Activities of different types of Thai honey on pathogenic bacteria causing skin diseases, tyrosinase enzyme and generating free radicals

Abstract

Honey is a natural product obtained from the nectar that is collected from flowers by bees. It has several properties, including those of being food and supplementary diet, and it can be used in cosmetic products. Honey imparts pharmaceutical properties since it has antibacterial and antioxidant activities. The antibacterial and antioxidant activities of Thai honey were investigated in this study.
The honey from longan flower (source No. 1) gave the highest activity on MRSA when compared to the other types of honey, with a minimum inhibitory concentration of 12.5% (v/v) and minimum bactericidal concentration of 25% (v/v).
Honey demonstrates tremendous potential as a useful source that provides anti-free radicals, anti-tyrosinase and anti-bacterial activity against pathogenic bacteria causing skin diseases.

Full text

R E S E A R C H A R T I C L E Open Access
Activities of different types of Thai honey on
pathogenic bacteria causing skin diseases,
tyrosinase enzyme and generating free radicals
Kanyaluck Jantakee and Yingmanee Tragoolpua
*
Abstract
Background: Honey is a natural product obtained from the nectar that is collected from flowers by bees. It has
several properties, including those of being food and supplementary diet, and it can be used in cosmetic products.
Honey imparts pharmaceutical properties since it has antibacterial and antioxidant activities. The antibacterial and
antioxidant activities of Thai honey were investigated in this study.
Results: The honey from longan flower (source No. 1) gave the highest activity on MRSA when compared to the other
types of honey, with a minimum inhibitory concentration of 12.5% (v/v) and minimum bactericidal concentration of
25% (v/v).
Moreover, it was found that MRSA isolate 49 and S. aureus were completely inhibited by the 50% (v/v) longan honey
(source No. 1) at 8 and 20 hours of treatment, respectively. Furthermore, it was observed that the honey from coffee
pollen (source No. 4) showed the highest phenolic and flavonoid compounds by 734.76 mg gallic/kg of honey and
178.31 mg quercetin/kg of honey, respectively. The antioxidant activity of the honey obtained from coffee pollen was
also found to be the highest, when investigated using FRAP and DPPH assay, with 1781.77 mg FeSO
4
•7H
2
O/kg of
honey and 86.20 mg gallic/kg of honey, respectively. Additionally, inhibition of tyrosinase enzyme was found that
honey from coffee flower showed highest inhibition by 63.46%.
Conclusions: Honey demonstrates tremendous potential as a useful source that provides anti-free radicals,
anti-tyrosinase and anti-bacterial activity against pathogenic bacteria causing skin diseases.
Keywords: Anti-bacterial activity, Pathogenic bacteria, Anti-free radicals, Anti-tyrosinase
Background
Infection of skin by pathogenic bacteria is one of the
major health concerns in Thailand as well as the rest of
the world. Skin and wound infections are frequently
caused by pathogenic bacteria such as Staphylococcus
aureus,Staphylococcus epidermidis,Micrococcus luteus,
Streptococcus pyogenes,Pseudomonas aeruginosa,Escheri-
chia coli,Klebsiella pneumoniae, methicillin resistant or
sensitive S. aureus (MRSA or MSSA), and vancomycin
resistant Enterococci [1-3]. The development of resistant
bacterial strains resulting from excessive use of antibiotics
has limited the efficacy of currently available antibiotics.
Therefore, it is imperative that new pharmaceutical agents
are developed for the effective treatment of diseases.
Honey is a natural product obtained from the nectar
that is collected from flowers by bees. It has several
properties, including those of being food and supple-
mentary diet, and it can be used in cosmetic products.
Honey imparts pharmaceutical properties since it has
antibacterial and antioxidant activities. Therefore, honey
has been in use since ancient times as medicine in a
wide variety of treatments [2]. In addition to high sugar
and low water activity content (a
w
), honey also contains
bioactive compounds and various enzymes [4]. Honey has
been reported to exhibit gastroprotective, antibacterial,
antifungal, antioxidant, and anti-inflammatory properties
[5-8]. It has been demonstrated to be effective against
several human bacterial pathogens, including Escherichia
coli,Enterobacter aerogenes,Salmonella typhimurium,
and Staphylococcus aureus [9]. The antibacterial activities
of honey were demonstrated by using osmotic effects,
* Correspondence: yingmanee.t@cmu.ac.th
Department of Biology, Faculty of Science, Chiang Mai University, Chiang Mai
50200, Thailand
© 2015 Jantakee and Tragoolpua; licensee BioMed Central. This is an Open Access article distributed under the terms of the
Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use,
distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public
Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this
article, unless otherwise stated.
Jantakee and Tragoolpua Biological Research 2015, 48:4
http://www.biolres.com/content/48/1/4

acidity, hydrogen peroxide generation, and phytochemical
components [3]. The hydrogen peroxide, which is produced
by the enzyme glucose oxidase, a major antibacterial agent,
showed different concentrations in the different types of
honey. In addition, it was found that some honey con-
sisted of many natural antibacterial components. The
well-studied New Zealand mānuka honey contained 1,2
dicarbonyl compound methylglyoxal (MGO), or the unique
mānuka factor (UMF), and the cationic antimicrobial
peptide bee defensin-1, which were identified as specific
antibacterial substances [10,11]. Several publications have
shown the antimicrobial properties of mānuka honey
and its activity against pathogenic bacteria including
Staphylococcus aureus,Pseudomonas aeruginosa, and
Escherichia coli [12,13]. In addition, honey contains many
substances including phenolic acids, flavonoids, ascorbic
acid, protein, carotenoid, and enzymes [14,15]. Phenolic
and flavonoids compounds are some of the most import-
ant groups of the antioxidant substances present in honey.
The diversity of the phenolic content and the antioxidant
capacity depends on the processing, handling, and storage
of honey [16]. Moreover, many phytochemicals in natural
substances have been shown as skin-whitening agents
by inhibition of tyrosinase enzyme in melanin synthesis
pathway [17]. In the biosynthetic pathway of melanin
formation, tyrosinase has a primary role in the reaction
by oxidation of tyrosine to L-3,4-dihydroxyphenylalanine
(L-DOPA) and dopaquinone [18].
In Thailand, various types of honey are produced and
consumed on a large scale; however, only a few pieces of
research literature and information on the biological
properties of honey have been reported. For that reason,
this study aimed to investigate the efficacy of different
types of Thai honey on inhibition of pathogenic bacteria on
skin. Additionally, the polyphenol, flavonoid, antioxidant
and anti-tyrosinase properties of honey were determined.
Results
Viscosity and pH of honey
The viscosity and pH values were shown in Table 1. The
pH value of honeys ranged from 3.37 ± 0.08 to 4.06 ± 0.03.
Additionally, honeys showed the viscosity of 1,083.50 ±
12.02 to 4,892.00 ± 63.64 Centipoise. Mānuka honey
showed very high viscosity so the viscosity could not be
measured by the viscometer used in this study.
Determination of activity of honey on bacteria using the
agar well diffusion assay
The results of the antibacterial assays of 16 types of honey
from 6 sources are shown in Table 2. The honey samples
at the concentration of 100% inhibited S. aureus and
MRSA.However, Corynebacterium sp. was not inhibited
by honey obtained from the pollens of forest flora (source
No.3), and honey obtained from the pollens of lychee,
sunflower, and sesame (source No.6). In addition, honey
from the pollens of longan, lychee, and polyflora (source
No.1) and honey from polyflora (source No.5, No.6)
showed activity on P. acnes. Moreover, all the tested bac-
teria were found to be inhibited by mānuka honey. Upon
testing the inhibiton of bacteria with honey obtained from
the longan pollen (source No.3), it was found that the
antibacterial activity against MRSA 49 was higher than
that exhibited by mānuka honey, with diameters of the
inhibition zone of 24.00 ± 5.20 mm. However, after
treatment of all Thai honey that used in this study with
catalase enzyme, the inhibition zones were not observed
except the treatment of mānuka honey. S. aureus, Coryne-
bacterium sp., MRSA 49, MRSA 50 and P. acneswere
inhibited by mānuka honey after treatment with cata-
lase enzyme with diameters of the inhibiton zones of
20.33 ± 0.58, 24.00 ± 1.00, 23.33 ± 2.08, 20.33 ± 0.58 and
11.33 ± 0.58 mm., respectively.
Phenol was also used as positive control. After treat-
ment S. aureus, Corynebacterium sp., MRSA 49, MRSA
50 and P. acnes by phenol at 12%, the inhibition zones
were 25.67 ± 0.58, 25.33 ± 1.52, 21.00 ± 1.73, 23.00 ± 3.60
and 28.67 ± 0.58 mm., respectively. In addition, it was
observed that M. luteus,B. subtilis,S. epidermidis, and
Ps. aeruginosa were not inhibited by honey.
Determination of minimum inhibitory concentration (MIC)
and minimum bactericidal concentration (MBC) of honey
on bacteria
The MIC and MBC values of honey obtained from the dif-
ferent pollens and the MIC and MBC values of mānuka
Table 1 pH and viscosity values of honey
Source Type of pollen Vicosity (cP) pH
1 Longan 2599.50 ± 3.54 4.02 ± 0.06
Lychee 1597.50 ± 19.09 4.06 ± 0.03
Polyflora 2940.00 ± 25.46 3.87 ± 0.11
2 Longan 3570.50 ± 47.38 3.98 ± 0.02
Sunflower 2624.00 ± 9.90 3.77 ± 0.12
Polyflora 2824.00 ± 41.01 3.98 ± 0.07
3 Longan 3481.50 ± 58.69 3.87 ± 0.10
Lychee 3332.50 ± 106.77 4.02 ± 0.04
Polyflora 2993.00 ± 42.43 3.84 ± 0.05
Forest flora 1595.00 ± 28.28 3.88 ± 0.12
4 Coffee 1083.50 ± 12.02 4.00 ± 0.09
5 Polyflora 1641.00 ± 14.14 3.84 ± 0.07
6 Lychee 1759.50 ± 21.92 3.37 ± 0.08
Sunflower 2669.50 ± 27.58 3.76 ± 0.08
Polyflora 4892.00 ± 63.64 3.79 ± 0.16
Sesame 2953.00 ± 53.74 3.57 ± 0.04
Mānuka honey - 3.85 ± 0.06
Jantakee and Tragoolpua Biological Research 2015, 48:4 Page 2 of 11
http://www.biolres.com/content/48/1/4

honey were determined and compared for antibacterial
activity using the broth dilution method. Most honey
showed antibacterial activity against all of the pathogenic
bacteria. The MIC values of honey ranged from 12.5% to
50%, and the MBC values ranged from 25% to >50%,while
the MIC values observed from mānuka honey were lower,
and ranged between 3.125% and 25% (Table 3). However,
the honey obtained from the longan and polyflora pollens
(source No.1) showed lower MIC and MBC values of
12.5% and 25%, respectively, on MRSA 49 and MRSA 50
when compared to the other varieties of honey.
Determination of time–kill endpoints of honey on
bacterial growth
The time–kill assay of honey obtained from longan
flowers (source No.1) at a concentration of 50% (v/v) was
performed against S.aureus and MRSA49 after treatment
for 10 minutes, 20 minutes, 30 minutes, 60 minutes, 90
minutes, and 120 minutes, and every 2 hours until 24
hours. The results of the bacterial growth for each time
are presented in Figure 1. The MRSA49 growth was com-
pletely inhibited by the 50% honey at 8 hours, while the S.
aureus growth was completely inhibited at 20 hours after
the treatment.
Determination of total phenolic content of honey
The total phenolic compounds were determined using
the Folin–Ciocalteu method and reported as gallic acid
equivalents with reference to the standard curve (y =
8.8325x- 0.0433, R
2
= 0.9981). The honey from coffee
flower (source No.4) had, significantly, the highest total
phenolic content (p < 0.05), of 734.76 mg gallic/kg of
honey, when compared to the other types of honey, except
mānuka honey (Table 4).
Determination of flavonoid content of honey
The total flavonoid content of the honey samples were
in the range of 29.86 mg quercetin/kg of honey to
178.31 mg quercetin/kg of honey, which was expressed
using quercetin as the standard (Table 4). The honey
from coffee flower (source No.4) showed significantly
high flavonoid content, of 178.31 mg quercetin/kg of
honey. Therefore, it can be concluded that the level of
the flavonoid content in a honey sample is dependent on
the type of flora.
Table 2 Effect of honey on growth of pathogenic bacteria causing skin disease by agar well diffusion method
Source Type of
pollen
Zone of Inhibition (mm)
Bacterial strain
S. aureus MRSA 49 MRSA 50 Corynebacterium sp. P.acnes
1 Longan 15.67 ± 1.15 19.67 ± 1.53 21.33 ± 2.31 11.66 ± 0.58 8.67 ± 0.58
Lychee 12.00 ± 0.00 11.33 ± 1.53 13.33 ± 2.52 6.50 ± 0.00 9.33 ± 0.58
Polyflora 14.33 ± 1.15 17.33 ± 1.53 16.33 ± 1.15 9.33 ± 1.15 9.67 ± 0.58
2 Longan 13.00 ± 1.00 16.33 ± 0.58 15.00 ± 0.00 11.00 ± 0.00 0.00 ± 0.00
Sunflower 9.67 ± 0.58 11.33 ± 0.53 10.33 ± 0.58 7.00 ± 0.00 0.00 ± 0.00
Polyflora 11.67 ± 0.58 15.33 ± 0.58 15.00 ± 1.00 10.00 ± 0.00 0.00 ± 0.00
3 Longan 18.33 ± 0.58 24.00 ± 5.20 22.67 ± 0.58 16.00 ± 1.00 0.00 ± 0.00
Lychee 13.330.58 19.33 ± 1.15 17.33 ± 2.52 10.33 ± 0.58 0.00 ± 0.00
Polyflora 14.33 ± 1.15 20.33 ± 2.33 19.67 ± 1.53 9.33 ± 0.58 0.00 ± 0.00
Forest flora 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00
4 Coffee 13.00 ± 1.00 16.00 ± 0.00 15.33 ± 0.58 11.00 ± 1.00 0.00 ± 0.00
5 Polyflora 13.67 ± 1.15 21.00 ± 3.61 13.67 ± 1.15 11.67 ± 0.58 8.33 ± 0.58
6 Lychee 10.00 ± 0.00 10.67 ± 0.58 11.33 ± 0.58 0.00 ± 0.00 0.00 ± 0.00
Sunflower 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00
Polyflora 13.67 ± 1.15 14.00 ± 1.00 17.33 ± 1.15 10.33 ± 0.58 7.33 ± 0.58
Sesame 0.00 ± 0.00 10.33 ± 0.58 10.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00
Mānuka honey 20.33 ± 0.58 19.67 ± 2.52 22.00 ± 1.00 22.00 ± 3.00 11.67 ± 0.58
Control solution 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00
Gentamycin 2 mg/ml 23.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 28.33 ± 0.58 34.00 ± 1.00
Gentamycin 10 mg/ml ND 9.67 ± 0.58 10.00 ± 0.00 ND ND
Note: The data are given as mean ± standard deviation (SD) of triplicate experiments. The statistical comparison between values from the different types of honey
and the bacterial strains was done using post hoc Duncan’s test. The values were found to be significantly different (p < 0.05) when compa red between different
strains of bacteria and different types of honey. ND = not determined.
Jantakee and Tragoolpua Biological Research 2015, 48:4 Page 3 of 11
http://www.biolres.com/content/48/1/4

Determination of antioxidant activity of honey by 2,
2-diphenyl-1-picrylhydrazyl (DPPH) assay
The DPPH radical scavenging analysis was used to inves-
tigate the overall hydrogen or electron donating activity
of single antioxidants. The scavenging abilities of the
different varieties of honey are presented in Table 4. The
results demonstrated that the greatest antioxidant activ-
ity of 86.20 mg gallic/kg of honey was found in the
honey collected from the nectar of coffee flower (source
No.4), while the honey obtained from the lychee flower
(source No.3) demonstrated the lowest DPPH radical
scavenging activity (5.93 mg gallic/kg of honey).
Determination of antioxidant activity of honey by ferric
reducing antioxidant power assay (FRAP)
The FRAP assay was used to estimate the amount of
antioxidants or reductants in a sample. The assay is
based on the ability of the sample to reduce the Fe
3+
to
the Fe
2+
couple, which was applied to determine the dif-
ferences in the antioxidant profiles of the various types
of honey. The results of the FRAP assay and the values
are presented in Table 4. The honey from coffee flower
(source No.4) had, significantly, the highest value of anti-
oxidant ability, with 1781.78 mg FeSO
4
/kg of honey.
Determination of anti-tyrosinase activity of honey
The tyrosinase inhibitory activity of honeys is shown in
Table 5. The inhibitory effects on tyrosinase activity ranged
from 22.71 ± 6.82 - 87.73 ± 5.33%. The highest activity
of tyrosinase inhibition was found from mãnuka honey.
The tyrosinase inhibition activity of mãnuka honey was
expressed as 500.15 mg koji/kg honey (p < 0.05). However,
honey from coffee flower (source No. 4) had highest anti-
tyrosinase activity, when compared to the other types of
tested honey (p < 0.05).
Discussion
In this study, Thai honey from different types of floral and
sources were collected. The physical characteristics of
honey were observed by determination of pH and viscosity
in this study. All types of honey showed pH ranging from
3.37 ± 0.08 to 4.06 ± 0.03 which were similar to the previous
reports that various types of honey also showed acidic pH.
Moreover, honey showed the viscosity that ranged from
1,083.50 ± 12.02 to 4,892.00 ± 63.64 Centipoise. The viscos-
ity of honey was influenced by temperature, moisture con-
tent and the presence of crystals and colloids. Honey from
different sources of flora showed the different physical char-
acteristics such as liquid honey or creamed honey because
the honey contained wide ranges of compounds [19-21].
Table 3 Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values of honey
against pathogenic bacteria causing skin disease
Source Type of
pollen
MIC and MBC (% honey)
Bacterial strains
S. aureus MRSA 49 MRSA 50 Corynebacterium sp. P.acnes
MIC MBC MIC MBC MIC MBC MIC MBC MIC MBC
1 Longan 25 25 12.5 25 12.5 25 25 25 25 50
Lychee 25 25 25 50 25 25 25 50 25 50
Polyflora 25 25 25 25 25 25 12.5 25 25 50
2 Longan 25 50 25 25 25 25 25 50 25 50
Sunflower 25 50 25 50 25 25 25 50 25 50
Polyflora 25 >50 25 25 25 50 25 50 25 50
3 Longan 25 50 25 25 25 50 25 50 25 50
Lychee 25 50 25 25 25 50 25 50 25 50
Polyflora 25 50 25 25 25 25 25 50 25 50
Forest flower 25 >50 25 >50 25 >50 25 >50 25 >50
4 Coffee 25 25 25 25 25 50 12.5 50 25 50
5 Polyflora 25 50 25 25 25 25 12.5 50 50 50
6 Lychee 25 >50 25 >50 25 50 25 50 25 50
Sunflower 25 >50 25 >50 25 >50 25 >50 50 >50
Polyflora 25 50 25 50 25 50 25 50 25 50
Sesame 25 50 25 >50 25 50 25 50 50 50
Mānuka honey 6.25 6.25 6.25 6.25 6.25 6.25 3.125 6.25 25 25
Jantakee and Tragoolpua Biological Research 2015, 48:4 Page 4 of 11
http://www.biolres.com/content/48/1/4

Table 4 The phenolic and flavonoid contents and antioxidant activities of honey by DPPH and FRAP assays
Source Type of pollen Phenolic content
(mg GAE/kg of honey)
Flavonoid content
(mg quercetin /kg of honey)
DPPH (mg GAE/kg
of honey)
FRAP (mg FeSO
4
/kg
of honey)
1 Longan 301.93 ± 47.90 40.46 ± 11.49 14.04 ± 0.18 627.57 ± 91.61
Lychee 253.69 ± 50.34 59.77 ± 20.93 7.35 ± 0.10 440.16 ± 57.85
Polyflora 320.37 ± 30.78 66.56 ± 15.64 12.28 ± 0.28 295.60 ± 50.86
2 Longan 467.66 ± 16.19 81.91 ± 3.09 33.74 ± 2.90 899.37 ± 119.27
Sunflower 380.94 ± 14.37 67.23 ± 4.70 17.93 ± 0.44 860.40 ± 124.53
Polyflora 425.19 ± 58.77 51.62 ± 1.41 15.06 ± 0.29 568.12 ± 95.83
3 Longan 312.53 ± 32.54 53.88 ± 2.62 11.36 ± 0.25 696.83 ± 94.80
Lychee 280.44 ± 64.79 34.48 ± 1.17 5.93 ± 0.02 319.56 ± 28.41
Polyflora 350.05 ± 53.06 55.42 ± 2.23 17.03 ± 2.54 645.61 ± 87.77
Forest flower 315.72 ± 12.37 35.40 ± 1.46 7.10 ± 0.12 373.76 ± 52.45
4 Coffee 734.76 ± 155.50 178.31 ± 40.04 86.20 ± 0.64 1781.78 ± 218.70
5 Polyflora 674.81 ± 52.77 79.75 ± 1.70 48.18 ± 9.20 1016.11 ± 87.91
6 Lychee 311.87 ± 12.11 55.99 ± 2.17 8.71 ± 1.86 269.83 ± 9.20
Sunflower 274.89 ± 69.37 47.36 ± 2.84 13.39 ± 2.86 490.61 ± 44.47
Polyflora 234.66 ± 22.47 29.86 ± 6.69 6.15 ± 0.65 331.87 ± 17.50
Sesame 314.92 ± 16.82 54.97 ± 0.19 9.12 ± 1.05 279.76 ± 19.65
Mānuka honey 831.88 ± 120.26 162.87 ± 11.51 83.80 ± 11.29 954.10 ± 123.77
Note: The data are given as mean ± standard deviation (SD) of triplicate experiments. The statistical comparison between values from the different types of honey
applied using the post hoc Duncan test. The values were found to be significantly different (p < 0.05).
Figure 1 The time–kill kinetic of the 50% honey from longan flowers (source No. 1) on MRSA49 (A) and S. aureus (B).
Jantakee and Tragoolpua Biological Research 2015, 48:4 Page 5 of 11
http://www.biolres.com/content/48/1/4

In this research, we found that all honey had antibacter-
ial activity against S. aureus, MRSA, and Corynebacterium
sp. and P. acnes by the agar well diffusion assay. However,
all the varieties of honey were observed to exhibit the
greatest inhibition on the growth of MRSA. The high vis-
cosity of honey helped to provide the barrier and protect
bacterial infection in the host [22]. Moreover, the high
sugar content of the honey could affect the osmolarity and
inhibited microbial growth [4].
Additionally, the antibacterial activity of mānuka honey
(Leptospermum scoparium)againstS. aureus, MRSA, and
Pseudomonas sp. were demonstrated. The antibacterial
activity of honey depended on various factors that function
either singularly or synergistically [15,23,24]. The honey
consists of hydrogen peroxide, phenolic compounds, lower
pH, osmotic pressure, and other phytochemical content.
Honey has the ability to generate hydrogen peroxide-
related antimicrobial activity. The production of hydrogen
peroxide by transforming glucose substrate with the
glucose oxidase of honey depends on the enzyme level
and the floral sources of honey [19].
The result also revealed that the antibacterial activity
was associated with the region where the honey was pro-
duced, as well. Different varities of honey from different
countries and regions vary widely and significantly in their
antibacterial activity and in their action against pathogenic
bacteria. Most of gram positive bacteria; S. aureus, MRSA,
Corynebacterium sp., and P. acnes were inhibited by the
honey from different types of pollen. Therefore, the action
of honey on tested organism may be due to the difference
in the species of bacteria. In this study, the mode of action
of Thai honey against the bacteria depended on different
factors such as the osmolarity, acidity and hydrogen
peroxide generation. The formation of free free OH
-
radi-
cals can break DNA and oxidize thiol-groups of proteins
and lipids leaded to damage of bacterial cells. Moreover,
after treatment of Thai honey that used in this study with
catalase enzyme resulted in the removal of the antibacter-
ial activity whereas the antibacterial activity of mānuka
honey still retained.
In addition, the broth dilution method was used to
determine the MIC and MBC values because this
method had more efficiency to indicate quantitative
results compared to the agar well on diffusion method
[5,24]. Minimum inhibitory concentration (MIC) and
minimum bactericidal concentration (MBC) of honey
determine the antibacterial activity of honey. The MIC
values of all the varieties of honey (Table 3) were found
to be in the range of 12.5–50% and the MBC values
were found to be in the range of 25% to >50% (Table 3).
Mānuka honey showed higher antibacterial activity,
with MIC ranging from 3.125% to 25%. In this study,
the broth dilution method, bacteria were brought into
direct contact with honey, while in the agar well diffusion
method, the components relied on diffusion through
the agar. Moreover, the diluted honey in the broth dilu-
tion method could generate hydrogen peroxide since
the enzyme glucose oxidase oxidized glucose to gluconic
acid and hydrogen peroxide. Therefore, antibacterial activ-
ity of the different varieties of honey was from hydrogen
peroxide activity [25].
However, other studies showed that the inhibitory
activity of honey on pathogenic bacteria was observed
when the treatment of the bacteria with honey was con-
ducted at concentrations lower than 3% or at concentration
50% or higher [24,26]. Furthermore, mānuka honey, as pre-
vious reports have pointed out, demonstrates high anti-
microbial activity and is known to contain non-peroxide
components such as polyphenols and protein defensin-1
that exhibit antimicrobial activity [27]. Additionally, methyl-
glyoxal was also found as a major compound in mānuka
honey and could inhibit pathogenic bacteria such as E. coli
and S. aureus [11]. Besides, honey from the flowers of E.
cladocalyx (Blue gum), Erica species (Fynbos), and L. cordi-
folium (Pincushion) were seen to demonstrate antibacterial
activity similar to mānuka honey [28].
The investigation of the killing kinetic of honey obtained
from the longan flower at the concentration of 50% (v/v)
demonstrated the reduction in the survival of S. aureus
and MRSA49, although S. aureus exhibited more
Table 5 Anti-tyrosinase activity of honey from different
types of pollen
Source Type of pollen Tyrosinase
inhibition (%)
Kojic acid equivalents
(mg koji/kg honey)
1 Longan 60.09 ± 5.51 34.23
Lychee 56.51 ± 1.44 24.19
Polyflora 58.20 ± 4.76 28.50
2 Longan 53.27 ± 10.38 17.67
Sunflower 46.64 ± 3.85 9.29
Polyflora 45.01 ± 2.78 7.93
3 Longan 45.02 ± 8.01 7.94
Lychee 48.30 ± 9.65 10.91
Polyflora 40.17 ± 4.44 4.96
Forest flower 22.71 ± 6.82 0.91
4 Coffee 63.46 ± 16.30 47.49
5 Polyflora 44.57 ± 4.70 7.60
6 Lychee 51.87 ± 5.94 15.42
Sunflower 52.12 ± 4.05 15.81
Polyflora 49.61 ± 11.26 12.40
Sesame 52.53 ± 5.19 16.45
Mānuka honey 87.73 ± 5.33 500.15
Note: The data are given as mean ± standard deviation (SD) of triplicate
experiments. The statistical comparison between values from different types of
honey applied using the post hoc Duncan test. The values were found to be
significantly different (p < 0.05).
Jantakee and Tragoolpua Biological Research 2015, 48:4 Page 6 of 11
http://www.biolres.com/content/48/1/4

resistance to honey. Interestingly, methicillin resistant S.
aureus (MRSA) was found to have been completely inhib-
ited by the honey at 8 hours of treatment. Therefore, the
significant finding of the study was the potent activity of
Thai honey against antibiotic-resistant pathogenic bac-
teria, MRSA. The anti-bacterial activity of the honey is a
property that is of valuable interest in its function as an
agent in treating bacterial infections that are resistant to
the action of antibiotics.
Another study conducted using Revamil
®
medical-grade
honey (RS honey) produced in the Netherlands at a con-
centration of 40% (v/v) also demonstrated anti-bacterial
activity against resistant strains of bacteria such as MRSA,
methicillin-resistant S. epidermidis (MRSE), vancomycin-
resistant Enterococcus faecium (VREF), extended-spectrum
beta-lactamase (ESBL) producing P. aeruginosa,and
Burkholderia cepacia. The growth inhibition of MRSA
and ESBL E. coli was found to have taken place at
6 hours of incubation with the Revamil
®
honey. The RS
honey demonstrated high sugar concentration, H
2
O
2
production, MGO, low pH, and bee defensin-1, and
these were significant factors contributing toward the
bactericidal activity [12,29].
Natural products include several phytochemical contents
such as phenolic acid and flavonoids. These have been
reported to have antioxidant and antibacterial activities.
Moreover, the polyphenol compounds are essential for
improving the potential effects on human health. Phenolic
compounds are some of the most important groups that
can be found in plants and honey. In addition, the com-
position of phenolic and flavonoid compounds of honey
depends on floral sources, seasonal factors, and envir-
onmental factors [30]. As far as the quantitation of the
total phenolic compounds in Thai honey is concerned,
the phenolic compound was investigated using the
Folin–Ciocalteu assay, and gallic acid was used as the
standard. The assay measured the phenol and polyphenol
derivatives, and other electron-donating antioxidants [31].
The results revealed that the total phenolic compounds
existed in the wide range of 234.66 mg gallic acid/kg
honey to 831.88 mg gallic acid/kg honey. These differ-
ences in the antioxidant activities of honey depend on the
floral sources and the sources of collection. Furthermore,
the total phenolic content of five different types of Yemeni
honey demonstrated different values of antioxidant activ-
ity and phenolic compound, which can be attributed to
the fact that the differences in the botanical sources of the
honey and the colors of the honey were mainly due to the
differences in the content and composition of the phenolic
compounds in them [16,32]. Monofloral Cuban honey
showed phenolic content values ranging from 213.9 GAE/
kg of honey to 595.8 GAE/kg of honey [33].
In this study, it was found that the honey from coffee
flower had high phenolic content (734.76 mg gallic acid/
kg honey) when compared to honey from other sources.
The phenolic content of the honey from coffee flower
was found to be as high as that of mānuka honey
(831.88 mg gallic acid/kg honey). Thus, it can be con-
cluded that different varieties of honey contain several
different polyphenols, at different concentrations, which,
in turn, affects the efficacy of the honey [34].
Flavonoids are widely found in food products derived
from plant sources and honey, and they are a type of
phenolic compounds [35]. The potential of flavonoids is
considerable in its ability to protect the pBR322 plasmid
DNA, against reactive oxygen species (ROS) or against
H
2
O
2
-induced oxidative damage [36]. The investigation
conducted in this study regarding the total flavonoid
content of Thai honey showed that the flavonoid content
depended on the floral sources as well as on the sources
of collection. The honey collected from coffee flower
had the highest flavonoid content, which was not signifi-
cantly different from that of mānuka honey. Therefore,
in this study, the observation regarding total flavonoid
content correlated with that of total phenolic content.
These results are similar to the findings obtained in the
study conducted by Alvarez-Suarez et al. (2010) [33].
In addition, the correlation between total flavonoid
content and total phenolic content depends on the color
of fresh honey, for example, darker-colored varieties of
honey (buckwheat and heather) showed total phenolic
contents higher (71.7 μg/g to 202.6 μg/g) than those of
lighter-colored honey (rape honey). However, after long
periods of storage the color of the honey was observed
to become too dark from hydroxymethylfurfural (HMF)
[32]. Thus, antioxidant activity is also influenced by the
temperature of processing and the storage method of
the material. Documents on the varieties of African
honey and the varieties of European honey certify them
as containing flavonoids such as quercetin, hesperetin,
kaempferol, apigenin, isorhamnetin, and myricetin [37].
The antioxidant activities of honey from different types
of pollen were evaluated using DPPH and FRAP assays.
Boththeassaysshowedthathoneyfromcoffeeflower
had the highest antioxidant capacity compared to other
types of honey. Similar antioxidant properties were also
reported from Turkish red pine honey produced by
Marchalina hellenica, Saudi Arabian honey, Peruvian
honey, Malaysian tualang honey, American buckwheat
honey, Spanish honey, Portuguese honey, Cuban honey,
Venezuelan honey, and Ecuadorian honey [38-42]. Add-
itionally, the antioxidant properties of the types of
honey produced by stingless bees, for example, Trigona
carbonaria in Australia and Trigona laeviceps in Thailand,
were also reported [43,44]. The antioxidant activity is due
to the presence of various substances such as enzymes,
organic acids, amino acids, Maillard reaction products,
phenolic compounds, flavonoids, tocopherols, catechins,
Jantakee and Tragoolpua Biological Research 2015, 48:4 Page 7 of 11
http://www.biolres.com/content/48/1/4

ascorbic acid, and carotenoids [45]. The high antioxidant
capacity of honey correlated with the presence of phenolic
and flavonoid compounds. Schneider et al. (2013) [19]
also found a strong correlation between antioxidant cap-
acity and polyphenol content since polyphenols are the
major contributors to the antioxidant effect of honey.
Anti-tyrosinase activity of honey was tested and it
showed that the percentage of inhibition ranged from
22.71 ± 6.82 - 87.73 ± 5.33%. Other natural substances
such as ginger species and longan fruit extracts also
showed anti-tyrosinase activity. Previous study demon-
strated that favonols, galangin, kaempferol and quercetin
were found to inhibit tyrosinase enzyme [46-48]. Thus,
honey demonstrated efficacy to inhibit tyrosinase enzyme.
From the result in this study, the honey from coffee
pollen showed high degree of phenolic, flavonoid, anti-
oxidant and anti-tyrosinase activity, which was similar to
mānuka honey. Therefore, the antioxidant and anti-
tyrosinase activity of Thai and mānuka honey were
correlated. Although, in this study the presence of phen-
olic and flavonoid compounds were varied depending on
the types of honey but similar anti-bacterial activity was
also observed on different types of honey.
Moreover, treatment of all Thai honey used in this
study including honey from pollens of lychee, longan,
sesame, polyflora, sunflower, forest flower, and coffee with
catalase enzyme showed the removal of the antibacterial
activity in this study. Therefore, antibacterial activity of
Thai honey was due to hydrogen peroxide activity of the
honey. The highest antioxidant activity of honey from
coffee pollen was from phenolic and flavonoid compounds
sincetheywerethemostimportantgroupsoftheanti-
oxidant substances in honey [16]. The antibacterial activity
was not correlated with phenolic and flavonoid com-
pounds that found in honey.
However, honey from coffee pollen did not show high
anti-bacterial activity as the mānuka honey. The antibacter-
ial compound of Thai honey was different from mānuka
honey since the antibacterial activity of mānuka honey
was mainly from non-peroxide component such as
methylglyoxal [10].
The removal of hydrogen peroxide activity from Ulmo
honey from Chile was also shown to reduce its anti-
microbial activity when tested in the presence of catalase
[49]. Therefore, Thai honey and mānuka honey showed
different degree of anti-bacterial activity against patho-
genic bacteria causing skin diseases.
Conclusion
This study demonstrated the similar antibacterial activity
of Thai honey obtained from various types of flowers
against pathogenic bacteria causing skin diseases, includ-
ing S. aureus,MRSA,Corynebacterium sp., and P. acnes.
Antibacterial activity of Thai honey was due to peroxide
activity, whereas the activity of mānuka honey was from
non-peroxide activity of the honey. In addition, it was
demonstrated that the honey collected from coffee flower
showed the highest antioxidant activity and anti-tyrosinase
activity, a finding that correlates with the phytochemical
composition of honey which consists of phenolic and flavo-
niod compounds. Therefore, as is evident, in this study, it
was also established that the potential of honey for use as a
powerful source of antibacterial, antioxidant and anti-
tyrosinase activity is tremendous. Moreover, honey can be
used as an alternative therapeutic agent against pathogenic
bacteria causing skin diseases.
Methods
Honey collection and preparation
The honey was collected from different sources and types
of pollens, including lychee, longan, sesame, polyflora,
sunflower, forest flower, and coffee during February - April
2012. Thai honey samples were purchased from local
market and bee farm in July 2012. Also, mānuka honey
(comvita®), produced in New Zealand, was used as the
positive control in this study. Honey sample was kept
in the dark at room temperature. The control solution
contained 39% (w/v) d-fructose, 31% (w/v) d-glucose,
8% (w/v) maltose, 3% (w/v) sucrose, and 19% (w/v)
water, and it was kept in refrigerator [28].
Bacterial strains
Pathogenic bacteria causing infectious diseases on skin
were tested. Gram positive bacteria, such as Staphylococcus
aureus, methicillin resistant S. aureus (MRSA), Staphylo-
coccus epidermidis,Corynebacterium sp., Bacillus sp.,
Micrococcus luteus, and Propionibacterium acnes,and
Gram negative bacteria, such as Pseudomonas aeruginosa,
were obtained from the Microbiology Section, Department
of Medical Technology, Faculty of Associated Medical
Sciences, Chiang Mai University, Chiang Mai, Thailand,
while P. acnes DMST14916 were obtained from the
culture collection section of the Department of Medical
Science, Ministry of Public Health, Thailand. All the
bacteria were cultured in a liquid medium for 24–72
hours before testing with honey.
Viscosity and pH of honey
Viscosity was measured using a rotational viscometer
(Brookfield viscometer DV-III+ Rheometer, USA) with
SC4-29 spindle. The pH of honey was also measured by
pH meter (Denver instrument, USA).
Determination of activity of honey on bacteria using the
agar well diffusion assay
Screening of the antibacterial activity of honey was per-
formed using the agar well diffusion assay. The cultured
bacteria were adjusted to McFarland No. 0.5 (1 × 10
8
CFU/
Jantakee and Tragoolpua Biological Research 2015, 48:4 Page 8 of 11
http://www.biolres.com/content/48/1/4

ml) with 0.85% NaCl solution. Then, the Mueller Hinton
agar (MHA) plate was swabbed with the suspension. After
that, 60 μl of the 100% honey sample and the control sugar
solution were added to each well, by comparing it to
phenol, which was used as the positive control. Gentamycin
was also used as a drug control. The peroxide activity of
honey on bacteria was also determined by the agar well
diffusion assay after treatment of the 100% honey (100 μl)
with 5 μl of catalase enzyme, 2 mg/ml (Sigma, 4000 U/mg)
[50]. The plates were incubated at 37°C for 18–24 hours.
After incubation, the inhibition zones were determined and
the diameters of the zones were recorded.
Determination of minimum inhibitory concentration (MIC)
and minimum bactericidal concentration (MBC) of honey
on bacteria
The minimum inhibitory concentration (MIC) was deter-
mined using the broth dilution method. The different sam-
ples of honey were diluted two-fold, from 100% to 50%,
25%, 12.5%, 6.25%, and 3.12%, with sterile Mueller Hinton
broth (MHB). After that, the bacterial culture was adjusted
to McFarland No. 0.5 for approximately 1 ×10
8
CFU/ml.
Thereafter, the bacterial culture was added to each dilution
of honey and incubated at 37°C for 18–24 hours. After the
incubation, the turbidity of the solution was observed and
recorded as the lowest concentration of honey to inhibit
the bacterial growth. The culture that completely inhibited
visible growth on account of honey was streaked on an
MHA plate for evaluating the minimum bactericidal con-
centration (MBC) after incubation at 37°C 18–24 hours.
The MBC was determined as the minimum bactericidal
concentration.
Determination of time–kill endpointsof honey on bacterial
growth
The time–kill studies were performed for a period of
24 hours, and stock honey was prepared to 2 MIC.
Then, the bacteria were adjusted to McFarland No. 0.5
and mixed with honey in the ratio of 1:1 and incubated
at 37°C, while inoculums without honey were used as
the control. The samples were taken from the experi-
ment at 0 minutes, 10 minutes, 20 minutes, 30 minutes,
90 minutes, and 120 minutes, and every 2 hours until
24 hours. At the end of each time period, the sample
was ten-fold serial diluted with 0.85% NaCl, and 100 μl
of each dilution was spread onto the MHA plates. After
incubation at 37°C for 24 hours, the bacterial colonies
were counted and recorded.
Determination of total phenolic content of honey
The Folin–Ciocalteu method was used to determine the
total phenolic content [35]. Honey (250 μl) at a concentra-
tion of 100 mg/ml was mixed with 1.25 ml of water,
250 μl of 95% ethanol, and, then, 125 μl of 50% Folin–
Ciocalteu. The solution was incubated at room temp-
erature for 5 minutes. After that, 5% of Na
2
CO
3
was
added to the solution and incubated for 1 h, and the
reaction was measured to determine the absorbance at
the wavelength 725 nm. The total phenolic content was
expressed in mg of gallic acid equivalents (mg GAE/kg
of honey).
Determination of flavonoid content of honey
The total flavonoid content was determined using the
colorimetric method discussed in Ghasemi et al. (2009)
[35]. The honey sample was diluted with methanol to a
final concentration of 100 mg/ml. Five hundred microliters
of the honey solution was mixed with 0.1 ml of 10% alu-
minium chloride, 1.5 ml methanol, 0.1 ml 1 M potassium
acetate, and 2.8 ml distilled water. Thereafter, the solution
was incubated at room temperature for 30 minutes.
After the incubation, the absorbance of the reaction was
measured at the wavelength 415 nm. The total flavon-
oid content was expressed as mg quercetin equivalents
(mg quercetin/kg of honey).
Determination of antioxidant activity of honey by 2,
2-diphenyl-1-picrylhydrazyl (DPPH) assay
The antioxidant activity of the different types of honey
was evaluated using the DPPH radical scavenging assay
[35]. The honey was prepared to various concentrations
by dissolving in methanol. A volume of 0.5 ml of the
honey was mixed with 1.5 ml of the DPPH reagent and
incubated in the dark at room temperature for 20 mi-
nutes. The absorbance of the reaction was measured at
the wavelength 517 nm. The DPPH without honey was
used as the control and methanol was used as the blank
solution. The scavenging activity was evaluated. The
IC
50
(mg/ml) value was calculated using the following
equation:
% Inhibition ¼Acontrol −Atest
ðÞ=Acontrol
½100
Finally, the antioxidant activity of honey was reported as
the gallic acid equivalent antioxidant capacity (mg GAE/
kg of honey).
Determination of antioxidant activity of honey by ferric
reducing antioxidant power assay (FRAP)
The FRAP assay was performed according to the method
discussed in Benzie and Strain (1996) [51]. The different
samples of honey were dissolved with deionized water to
100 mg/ml. The FRAP reagent included 30 mM acetate
buffer pH 3.6, 10 mM TPTZ solution, 20 mM FeCl
3
solu-
tion, and deionized water. A volume of 0.5 ml of the honey
solution was mixed with 1.5 ml of the FRAP reagent.
Thereafter, the mixture was incubated in the dark at
room temperature for 15 minutes. The absorbance of
Jantakee and Tragoolpua Biological Research 2015, 48:4 Page 9 of 11
http://www.biolres.com/content/48/1/4

the reaction was measured at the wavelength of 593 nm.
The FRAP without honey was used as the control and
deionized water was used as the blank solution. The anti-
oxidant activity of honey was calculated from the ferrous
sulfate standard curve, which has the antioxidant stand-
ard, and reported as mg ferrous sulfate/kg of honey.
Determination of anti-tyrosinase activity of honey
The method was evaluated using the dopachrome
micro-plate [52]. Honey was dissolved in 20% ethanol to
a final concentration of 50%. This stock solution, 50 μl
was combined with 150 μl of 0.02 M phosphate buffer
(pH 6.8) and 50 μl of mushroom tyrosinase (313 Units/
ml in phosphate buffer, Sigma Chemical), and incubated
for 10 minutes. After that, 0.32 mM 3,4-Dihydroxy-L-
phehylalanine (Sigma Chemical), 50 μl was used as sub-
strate and added to the each well. Anti-tyrosinaseactivity
was evaluated by measuring absorbance at 492 nm before
incubation at 25°C and after incubation for 2 min. Kojic
acid (Merck Millipore) was used as the standard inhibitor
of tyrosinase enzyme. The % inhibition was calculated
using the following equation:
% Inhibition ¼A−BðÞ–C−DðÞ½=A−Bfg100
Where A was the optical density (OD
492
) of the control
(L-Dopa mixed with tyrosinase enzyme in buffer); B repre-
sented the blank (L-Dopa in buffer); C represented the
reaction ofL-Dopa with tyrosinase enzyme and honey in
buffer and D represented the blank of C (L-Dopa mixed
with honey in buffer)
Statistical analysis
The results were expressed as mean ± standard deviation.
The ANOVA test was used to analyze the variance of data
(SPSS software version 17.0 for Windows). The analysis
average of the treatment using multiple comparisons was
determined by using Dancan’s multiple range tests, and
the data were compared using the p values: p < 0.05 was
considered statistically significant. The least significant
difference (LSD) was used to determine the difference
between the methods used to the investigation of the
various antioxidant capacities.
Competing interests
The authors declare that they have no competing interests.
Authors’contributions
YT carried out conception and design of the study, acquisition of data,
analysisand interpretation of data, drafting the manuscript and revising. KJ carried
out the experiment, analysis, and interpretation of data, statistical analysis, and
drafting the manuscript. All authors read and approved the final manuscript.
Acknowledgements
We would like to thank the Faculty of Science, Chiang Mai University,
Thailand. The Thailand Research Fund-Research and Researchers for Industry
(RRi) Master Scholarship (MSD56I0061) and Bee Products Industry Co., Ltd.,
are also acknowledged for their financial support.
Received: 16 August 2014 Accepted: 7 January 2015
Published: 16 January 2015
References
1. Halco´n L, Milkus K. Staphylococcus aureus and wounds: a review of tea tree
oil as a promising antimicrobial. Am J Infect Control. 2004;32:402–8.
2. Molan PC. Why honey is effective as a medicine. 1 its use in modern
medicine. Bee World. 1999;80(2):80–92.
3. Mandal MD, Mandal S. Honey: its medicinal property and antibacterial
activity. Asian Pac J Trop Biomed. 2011;1(2):154–60.
4. Cooper RA, Molan PC, Harding KG. The sensitivity to honey of gram positive
cocci of clinical significance isolated from wounds. J Appl Microbiol.
2002;93:857–63.
5. Erejuwa OO, Gurtu S, Sulaiman SA, AB Wahab MS, Sirajudeen KN, Salleh MS.
Hypoglycemic and antioxidant effects of honey supplementation in
streptozotocin-induced diabetic rats. Int J Vitam Nutr Res. 2010;80:74–82.
6. Tan HT, Rahman RA, Gan SH, Halim AS, Hassan SA, Sulaiman SA, et al. The
antibacterial properties of Malaysian tualang honey against wound and
enteric microorganisms in comparison to manuka. BMC Complement Altern
Med. 2009;9:34.
7. Koc AN, Silici S, Kasap F, Hormet-oz HT, Mavus-buldu H, Ercal BD. Antifungal
activity of the honeybee products against Candida spp. and Trichosporon
spp. J Med Food. 2011;14:128–34.
8. Kassim M, Achoui M, Mustafa MR, Mohd MA, Yusoff KM. Ellagic acid,
phenolic acids and flavonoids in Malaysian honey extracts demonstrate
in vitro anti- inflammatory activity. Nutr Res. 2010;30:650–9.
9. Visavadia BG, Honeysett J, Danford MH. Manuka honey dressing: An
effective treatment for chronic wound infections. Br J Oral Maxillofac Surg.
2006;44:38–41.
10. Mavric E, Wittmann S, Barth G, Henle T. Identification and quantification of
methylglyoxal as the dominant antibacterial constituent of manuka
(Leptospermum scoparium) honeys from New Zealand. Mol Nutr Food Res.
2008;52:483–9.
11. Kwakman PHS, De Boer L, Ruyter-spira CP, Creemers-Molenaar T, Helsper
PFG, Vandenbroucke-Grauls CMJE, et al. Helsper PFG, vandenbroucke-Grauls
CMJE, Zaat SAJ, Te Velde AA: Medical-grade Honey enriched with antimicrobial
peptides has enhanced activity against antibiotic resistant. Eur J Clin Microbiol.
2011;30:251–7.
12. Molan PC. The evidence supporting the use of honey as a wound dressing.
Int J Low Extrem Wounds. 2006;5(1):40–54.
13. Green AE. Wound healing properties of honey. Brit J Surg. 1988;75(12):1278.
14. Meda A, Lamien CE, Romito M, Millogo J, Nacoulma OG. Determination of
the total phenolic, flavonoid and proline contents in Burkina Fasan honey,
as well as their radical scavenging activity. Food Chem. 2005;91:571–7.
15. Molan PC, Betts JA. Clinical usage of honey as a wound dressing: an update.
J Wound Care. 2004;1:353–6.
16. Al-mamary M, Al-meeri A, Al-habori M. Antioxidant activities and total
phenolic of different types of honey. Nutr Res. 2002;22:1041–7.
17. Chang TS. An updated review of tyrosinase inhibitors. Int J Mol Sci.
2009;10:2440–75.
18. Kim YJ, Uyama H. Tyrosinase inhibitors from natural and synthetic sources:
structure, inhibition mechanism and perspective for the future. Cell Mol Life
Sci. 2005;62:1707–23.
19. Schneider M, Coyle S, Warnock M, Gow I, Fyfe L. Anti-microbial activity and
composition of Manuka and Portobello honey. Phytother Res. 2013;27:1162–8.
20. Yanniotis S, Skaltsi S, Karaburnioti S. Effect of moisture content on the
viscosity of honey at different temperatures. J Food Eng. 2006;72:372–7.
21. Ren Z, Bian X, Lin L, Bai Y, Wang W. Viscosity and melt fragility in
honey- water mixtures. J Food Eng. 2010;100:705–10.
22. Lusby PE, Coombes AL, Wilkinson JM. Bactericidal activity of different
honeys against pathogenic bacteria. Arch Med Res. 2005;36:464–7.
23. Moussa A, Noureddine D, Abdelmelek M, Saad A. Antibacterial activity of various
honey types of Algeria against Pathogenic Gram-negative Bacilli,Escherichia coli
and Pseudomonas aeruginosa. Asian Pac J Trop Dis. 2012;2(3):211–4.
24. Mulu A, Tessema B, Derbie F. In vitro assessment of the antimicrobial potential of
honey on common human pathogen. Ethiop J Health Dev. 2004;18(2):107–12.
25. Bang LM, Buntting C, Molan P. The effect of dilution on the rate of
hydrogen peroxide production in honey and its implications for wound
healing. J Altern Complement Med. 2003;9(2):267–73.
26. Wilkinson JM, Cavanagh HMA. Activity of 13 honeys against Escherichia coli
and Pseudomonas aeruginosa. J Med Food. 2005;8:100–3.
Jantakee and Tragoolpua Biological Research 2015, 48:4 Page 10 of 11
http://www.biolres.com/content/48/1/4

27. Weston RJ, Mitchell KR, Allen KL. Antibacterial phenolic components of New
Zealand manuka honey. Food Chem. 1999;64:295–301.
28. Basson NJ, Grobler SR. Antimicrobial activity of two South African honeys
produced from indigenous Leucospermum cordifolium and Erica species on
selected microorganisms. BMC Complement Altern Med. 2008;8:1–4.
29. Kwakman PHS, Te Velde AA, De Boer L, Vandenbroucke-Grauls CMJE, Zaat
SAJ. Two major medicinal honeys have different mechanisms of bactericidal
activity. PLoS One. 2011;6(3):1–7.
30. Estevinho L, Pereira AP, Moreira L, Dias LG, Pereira E. Antioxidant and
antimicrobial effects of phenolic compounds extracts of Northeast Portugal
honey. Food Chem Toxicol. 2008;46:3774–9.
31. Denisov ET, Afanas'ev IB. Oxidation and Antioxidants in Organic Chemistry
and Biology. New York: Taylor and Francis; 2005.
32. Kaskoniene V, Maruska A, Kornysova O, Charczun N, Ligor M, Buszewski B.
Quantitative and qualitative determination of phenolic compounds in
honey. Chemine Technologija. 2009;3(52):1–7.
33. Alvarez-suarez JM, Tulipani S, Díaz D, Estevez Y, Romandini S, Giampieri F,
et al. Antioxidant and antimicrobial capacity of several monofloral Cuban
honeys and their correlation with color, polyphenol content and other
chemical compounds. Food Chem Toxicol. 2010;48:2490–9.
34. Beretta G, Granta P, Ferrero M, Orioli M, Facino RM. Standardization of
antioxidant properties of honey by a combination of spectrophotometric/
fluorimetric assaysand chemometrics. Anal Chim Acta. 2005;533:185–91.
35. Ghasemi K, Ghasemi Y, Ebrahimzadeh MA. Antioxidant activity, phenol and
flavonoid contents of 13 citrus species peels and tissues. Pak J Pharm Sci.
2009;22(3):277–81.
36. Kalpana KB, Srinivasan M, Menon VP. Antioxidant potential of aminothiazole
derivative and its protective effect on H
2
O
2
-induced oxidative damage on
pBR322 DNA and RBC cellular membrane. Mol Cell Biochem. 2008;314:95–103.
37. Serem JC, Bester MJ. Physicochemical properties, antioxidant activity and
cellular protective effects of honeys from southern Africa. Food Chem.
2012;133:1544–50.
38. Al-hindi RR, Bin-masalam MS, El-Shahawi MS. Antioxidant and antibacterial
characteristics of phenolic extracts of locally produced honey in Saud Arabia.
Int J Food Sci Nutr. 2011;62:513–7.
39. Khalil MI, Alam N, Moniruzzaman M, Sulaiman SA, Gan SH. Phenolic acid
composition and antioxidant properties of Malaysian honeys. J Food Sci.
2011;76:C921–8.
40. Kishore RK, Halim AS, Syazana MS, Sirajudeen KN. Tualang honey has higher
phenolic content and greater radical scavenging activity compared with
other honey sources. Nutr Res. 2011;31:322–5.
41. Jerkovic I, Marijanovic Z. Oak (Quercus frainetto Ten.) honeydew
honey- approach to screening of volatile organic composition and
antioxidant capacity (DPPH and FRAP assay). Molecules. 2010;15:3744–56.
42. Erejuwa OO, Sulaiman SA, Wahab MS. Honeya novel antioxidant. Molecules.
2012;17:4400–23.
43. Oddo LP, Heard TA, Rodriguez-Malaver A, Perez RA, Fernandez-Muino M,
Sancho MT, et al. Composition and antioxidant activity of Trigona carbonaria
honey from Australia. J Med Food. 2008;11:789–94.
44. Suntiparapop K, Prapaipong P, Chantawannakul P. Chemical and biological
properties of honey from Thai stingless bee (Tetragonula leaviceps). J Apicult
Res. 2012;51(1):45–51.
45. Socha R, Juszczak L, Pietrzyk S, Fortuna T. Antioxidant activity and phenolic
composition of herbhoneys. Food Chem. 2009;113:568–74.
46. Kubo I, Kinst-Hori I, Chaudhuri SK, Kubo Y, Sánchezb Y, Ogurab T. Flavonols
from heterotheca inuloides: tyrosinase inhibitory activity and structural criteria.
Bioorg Med Chem. 2000;8:1749–55.
47. Rangkadilok N, Sitthimonchai S, Worasuttayangkurn L, Mahidol C,
Ruchirawat M, Satayavivad J. Evaluation of free radical scavenging and
antityrosinase activities of standardized longan fruit extract. Food Chem
Toxicol. 2007;45:328–36.
48. Chan EWC, Lim YY, Wong LF, Lianto FS, Wong SK, Lim KK, et al. Antioxidant
and tyrosinase inhibition properties of leaves and rhizomes of ginger
species. Food Chem. 2008;109:477–83.
49. Sherlock O, Dolan A, Athman R, Power A, Gethin G, Cowman S, et al.
Comparison of the antimicrobial activity of Ulmo honey from Chile and
Manuka Honey against methicillin-resistant Staphylococcus aureus, Escherichia
coli and Pseudomonas aeruginosa. BMC Complement Altern Med. 2010;10:47.
doi:10.1186/1472-6882-10-47.
50. Allen KL, Molan PC, Reid GM. A survey of the antibacterial activity of some
New Zealand honeys. J Pharm Pharmacol. 1991;43:817–22.
51. Benzie IFF, Strain JJ. The Ferric reducing ability of plasma (FRAP) as a measure
of “antioxidant power”: The FRAP assay. Anal Biochem. 1996;239:70–6.
52. Potduang B, Meeploy M, Giwanon R, Benmart Y, Kaewduang M,
Supatanakul W. Biological activities of Asparagus racemosus. Afr J Tradit
Complement Altern Med. 2008;5(3):230–7.
doi:10.1186/0717-6287-48-4
Cite this article as: Jantakee and Tragoolpua: Activities of different types
of Thai honey on pathogenic bacteria causing skin diseases, tyrosinase
enzyme and generating free radicals. Biological Research 2015 48:4.
Submit your next manuscript to BioMed Central
and take full advantage of:
• Convenient online submission
• Thorough peer review
• No space constraints or color figure charges
• Immediate publication on acceptance
• Inclusion in PubMed, CAS, Scopus and Google Scholar
• Research which is freely available for redistribution
Submit your manuscript at
www.biomedcentral.com/submit
Jantakee and Tragoolpua Biological Research 2015, 48:4 Page 11 of 11
http://www.biolres.com/content/48/1/4