have been
suggested as possible marker compounds for honey from Portugal.
1-Octen-3-ol or 2,6,6-trimethyl-2,4-cycloheptadien-1-one have
been suggested as possible marker compounds for honey from
Spain, while pentanal and cis-linalool oxide were not found in the
Spanish honeys. However, it should be noted that Radovic and
others (2001) analyzed only 3 English samples, 1 from Denmark,
2 from Portugal, and 2 from Spain and therefore their data may be
considered as rather preliminary; considerably more samples from
the same country should be evaluated in order to obtain more
reliable conclusions.
It was reported that some types of honey may be distinguished
by 1 characteristic compound. Methyl anthranilate was reported
as a marker of citrus honey (Ferreres and others 1994b; White
and Bryant 1996; Piasenzotto and others 2003; Soria and others 2003; Alissandrakis and others 2005a, 2007a), isophorone of
strawberry-tree honey (de la Fuente and others 2007b), and honeys
from the Ericaceae family (Guyot and others 1999). The main compounds suggested as possible markers of various honeys are listed in
Table 1, while the structures of some important volatile constituents found in honey are given in Figure 2. Until now, published data demonstrated that only few types of honeys contain
1 characteristic compound, while the majority of the previously
tested honey samples were characterized by several compounds.
The majority of studies were performed with chestnut, heather,
eucalyptus, and citrus honeys of various geographic origins, however, the results obtained for the same origin samples in many
cases are rather different (Table 1). For instance, Castro-Vázquez
and others (2007) pointed out the isomers of sinensal as the best
floral markers for citrus honeys as the compounds responsible
for “sweet” and “orange-like” aroma notes. Later, the same authors extended their study by applying multivariate analysis and
in addition to sinensal isomers distinguished linalool derivatives,
lilac aldehydes, lilac alcohols, limonyl alcohol, and α-4-dimethyl3-cyclohexene-1-acetaldehyde as other constituents involved in
the formation of citrus honey aroma (Castro-Vázquez and others
2009). Methyl anthranilate and sinensal isomers were identified
only in citrus honey samples, while, for example, phenylacetaldehyde was also attributed as a characteristic compound of chestnut,
eucalyptus, and lavender honeys; heptanal was found in lavender
and acacia honeys; nonanal was determined in 4 different honeys,
622 Comprehensive Reviews in Food Science and Food Safety r Vol. 9, 2010
c 2010 Institute of Food Technologists®
c 2010 Institute of Food Technologists®
Lime tree
Blue gum
Eucalyptus
Buckwheat
Heather
Calluna vulgaris
Heather
Erica arborea
Chestnut
Sage
Christ’s thorn
Honey source
Rhododendron
Benzoic acid, decanoic acid, high levels of cinnamic acid,
isophorone, 4-(3-oxobut-1-enylidene)-3,5,5trimethylcyclohexen-2-en-1-one
Phenylacetic acid, dehydrovomifoliol, (4-(3-oxo-1-butynyl)3,5,5-trimethylcyclohexen-2-en-1-one)
4-Methylbicyclo[2,2,2]octan-1-ol or 4-ethylphenyl acetate
High contents of benzene and guaiacol, p-anisaldehyde,
propylanisole and p-cresol
α-Isophorone, 2-hydroxy-3,5,5-trimethylcyclohexanone,
furfuryl alcohol, benzyl alcohol, 2-phenylethanol
3-Methylbutanal, 3-hydroxy-4,5-dimethyl-2(5H)-furanone
(sotolon), β-damascenone
8,9-Dehydrotheaspirone and 3-oxo-α-ionone
1-Octene or 2,3-pentanedione
Nonanal, ethylphenyl acetate, phenethyl alcohol
Acetoin, 5-hydroxy-2,7-dimethyl-4-octanone, p-cymene
derivatives, 3-caren-2-ol and spathulenol
Acetoin, diacetyl, 2,3-pentanedione, dimethyldisulfide
2-Hydroxy-5-methyl-3-hexanone,
3-hydroxy-5-methyl-2-hexanone
2,3-Pentanedione, acetoin, 1-hexyl alcohol,
2-acetyl-5-methylfuran, furfuryl propionate,
2-phenylacetaldehyde, nerolidol
2-Methylfuran or α-terpinene or α-pinene oxide or
α-terpene or methyl isopropyl benzene or
4-methylacetophenone
Ethylmethylphenol, carvacrol, estragole
Compound
n-Decane, lilac aldehyde, 2-aminoaceto phenone,
benzenedicarboxylic, nonanal, isobutylphthalate,
damascenone
Nonanal, 4 isomers of lilac aldehyde, decanal, methyl
nonanoate, hexanoic and 2-ethylhexanoic acids
Tetrahydro-2,2,5,5-tetramethylfuran, 3-hexenyl ester of
butanoic acid, 2-methylbenzene, maltol, methyl ester of
3-furanocarboxylic acid, benzeneacetic acid
Benzoic acid, phenylacetic acid, p-anisaldehyde,
a-isophorone, 4-ketoisophorone, dehydrovomifoliol,
2,6,6-trimethyl-4-oxocyclohex-2-ene-1-carbaldehyde,
2,2,6-trimethylcyclohexane-1,4-dione, and coumaran
High percentage of phenylacetic acid, low percentage of
benzoic acid, also 4-aminoacetophenone,
2-aminoacetophenone
n-Decane, phenylacetaldehyde, phthalic acid, αα-dimethylphenyl acetate, p-anisaldehyde
2-Methyldihydrofuranone or α-methylbenzyl alcohol or
3-hexen-1-ol and dimethylstyrene
2-Methylcyclopentanol, diethylphenol
Acetophenone, 1-phenylethanol, 2-aminoacetophenone
Acetophenone, 2-aminoacetophenone
Table 1– The most important volatile compounds in unifloral honeys.
TA trap)
ES
SPME
AU
EU
TR
ES
ES
ES
Catalonia and Basque area (ES)
EU
FR
Solvent
Purge-and-trap∗ (TenaxTM TA trap)
SPME
Likens–Nickerson
SPME
SPME
Likens–Nickerson
Purge-and-trap∗ (TenaxTM TA trap)
Likens–Nickerson
U.S.A. and ENG
EU
ES
Purge-and-trap∗ (TenaxTM TA trap)
Likens–Nickerson
Solvent
EU
EU
IT
FR and IT
Catalonia and Basque area (ES)
Likens–Nickerson
Likens–Nickerson
SPME
Likens–Nickerson
Likens–Nickerson
EU
TR
(TenaxTM
SPME
Purge-and-trap∗∗
HR
HR
Solvent
Solvent
HR
HR
GO∗
TR
SPME
SPME
Extraction method
SPME
Guyot and others 1998
(Continued)
Radovic and others 2001
Bonvehı́ and Coll 2003
de la Fuente and others 2005
de la Fuente and others 2007a
D’Arcy and others 1997
Radovic and others 2001
Senyuva and others 2009
Castro-Vázquez and others 2009
Zhou and others 2002
de la Fuente and others 2005
Radovic and others 2001
Castro-Vázquez and others 2009
Guyot and others 1999
Guyot and others 1999
Guidotti and Vitali 1998
Guyot and others 1998
Bonvehı́ and Coll 2003
Radovic and others 2001
Senyuva and others 2009
Jerković and others 2007
Jerković and others 2006
Lušić and others 2007
Jerković and others 2009a
Reference
Senyuva and others 2009
Floral markers of honey . . .
Vol. 9, 2010 r Comprehensive Reviews in Food Science and Food Safety 623
624 Comprehensive Reviews in Food Science and Food Safety r Vol. 9, 2010
Compound
Lilac aldehydes, dill ether, methyl anthranilate, 1-p-menthen-9-al
isomers
Methyl anthranilate
(E)-Linalool oxide, lilac aldehydes, lilac alcohols, limonyl alcohol,
sinensal isomers, α-4-dimethyl-3-cyclohexene-1-acetaldehyde
Limonene diol, methyl anthranilate
Lilac aldehydes, methyl anthranilate
α and β sinensal
Heptanal
Hexanal, nerolidol oxide, coumarin, high concentrations of hexanol
and hotrienol
n-Hexanal, n-heptanal, phenylacetaldehyde, coumarin
n-Hexanal, n-heptanal, hexanol, phenylacetaldehyde
cis-Linalool oxide and heptanal
α-Pinene or 3-methyl-2-butanol
Cinnamaldehyde, cinnamyl alcohol, cinnamic acid, neryl and geranyl
nitrile, benzenepropanol, homovanillyl alcohol, (E)- and
(Z)-p-methoxy-cinnamic acid, 2-methyl-p-phthalaldehyde,
coniferaldehyde, p-coumaric acid, ferulic acid, scopoletin, scoparone
Dimethyl disulfide
Methyl salycilate
Lilac aldehyde isomers
Methyl-p-anisaldehyde, p-anisaldehyde, p-cresol, trimethoxybenzene,
5-hydroxy-2-methyl-4H-pyran-4-one, lilac aldehyde isomers (A, B, C,
D), phenethyl alcohol, benzenacetaldehyde
Furfuryl mercaptan, benzyl alcohol, δ-decalactone, eugenol, benzoic
acid, isovaleric acid, phenylethyl alcohol, 2-methoxyphenol
3,4,5-Trimethoxybenzaldehyde
1-Phenyl-2,3-butanedione, 3-hydroxy-4-phenyl-2-butanone,
3-hydroxy-1-phenyl-2-butanone, phenylacetonitrile, and carvacrol
3-Hydroxy-4-phenyl-2-butanone, 3-hydroxy-1-phenyl-2-butanone,
2-methylpropionic acid, 4-(4-hydroxy-2,2,6-trimethyl-7oxabicyclo[4.1.0]hept-1-yl)-3-buten-2-one
1,3-Diphenyl-2-propanone, (3-methylbutyl)benzene, 3,4,5-trimethoxy
benzaldehyde, 3,4-dimethoxy benzaldehyde, vanilline, thymol
Benzene propanol, benzyl alcohol, nonanol, hexanol, 4-methoxyphenol
3,5-Dihydroxytoluene, tridecane
Isovaleric acid, γ-decalactone, benzoic acid, vanillin
Benzaldehyde, 2,3-methylbutanoic acid, benzonitrile, 2-phenylethanol
α-Isophorone, β-isophorone, 4-oxoisophorone
Acetonitrile, methyl-2-buten-1-ol, n-hexanol, 3-hexanol, 1-propyne,
2-furanmethanol, 5-methyl-2(5H)-furanone, 4-methylphenol,
hexadecanoic acid, methylheptanoate
n-Decane, nonanal, α-α-dimethylphenyl acetate, nonanol, 2-methyl
heptanoic acid
(E)-2,6-dimethyl-6-hydroxy-2,7-octadienoic acid,
(E)-2,6-dimethyl-3,7-octadiene-2,6-diol,
(Z)-2,6-dimethyl-2,7-octadiene-1,6-diol,
(E)-2,6-dimethyl-6-hydroxy-2,7-octadienal, lilac aldehydes,
lilac alcohols
3-Carene and unidentified compound (m/z 55, 79, 91, 107, 123, 162)
trans-Oak lactone, aminoacetophenone, propylanisol
EU
ES
ES
JP
BR
Purge-and-trap∗ (TenaxTM TA trap)
SPME
Likens–Nickerson
Purge-and-trap∗ (TenaxTM TA trap)
Purge-and-trap∗ (TenaxTM TA trap)
NZ
TR
ES
Purge-and-trap∗ (TenaxTM TA trap)
Solvent
TR
PS
PS
BR
BR
Sardinia (IT)
HR
PS
Solvent
SPME
SPME
SPME
Purge-and-trap∗ (TenaxTM TA trap)
Solvent
Purge-and-trap∗ (CarbopackTM trap)
SPME
SPME
GR
FR
PT and FR
EU
EU
GR
Dynamic headspace
Likens–Nickerson
Purge-and-trap∗ (TenaxTM TA trap)
Purge-and-trap∗ (TenaxTM TA trap)
Solvent
Solvent
IT
ES
ES
EU
ES
SPME
SPME
Likens–Nickerson
Purge-and-trap∗ (TenaxTM TA trap)
Likens–Nickerson
TR
GR
Florida (U.S.A.)
ES
Likens–Nickerson
Likens–Nickerson
SPME
SPME
GO∗
GR
Extraction method
SPME
Tananaki and others 2007
Castro-Vázquez and others 2006
Wilkins and others 1993
Senyuva and others 2009
Odeh and others 2007
Odeh and others 2007
Moreira and others 2002
Moreira and de Maria 2005
Bianchi and others 2005
Lušić and others 2007
Odeh and others 2007
Alissandrakis and other 2009
Mannas and Altug 2007
Alissandrakis and other 2007b
Moreira and others 2002
Radovic and others 2001
de la Fuente and others 2007b
Castro-Vázquez and others 2009
Shimoda and others 1996
Bouseta and others 1992
Guyot-Declerck and others 2002
Radovic and others 2001
Radovic and others 2001
Alissandrakis and others 2005b
Piasenzotto and others 2003
de la Fuente and others 2005
Castro-Vázquez and others 2007
Radovic and others 2001
Castro-Vázquez and others 2009
White and Bryant 1996
Castro-Vázquez and others 2009
Reference
Alissandrakis and others 2007b
∗ Geographic origin: EU = Europe (honey from 3 and more Europe countries); AU = Australia; BR = Brasil; CN = China; ENG = England; ES = Spain; GR = Greece; HU = Hungary; HR = Croatia; IR = Iran; IT = Italy; FR = France; JP = Japan; NZ = New Zealand; PS = Palestine; PT = Portugal; RO =
Romania; SK = Slovakia; TR = Turkey; U.S.A. = United States of America.
∗∗ Dynamic headspace extraction with desorption tube, filled with corresponding sorbent.
Turkish pine
Oak honeydew
Nodding thistle
Honeydew
Thymelaea hirsute
Tolpis virgata
Marmeleiro
Cambara
Strawberry-tree
Fir honeydew
Thymus capitatus
Thyme
Cashew
Rape
Willow
Rosemary
Haze
Acacia
Sunflower
Cotton
Lavender
Honey source
Citrus
Table 1– (Continued)
Floral markers of honey . . .
c 2010 Institute of Food Technologists®
Floral markers of honey . . .
Figure 2–Structures of selected volatile
compounds found in honey.
HO
O
O
O
O
O
methyl-p-anisaldehyde
trans-oak lactone
OH
cinnamic acid
OH
O
OH
OH
O
eugenol
isovaleric acid
limonene diol
O
O
O
O
NH2
methyl anthranilate
α-pinene oxide
isophorone
O
O
O
O
O
OH
dill ether
lilac aldehyde
methyl salicylate
originating from rhododendron, Christ’s thorn, eucalyptus, and
honeydew (Table 2).
Probably, the number of volatile compounds suggested as floral
markers of particular honey samples might be reduced by applying standardized isolation and analysis protocols, which are not
currently used for the scientific purposes. For instance, due to a
high concentration of carbohydrates special pretreatment of honey
is required before the isolation of volatile compounds, except for
the headspace method (Soria and others 2004; Jerković and others
2009a, 2009b). However, at least 4 different techniques for the
extraction of honey volatile compounds with various modifications were used and this might influence the results. The most
frequently used techniques in the reviewed studies were a SPME
(Kaškonienė and others 2008; Senyuva and others 2009); simultaneous steam distillation-dichlormethane extraction, also called
Likens-Nickerson (Bouseta and Collin 1995; Castro-Vázquez and
others 2009), purge and trap technique (Radovic and others 2001;
Moreira and others 2002), ultrasonic solvent extraction (Jerković
and Marijanović 2009; Jerković and others 2009b), and others
(extraction methods used for the analysis of floral markers are also
listed in Table 1). The SPME was used not only for the extraction
of aroma compounds, but also for the determination of honey
pollutants (Bentivenga and others 2004). It may be concluded
that none of the applied methods can be considered as an ideal
and absolutely representative; all of them possess advantages and
disadvantages, which may have bigger or smaller impact on the
identification and interpretation of the compositional peculiarities
of volatile compounds. Some isolation techniques, such as Likens-
c 2010 Institute of Food Technologists®
Nickerson are relatively long (up to 2 h) and require rather high
quantity of the sample (Bouseta and Collin 1995); in addition,
isolation of low molecular weight and higher boiling temperature
compounds, for example, benzoic acid is rather limited and the
use of high temperatures results in the formation of thermally
produced artifacts, such as Strecker degradation and Maillard reaction products (Jerković and Marijanović 2009; Jerković and others
2009a). In case of solvent extraction, the loss of volatiles and/or
formation of new compounds may also occur during solvent evaporation (Kaškonienė and other 2008). Different modifications of
headspace analysis, for example, using static and dynamic conditions enable to avoid above-mentioned disadvantages of methods
involving the use of organic solvents. However, solvent extraction
isolates a higher number of compounds, including less volatile
ones, among which marker compounds may be found; therefore,
the use of both techniques is advisable in comprehensive analysis
of honey.
Table 1 shows the most important volatile compounds in some
unifloral honeys from different geographic regions, identified by
various authors with different isolation techniques. Not all volatile
compounds have a significant impact on honey aroma. In general,
the impact of a given compound depends on the extent to which
the concentration exceeds its odor threshold; however, the possibilities of synergistic and/or antagonistic interactions between
different components should also be taken into account. Thus,
even compounds present in low concentrations may contribute
to honey aroma. However, the use of combined instrumental
and sensory assessment methods, such as gas chromatography and
Vol. 9, 2010 r Comprehensive Reviews in Food Science and Food Safety 625
Floral markers of honey . . .
Odor description
Bitter almond, fragrant, aromatic, sweet
Harsh, green (reminiscent of hyacinth on dilution)
Pungent, warm
Citrus, pine
Strong, penetrating, sweet, orange peel odor; citrus taste
Cabbage, sulfur, gasoline
Vegetable, cabbage, putrid
Powerful, fish, diffusive, penetrating (reminiscent of fresh onion)
Ethereal, sharp, wine-brandy-like, reminiscent of pineapple
Bread, almond, sweet, woody, fragrant
Rancid, sour sweet-like, fatty
Hyacinth
Sweet, floral, lavender, refreshing, citrus, orange, clean
Sweet, musty, aldehydic
Sweet, fruity, pineapple
Sweet woody, floral, creamy
Flowery, sweet champagne
Flowery, fresh
Phenolic Compounds as Floral Markers
∗ NA = not available.
Compound
Benzaldehyde
Benzeneacetaldehyde
Carvacrol
p-Cymenene
Decanal
Dimethyl sulfide
Dimethyl disulfide
Dimethyl trisulfide
Ethyl acetate
Furfural
Heptanoic acid
Hotrienol
Linalool
2-Methylbutanal+3-methylbutanal
Ethyl butyrate
trans-Linalool oxide
Acetic acid phenylethyl ester
Lilac aldehyde
Table 2– Odor description of some volatile compounds found in honeys.
Odor threshold
0.10 to 4.60 ppm
4.00 ppb
2.30 ppm
NA∗
0.10 ppb
NA
NA
NA
0.01 to 5.00 ppm
0.28 to 8.00 ppm
0.64 to 10.40 ppm
NA
4.00 to 10.00 ppb
NA
NA
0.32 ppm
NA
0.20 to 22.00 ng
Reference
Burdock 2001
Burdock 2001
Burdock 2001
Burdock 2001; Flavornet 2004
Leffingwell 2002
Flavornet 2004
Flavornet 2004
Burdock 2001
Burdock 2001; Leffingwell 2002
Burdock 2001; Flavornet 2004
Burdock 2001
Flavornet 2004
Burdock 2001; Flavornet 2004; Leffingwell 2002
Wardencki and others 2009
Wardencki and others 2009
Burdock 2001; Flavornet 2004; Leffingwell 2002
Wardencki and others 2009
Leffingwell 2002
olfactometry, which may provide important information on a specific compound (or compounds) responsible for aroma are rather
scarce in the studies of volatile constituents of honey (Zhou and
others 2002; Moreira and De Maria 2005; Wardencki and others
2009). Some individual volatile compounds identified in various
honey samples can be characterized by a wide range of aroma
descriptors, for example, from bitter, rancid, or fishy, to sweet
and flowery (Table 2). Although the variability of honey flavors
depends mainly on its floral origin, the techniques used in the
isolation of volatiles as well as their detection may also play an
important role in analysis results (Anklam 1998; Pérez and others 2002; Alissandrakis and others 2005a). There are many isolation techniques of volatile components, which may be applied
to honey. Depending on the isolation technique, identified compounds in the same honey types, even from the same country, can
be different.
As mentioned above, in some studies, the same volatile compound was attributed to several honeys; for example, lilac aldehydes were suggested as characteristic constituents of rhododendron
(Senyuva and others 2009), citrus (de la Fuente and others 2005;
Alissandrakis and others 2007a), rosemary (Castro-Vázquez and
others 2009), and haze honeys (Shimoda and others 1996), while
other important volatile components for these honeys were different (Table 1). However, in the majority of published articles several
different aroma compounds present at different proportions were
specified for distinguishing honeys of different botanical origin.
Taking into account the fact that apart from a floral source, several
other factors, such that of geographic origin, climatic conditions,
maturity, and processing of honey, as well as isolation technique
and contribution of other plants to the production of the honey
may influence the composition of honey volatile compounds, it
may be concluded that the measurement of volatile compounds
in most cases does not provide sufficient chemical information
for the precise determination of the botanical and/or geographic
origin of honey. Possibly, further developments in analysis techniques will enable to identify and quantify more specific volatile
constituents of honey, which might be present in very low concentrations and, which might provide more reliable information
on the floral source of honey. The reliability of the results may also
be improved by increasing the amount of samples and comparing
with honeys from other floral origins, because volatile compounds
present in very low concentrations may be the result of contribution of minor floral nectar and could therefore be misinterpreted.
The main sources of honey phenolic compounds are plants.
Plants biosynthesize a great number of various phytochemicals,
which may possess health-promoting properties, antioxidants being the major group of bioactive constituents; they are considered as natural agents, which might reduce the risk of oxidative
damage in living cells (The National Honey Board 2002). Many
studies show that honey, depending on the floral source, possesses
higher or lower antioxidant, antibacterial, or radical-scavenging
activity (Weston and others 1999; Gheldof and Engeseth 2002;
Baltrušaitytė and others 2007a, 2007b). Bioactive substances may
be transferred from the plant to the nectar and further to honey by
imparting to the final product different properties that may reasonably depend on the floral origin of honey. For instance, different
flavonoid profiles were reported for honeys collected from various
floral sources. The main phytochemicals reported in honey are
flavonoids: pinobanksin, pinocembrin, quercetin, chrysin, galangin, luteolin, and kaempferol (Martos and others 2000; Gheldof
626 Comprehensive Reviews in Food Science and Food Safety r Vol. 9, 2010
c 2010 Institute of Food Technologists®
Floral markers of honey . . .
Figure 3–The major phenolic compounds
identified in honey.
O
HO
HO
HO
O
OH
OH
O
HO
OH
O
O
O
homogentisic acid
ferulic acid
syringic acid
OH
OH
HO
HO
O
O
OH
OH
OH
OH
O
galangin
O
quercetin
OH
OH
HO
OH
O
HO
O
OH
OH
OH
OH
O
OH
HO
O
kaempferol
myricetin
HO
OH
O
OH
O
OH
O
N
OH
OH
OH
HO
OH
kynurenic acid
O
O
rosmarinic acid
abscisic acid
and others 2002); while pinocembrin, pinobanksin, and chrysin
are characteristic flavonoids of propolis; these flavonoids have
been determined in the majority of previously analyzed European
honey samples (Andrade and others 1997a; Yao and others 2004a).
The structures of the most important honey phenolic compounds
are presented in Figure 3.
The composition of phytochemicals has an influence on the
biological activity of honey; usually, the same compounds have
antioxidant and antimicrobial activity. For example, syringic acid,
methyl syringate, cinnamic acid, caffeic acid, phenyllactic acid,
pinocembrin, pinobanksin, chrysin, and galangin were attributed
to the antibacterial substances of New Zealand manuka honey
(Weston and others 1999), while Gheldof and others (2002) classified these compounds as antioxidants.
It was reported that the composition of honey depends on the
floral source used to collect nectar; however, seasonal and environmental factors, as well as processing, may also have an effect on the
composition of phenolic compounds in honey (Frankel and oth-
c 2010 Institute of Food Technologists®
ers 1998; Gheldof and Engeseth 2002; Gheldof and others 2002;
Schramm and others 2003; Yao and others 2003). Some studies have suggested possible correlations between floral origin and
flavonoid profiles (Anklam 1998; Yao and others 2004b). Therefore, predominance of some individual components or a group of
compounds in honey may be a promising marker for the determination of honey botanical origin. For example, quercetin was
suggested as a marker for sunflower honey (Tomás-Barberán and
others 2001); 8-methoxykaempferol was the main compound in
rosemary (Ferreres and others 1994a); flavanone hesperitin in citrus honey (Ferreres and others 1993). Naringenin (Andrade and
others 1997b) and luteolin (Ferreres and others 1994a) were suggested as markers of lavender honey. Characteristic compounds to
some unifloral honeys are listed in Table 3. In the case of phenolic components, the extraction method may also have an impact
on the measurement of the compounds selected as candidates of
being floral markers for some types of honey (Michalkiewicz and
others 2008). Geographic origin of honey is another important
Vol. 9, 2010 r Comprehensive Reviews in Food Science and Food Safety 627
Floral markers of honey . . .
Table 3– Phenolic and some other compounds characteristic to unifloral honeys.
Honey source
Compound
Extraction method
GO∗
Strawberry-tree
Homogentisic acid
Gluconic acid
2,5-Dihydroxyphenylacetic acid and
α-isophorone
Homogentisic, (±)-2-cis,4-trans,
(±)-2-trans,4-trans-abscisic acids, unedone
Methyl syringate
Kaempferol
8-Methoxykaempferol
Kaempferol, 8-methoxykaempferol
Kaempferol rhamnosides
Ferulic acid, acacetin
Cinnamic acid derivatives
cis, trans- and trans, trans- abscisic acids,
ellagic acid
Myricetin, tricetin, luteolin, quercetin,
kaempferol
Benzoic acid derivatives
Quercetin and 6 unidentified compounds
Gallic acid
Abscisic acid
Caffeic acid, p-coumaric acid, ferulic acid
4-Hydroxybenzoic, DL-p-hydroxyphenyllactic,
ferulic, and phenylacetic acids
Quercetin, luteolin, quercetin 3-methyl ether
Gallic acid, abscisic acid
2-Methoxybenzoic and trimethoxybenzoic
acids, and methylglyoxal
Methoxyphenyllactic acid
cis, trans- and trans, trans- abscisic acids,
quercetin, kaempferol,
8-methoxykaempferol, unknown compound
Quercetin, hesperetin, and chrysin
Hesperitin
Solvent
SPE
No extraction∗∗
Sardinia (IT)
IT
EU
Cabras and others 1999
Cherchi and others 1994
Donarski and others 2010
Solvent
Sardinia (IT)
Tuberoso and others 2010
Solvent
SPE
SPE
SPE
SPE
SPE
SPE
SPE
Sardinia (IT)
ES
ES
EU
IT and SK
Transylvania (RO)
EU
EU
SPE
EU
Martos and others 2000
SPE
SPE
SPE
SPE
SPE
SPE
EU
EU
AU
HR
IT
EU
Dimitrova and others 2007
Tomás-Barberán and others 2001
Yao and others 2004a
Kenjeric and others 2008
Cherchi and others 1994
Dimitrova and others 2007
SPE
SPE
SPE
AU and NZ
NZ
NZ
SPE
SPE
NZ
EU
Stephens and others 2010
Tomás-Barberán and others 2001
SPE
SPE
IR
EU
Hesperitin, methyl anthranilate
Caffeic acid, p-coumaric acid, ferulic acid, and
hesperetin
Quercetin, quercetin 3, 3-dimethyl ether,
myricetin, luteolin
p-Coumaric, ferulic, and caffeic acids
Quercetin
cis, trans- and trans, trans- abscisic acids
Ellagic acid
DL-β-Phenyllactic acid, phenylacetic and
benzoic acids
Myricetin, myricetin 3-methyl ether, myricetin
3’-methyl ether, tricetin
Ellagic, p-hydroxybenzoic, syringic, and
o-coumaric acids
Rosmarinic acid
3-Hydroxybenzoic acid
Naringenin
Luteolin
Gallic and caffeic acids
Gallic acid
SPE
Solvent
ES
CN
Hadjmohammadi and others 2009
Ferreres and others 1993;
Tomás-Barberán and others 2001
Ferreres and others 1994b
Liang and others 2009
SPE
AU
Yao and others 2004b
SPE
SPE
SPE
SPE
SPE
EU
EU
PT
EU
EU
Dimitrova and others 2007
Tomás-Barberán and others 2001
Ferreres and others 1996
Andrade and others 1997b
Dimitrova and others 2007
SPE
PT
Ferreres and others 1994c
SPE
PT
Andrade and others 1997a
SPE
SPE
SPE
SPE
SPE
SPE
EU
EU
EU
ES
EU
PT
Andrade and others 1997b
Dimitrova and others 2007
Andrade and others 1997b
Ferreres and others 1994a
Dimitrova and others 2007
Andrade and others 1997a
Asphodel
Rosemary
Acacia
Eucalyptus
Sage
Chestnut
Leptospermum
Manuka
Kanuka
Rape
Citrus
Sunflower
Heather
Thyme
Lime tree
Lavander
Reference
Tuberoso and others 2009
Gil and others 1995
Ferreres and others 1994a
Tomás-Barberán and others 2001
Truchado and others 2008
Bobis and others 2007
Dimitrova and others 2007
Tomás-Barberán and others 2001
Yao and others 2003
Yao and others 2003
Stephens and others 2010
∗ Geographic origin: abbreviations see Table 1.
∗∗ Without extraction, only preparation for 1 H NMR spectroscopic analysis with some reagents.
factor in the composition of phenolic constituents; for example,
chestnut and citrus honeys collected in different regions were
characterized by different compounds (Ferreres and others 1993,
1994b; Dimitrova and others 2007; Hadjmohammadi and others
2009).
Methodological aspects of determining of phenolic compounds
(Alvarez-Suarez and others 2009) and phenolic acids (Pyrzynska
and Biesaga 2009) were the subjects of recent reviews. SPE is
very often used to extract phenolic compounds from a sugar matrix. SPE on Amberlite XAD-2 sorbent described by Ferreres
and others (1994a) is probably the most popular technique applied by many researchers, but the use of C18 cartridges has also
been reported (Pulcini and others 2006; Michalkiewicz and others
2008). Various solvents have been applied for extractions of phe-
nolic compounds (Cabras and others 1999; Dimitrova and others
2007; Tuberoso and others 2009). Daher and Gülaçar (2008) used
a SPME in their study, while this method is not so popular for the
extraction of flavonoids and nonflavonoid phenolic compounds.
It was shown that the choice of the extraction techniques of phenolic compounds might give different results for the same type of
honey. For example, 2 different characteristic compounds were reported for strawberry-tree honey from Italy, which was extracted
by different methods (Cherchi and others 1994; Cabras and others
1999); sunflower and lavender honeys, extracted by the SPE were
also characterized by different compounds (Table 3). However,
it is possible that in the above-mentioned cases some unconsidered factors, such as bee subspecies, the contribution of nectar or
honeydew from other plants, climatic conditions, and others also
628 Comprehensive Reviews in Food Science and Food Safety r Vol. 9, 2010
c 2010 Institute of Food Technologists®
c 2010 Institute of Food Technologists®
Weston and Brocklebank 1999
NZ
Melezitose, panose, and maltotriose
pulsed amperometric detector; ∗∗∗∗ refractive index detector.
TR
HPLC-PAD
Senyuva and others 2009
HPLC –RIF∗∗∗∗
Heather, spike lavender, French lavender, and forest
honeys
Rhododendron, chestnut, honeydew, anzer (Thymus
spp.), eucalyptus, gossypium, and multifloral
Honeydew
Rosemary, citrus, lavender, sunflower, eucalyptus,
heather, forest
Acacia
FR
FR
Acacia and chestnut
Acacia, chestnut, rape, lavender, fir, linden, sunflower
FR
Rape and sunflower
∗ Geographic origin: abbreviations see Table 1.
∗∗ Gas chromatographic analysis of honey was performed after derivatization reaction with trimethylsilyloxime; ∗∗∗
Nozal and others 2005
HPLC-PAD
Province of Soria (ES)
Horváth and Molnár-Perl 1997
GC-MS
HU
Mateo and Bosch-Reig 1998
GC-FID
ES
Cotte and others 2004a
Cotte and others 2004a
GC-FID and LC-PAD
GC-FID and LC-PAD
Cotte and others 2004a
Cotte and others 2004a
GC-FID and LC-PAD∗∗∗
FR
GC-FID and LC-PAD
Reference
Mateo and Bosch-Reig 1997
Analysis method
GC-FID∗∗
Suggested compounds or criteria
for differentiation
Fructose, glucose, sucrose, maltose, and
glucose/water ratio
High content in trisaccharides, particularly raffinose,
melezitose, and erlose
High glucose concentration, absence of raffinose,
melezitose, and erlose
High fructose/glucose ratio
Glucose, fructose, raffinose, trehalose, F/G,
maltose/trehalose, and erlose/maltulose
Fructose and sucrose, electrical conductivity, color (x,
y, L), water content
High fructose, sucrose, and minor oligosacharides
content
Erlose, nigerose, trehalose, melezitose, isomaltose,
and panose
Maltose and raffinose
Sugars (saccharides) are the main components of honey. The
attempts to differentiate various honey types by carbohydrate
composition so far were based mostly on quantitative composition of mono- and oligosaccharides, while studies on the possible qualitative differences in sugar composition are rather scarce.
Monosaccharides fructose and glucose are the major constituents
of honeys, the former being the dominant component in almost
all types, except for some honeys of dandelion (Taraxacum officinale), blue curl (Trichostema lanceolatum), and rape (Brassica napus) origin, where glucose is present in higher amounts (Cavia
and others 2002). The concentrations of fructose and glucose, as
well as their ratio, have been advanced as useful indicators for
the classification of unifloral honeys (Persano Oddo and others
1995; Persano Oddo and Piro 2004). Besides these 2 main constituents, there are also oligosaccharides. Up to the 1990s, about
25 oligosaccharides (disaccharides, trisaccharides, tetrasaccharides)
have been found in honey (Anklam 1998); however, the number of identified oligosaccharides is now increasing. For instance,
most recently 25 trisaccharides and 10 tetrasaccharides, which are
formed mainly by the action of honey enzymes, were reported for
Spanish and New Zealand honeys by Ruiz-Matute and others
(2010). The trisaccharides planteose and α-3 -glucosylisomaltose
were reported in honey for the 1st time by the same authors.
Thus, new developments in analytical techniques enhance the
possibilities of searching for more precise and representative floral/botanical origin markers. Dvash and others (2002) used NIR
spectroscopy for the analysis of an avocado (Persea Americana Mill.)
honey and found that carbohydrate alcohol perseitol (d-glycerod-galacto-heptitol) in spite of its low concentration could be used
as a marker for avocado honey: the average value of this compound in 109 analyzed honey samples was 0.48 g/100 g. The
same compound was reported in an avocado honey by de la Fuente
and others (2007b) and it was present even at a higher amount,
0.75 g/100 g.
Published data show that it is difficult to specify 1 or more
carbohydrates, which could serve as floral markers for honey. Instead, some authors have suggested to use the ratio of particular
carbohydrates and also other criteria, such as water activity, electrical conductivity, together with the amount of carbohydrates for
differentiation of honeys (Table 4).
Cellobiose, gentiobiose, isomaltose, kojibiose, laminaribiose,
maltose, maltulose, melibiose, nigerose, palatinose, trehalose, trehalulose, turanose, and sucrose are the main disaccharides found
in honey (Sanz and others 2004; de la Fuente and others 2007b).
However, it would be rather difficult to identify the predominant
disaccharide or certain combinations in the previously studied
honey types. For example, maltulose and turanose were found
Table 4– Carbohydrate composition, their ratios, and other criteria suggested for the discrimination of unifloral honeys.
Carbohydrates as Floral Markers
Honey sources
Rosemary, orange blossom, lavender, sunflower,
eucalyptus, heather, honeydew
Fir
had an influence on the accumulation of individual compounds in
honey.
It may be concluded that the presence of many factors, which
might have an effect on the composition of phenolic constituents
in honey as well as structural variety of such constituents that are
biosynthesized by the floral sources of honey, makes it difficult
to use the presence of phenolic compounds for the precise determination of the botanical origin of honey. However, it may
be expected that more future and comprehensive investigations
on the effects of these factors on honey composition will provide
more reliable indicators for the interconnection of honey and its
botanical source by using key phenolic compounds present both
in the plant and the honey.
GO∗
ES
Floral markers of honey . . .
Vol. 9, 2010 r Comprehensive Reviews in Food Science and Food Safety 629
Floral markers of honey . . .
in many honey samples, however their concentrations varied to
a wide extent. Thus, Sanz and others (2004) found the highest amounts of maltulose and turanose (0.66 to 3.52 and 0.72 to
2.87 g/100 g of honey, respectively) in 10 samples of honey from
different regions of Spain and commercially available nectar and
honeydew honeys (unfortunately, botanical composition was unknown). The same compounds were predominant in other Spanish
honeys of different origin (de la Fuente and others 2007b): the
amount of maltulose varied from 0.55 in Satureja montana to 4.34
g/100 g in an avocado honey, while that of turanose from 0.89
in Satureja montana to 4.56 g/100 g in honey of fir origin. Mateo
and Bosch-Reig (1997) reported that the total content of maltose,
nigerose, and turanose was highest in their tested Spanish honey
samples of rosemary, lavender, sunflower, eucalyptus, heather, and
honeydew origin, except for orange blossom honey in which sucrose was the highest out of all disaccharides. In addition, the
amount of kojibiose was higher than that of sucrose, maltulose,
or isomaltose in all the analyzed samples, except for orange blossom honey. In France, Cotte and others (2004a) determined the
predominant disaccharides in some honeys: maltose and turanose
in acacia; maltulose and turanose in chestnut and linden; turanose
and trehalose in fir; and sucrose and maltose in lavender origin
honey. Maltose was the major disaccharide present in 80 genuine
Brazilian honey samples (mostly Eucalyptus spp., extra-floral, and
multifloral honeys) with a mean value of 3.05 g/100 g (Da Costa
Leite and others 2000); possibly, in this case maltose may be considered for a geographic classification of honey. It was suggested,
that the dominating disaccharide might depend on the botanical source of honey (Persano Oddo and others 1995; Nozal and
others 2005); however, geographic region may be another factor
influencing the composition of honey carbohydrates.
It is known that there are differences in the composition of carbohydrates between honeydew and blossom honeys. Honeydew
honey is characterized by a higher concentration of oligosaccharides, mainly the trisaccharides melezitose and raffinose, which
usually are not found in blossom honeys (Bogdanov and others
2004). Panose, isomaltotriose, maltotriose, and erlose also may be
found in some kinds of honey such as heather, spike lavender,
French lavender, thyme, forest, and multifloral collected in the
province of Soria in Spain (Nozal and others 2005).
The advantages of discriminant analysis were suggested for an
advanced classification of honeys by their carbohydrate composition. Thus, Nozal and others (2005) used canonical discriminant
analysis for the identification of 5 kinds of honeys out of 65.
Heather honey was noted for the presence of erlose and nigerose;
forest honey contained higher amounts of trehalose and melezitose; spike lavender honey was specified by isomaltose; while
French lavender and thyme honeys were characterized by panose.
The ratio between some carbohydrates is another indicator
that may be used to ascertain honey authenticity (Horváth and
Molnár-Perl 1997; Nozal and others 2005). Thus, the ratios of
fructose/glucose, maltose/isomaltose, sucrose/turanose, and maltose/turanose, maltotriose/raffinose+erlose+melezitose were used
for the authentification of unifloral honeys. For example, an acacia honey was distinguished by a high maltose/isomaltose ratio
(11:1 to 25:9), while this ratio for linden and honeydew honeys
was remarkably lower, 2.2 and 2.0 to 2.5, respectively (Horváth
and Molnár-Perl 1997). Rape, dandelion, and blue curl origin
honeys were distinguished by a low fructose/glucose ratio (<1)
due to the predominant monosaccharide glucose in these honeys (Horváth and Molnár-Perl 1997; Cavia and others 2002).
However, Kaškonienė and others (2010) determined that the
fructose/glucose ratio was also lower than 1 for willow honey,
except for 1 sample out of 7 analyzed; in the same study, it was
found that willow pollen content in honey strongly correlated
with the electrical conductivity of honey. de la Fuente and others
(2007b) analyzed a sample of honey of willow origin from Spain
and obtained different results on monosaccharide amounts and
their ratio, which might indicate on the prevailing influence of geographic origin on sugar composition than the floral source itself;
however considering the fact that only 1 willow sample was analyzed, this assumption may be regarded as a very preliminary one.
Senyuva and others (2009) analyzed 70 honey samples of 9
different floral types, namely rhododendron, chestnut, honeydew, anzer (Thymus spp.), eucalyptus, gossypium, citrus, sunflower, and multifloral from 15 different geographic regions of
Turkey and found that a combination of the contents of raffinose,
maltose, and sucrose with volatile compounds like phthalic acid
or 2-methylheptanoic acid in chemometric analysis was a sufficiently informative indicator for differentiating honeys according
to botanical and geographic origin. However, it should be noted
that phthalic acid is not a natural component and therefore it was
probably a contaminant.
It may be concluded that sugar composition is a more reliable indicator for honey classification and authentication only in the case
of unifloral honeys with very high amount of dominating plant;
when the fraction of dominating plant of nectar source reduces,
the interpretation of the results of the measurement of saccharides
becomes more difficult and almost unusable in determining floral
origin of such honeys.
Amino Acids as Possible Floral Origin Markers
Amino acids are another group of compounds that were used
for the characterization and differentiation of honey sources. Proline is the main amino acid present in honey; it is added by the bee
(Anklam 1998; Bogdanov and others 2004) and its amount varies
depending on the floral source. For instance, the highest values
were reported in honey of Thymus origin, while the lowest were in
honey samples from Rhododendron, Brassica, and Robinia (Persano
Oddo and Piro 2004). Cotte and others (2004b) suggested that
high amounts of threonine and phenylalanine are characteristic to
sunflower and lavender honeys, respectively, while, Hermosı́n and
others (2003) found that lavender honey contained a high concentration of tyrosine. Probably, the content and the composition of
amino acids in honey are also dependent on its geographic origin;
Cotte and others (2004b) analyzed honey samples from France,
while Hermosı́n and others (2003) studied honey from Spain.
Iglesias and others (2004) used amino acids for the discrimination
of honeydew and floral honeys from a small geographic area and
found that 21 measured amino acids were in higher concentration in honeydew honeys than in floral ones, except for tyrosine
and phenylalanine. The same study showed that the content in
glutamic acid and tryptophan enables the differentiation between
floral honeys and honeydew honeys.
Fifty-six honey samples from 3 different regions in Argentina
were differentiated by the content of 13 free amino acids using
statistical analysis methods; the cluster analysis showed that the
analyzed samples may be grouped into 9 clusters related to sampling regions and more strictly to apiarian flora around apiaries
(Cometto and others 2003). Phenylalanine and tyrosine, as well
as some other amino acids, were suggested by principal component analysis (PCA) as positively or negatively correlating amino
acids with a column group of honeys. The authors used between
5 and 7 amino acids to distinguish honeys from different regions
630 Comprehensive Reviews in Food Science and Food Safety r Vol. 9, 2010
c 2010 Institute of Food Technologists®
Floral markers of honey . . .
and concluded that the applied method can be used to verify
both the botanical or geographic origin of honey. The study with
Turkish honeys also showed that the phenylalanine and tyrosine
contents may contribute to distinguishing honeys, with valine,
leucine, and isoleucine being other important amino acids for
recognizing botanical and geographic origins (Senyuva and others 2009). Furthermore, the authors suggested combining 1 or 2
amino acids with other phytochemicals such as volatile compounds
or oligosaccharides to achieve improved discrimination. Anyway,
data on honey amino acids are too scarce to make sound assumptions on the use of these components in the characterization of
different honey types.
Determination of proteins was also applied for tracing floral
origin of honey. Recently, Wang and others (2009) suggested
new method of determination geographic and botanical origin of
honey, which is based on matrix assisted laser desorption ionization time-of-flight mass spectrometry (MALDI TOF MS) technology using honey protein fingerprinting and barcoding. It may
be promising techniques, however nowadays it is rather expensive
and needs huge work to collect “barcodes” from different honeys to be used for their authentication. Almost 10 y ago, Baroni
and others (2002) applied immunoblot assays for the analysis of
honey proteins and showed that it was possible differentiating the
pollen originated from sunflowers and Eucalyptus sp. This method
was suggested as an alternative or complementary method to the
melissopalynological analysis.
Other Organic Compounds and Microelements as
Possible Floral Origin Markers
Other classes of chemical compounds have also been used for
the characterization of honeys. Beretta and others (2008) applied
1
H NMR to test the applicability of the techniques to assign reliable markers for European honeys and found that honeydew honey
could be characterized by the presence of aliphatic compounds (assessed by the characteristic resonance signals), chestnut honey by
kynurenic acid (4-hydroxyquinaldic acid) and its related metabolites, and linden honey by free and/or conjugated cyclohexa-1,3diene-1-carboxylic acid. Truchado and others (2009) used 1 H
NMR and 13 C NMR to analyze chestnut nectar, which is collected by the bees to make honey and identified kynurenic acid
and 4-quinolone-2-carboxylic acid; these compounds were also
found in chestnut honey and attributed as possible markers of this
kind of honey. Kynurenic acid as a biomarker of sweet chestnut
honey was also confirmed most recently by Donarski and others
(2010) by 1 H NMR spectroscopic analysis.
Analysis of microelements in honey in the last decade was
used mainly for the determination of environmental contamination of honey (Devillers and others 2002; Tuzen and others
2007; Yarsan and others 2007); however, some studies of elemental honey composition were also aimed at determining its
authenticity (Fernandez-Torres and others 2005; Chudzinska and
Baralkiewicz 2010). Rapid advances in high-resolution methods
and instrumentation have remarkably improved the opportunities to differentiate honeys according to their elemental composition, particularly at trace levels. A flame and graphite furnace
atomic absorption (Tuzen and others 2007), inductively coupled
plasma atomic emission (Devillers and others 2002), and inductively coupled plasma-mass spectrometry (ICP-MS) (Chudzinska
and Baralkiewicz 2010) procedures were used for the measurement of trace elements (microelements) in honey. For instance,
ICP-MS together with multivariate analysis was applied in the
analysis of honeydew, buckwheat, and rape honey samples from
c 2010 Institute of Food Technologists®
Poland, which were consequently classified by this technique into
2 main groups, honeydew honey and nectar honey (Chudzinska
and Baralkiewicz 2010). Honeydew honey was distinguished by
the content of K, Al, Ni, Cd, and Zn, while Na, Ba, and Pb were
selected as indicators for rape honey. Fernandez-Torres and others
(2005) found that the concentrations of Zn, Mn, Mg, and Na in
eucalyptus, heather, orange-blossom, and rosemary honeys from
Spain were strongly dependent on their botanical origin.
Some risks should be carefully considered when using microelements for the authentication of honey. For instance, some minerals,
such as Na and K are of floral origin because they are accumulated
in the plant cells and depend on the content of enzymes, while
some metals, particularly Pb and Cd, may be present in honey due
to the environmental contamination. Therefore, the concentration of such metals in honey may be dependent on contamination
of the environment to a larger extent than to the floral and/or
geographic origin of the honey.
Summary
Chemical analysis of honey may provide useful data that may be
associated and correlated with the floral origin of honey or honey
products, particularly when the product comes from a comparatively uniform floral source. The composition of volatile compounds, phenolic acids, sugars, and some other constituents have
been related to the floral origin of honey; however, it may be concluded that due to the other extrinsic as well as intrinsic factors,
which may influence honey composition, floral phytochemical
markers investigated up to now are not sufficiently reliable indicators for honey authentication by defining its floral source and
geographic origin. It is also evident that honey is too complicated
in terms of the floral sources used for its collection; the cases when
bees collect nectar from a single plant species are very rare. Consequently, when the amount of a dominant type of pollen (such
as rape) decreases, then the amount of another pollen (such as
willow, clover, dandelion, cornflower) increases. Every plant used
for collecting honey nectar may have an influence on the composition of honey. The higher the fraction of a mixture of minor
pollen, the more difficult is the differentiation of honeys by using
possible phytochemical indicators associated with the dominating
pollen. The data reviewed on floral markers suggest that better
classification of various honeys can be achieved by using groups
of compounds (volatiles, phenolic compounds, amino acids, and
carbohydrates), possibly including other supplementary parameters, such as electrical conductivity, color, and enzyme activity. In
addition, the application of multivariate statistical analysis on the
authentication and classification as was demonstrated by Tzouros
and Arvanitoyannis (2001) for many agricultural products, including honey, might be an important complementary tool for a more
reliable identification and quality control method of honey.
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634 Comprehensive Reviews in Food Science and Food Safety r Vol. 9, 2010
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