Iranian Honey Research Articles

Evaluation of the potential in vitro effects of propolis and honey on wound healing in human dermal fibroblast cells

Propolis and honey are two apiculture products with various pharmacological and biological properties that have been recognized since ancient times. The present investigation was an in vitro analysis of the impacts of individual and concomitant application of Iranian propolis and honey on the process of wound healing. The total phenolic contents of ethanolic extract of propolis (EEP) and honey were determined using Folin-Ciocalteu method. Cultured HDF cells were exposed to different concentrations of EEP, honey, and mixture of both substances, and then the percentage of wound closure and rate of cell migration were examined. Simultaneously, the viability of HDF cells exposed to the EEP and/or honey was assessed. In addition, the effects of both substances on the proliferation of the cultured HDF cells were investigated. The value of total phenolic contents of EEP was 111.388 ± 0.088 and for honey was 1.848±0.79 mg GAE/g. The optimum concentration of EEP and honey for wound healing, rate of cell migration, cell viability, and proliferation assays were observed at 100 µg/ml and 200 µg/ml. Similarly, the optimum concentrations of the mixture in all the tests were seen at 100 µg/ml and 200 µg/ml. The Iranian propolis and honey samples efficiently affected migration, proliferation, and viability of HDF cells in a dose-dependent manner, however, no synergistic effects of EEP and honey were observed.

Study of Human IgG and IgE Antibodies Against Bee (Apis mellifera) Venom

BACKGROUND: Bee venom contains various biomolecules, such as enzymes, peptides, and amines. The immune sys-tem produces IgG antibodies against bee venom proteins. However, IgE antibodies may also be developed in allergic individuals. OBJECTIVES: In this study, immune responses, including IgG and IgE reactions to bee venom were assessed in vari-ous individuals, using the immunoblotting technique. METHODS: Serum samples were collected from 20 people of three major groups, namely beekeepers, allergic individu-als, and normal people. Venom samples of honey bees and wild bees were collected from the suburbs of Tehran, Iran. Furthermore, commercial honey bee venom samples extracted from Apis mellifera and samples of wild bees extracted from Polistes and Vespula were purchased from France. Immunoblotting was carried out using the sera of subjects and anti-human IgG and IgE coupled to horseradish peroxidase. RESULTS: The results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed similar protein bands in Iranian and European honey bee venoms, including α-glucosidase (170 kDa), Api m (100 kDa), acid phosphatase (49 kDa), hyaluronidase (43 kDa), phospholipase A2 (17 kDa), and melittin (2 kDa). In wild bees, two bands were found with the molecular weights of 35 and 25 kDa belonging to antigen 5 and phospholipase A1, respectively. These were not observed in honey bee venoms. Immunoblot analysis revealed that all the mentioned proteins were immunogenic and al-lergenic in different individuals. Hyaluronidase, as well as phospholipases A1 and A2, were the major allergens in most individuals, while IgE reaction to melittin was only reported in one person. CONCLUSIONS: In conclusion, studies on antibodies against bee venoms can be useful in immunotherapy. Different people indicated distinct allergenic patterns. Therefore, further similar assays are recommended before, during, and after immunotherapy.

Protective effect of honey on learning and memory impairment, depression and neurodegeneration induced by chronic unpredictable mild stress

Introduction: Chronic stress, which has been prevalent in human life, induce structural changes in the hippocampus. Depression and impairment of memory are serious comorbidities of chronic stress. In this study, we evaluated the impact of an Iranian honey pretreatment on memory deficit, depression and the hippocampal neuronal loss in the chronic unpredictable mild stress (CUMS) model. Methods: Adult male Wistar rats were divided into the control groups that received water or honey (0.2 or 2g/kg) and CUMS groups that subjected different, randomly and unpredictable mild stressors for 4 weeks. Ten days before starting the CUMS procedures, the animals received honey (0.2 or 2g/kg, daily, orally), which was continued until sacrificing. Morris water maze and sucrose performance tests were used to evaluate the spatial learning and memory and depressive-like behavior in the animals respectively. Hippocampus and whole brain samples were collected for further biochemical and histological analysis. Results: Honey reversed the depression-like behavior and ameliorated the spatial memory deficit induced by CUMS. Also, honey decreased cell death in the hippocampus and reduced the malondialdehyde level in treated animals. Conclusion: These results revealed that honey diminished learning and memory deficits and depression in chronic stress conditions.

Correction to: Genetic characteristics of the Iranian honey bee, Apis mellifera meda, based on mitochondrial genes of ND4, ND4L and ND6 and their internal transcribed spacers

Honey bees are one of the most important social insects in the world. Genetic markers are powerful tools for differentiating honey bee subspecies. The purpose of the present research was the introduction of a new barcoding marker and assessment of the accuracy and discriminatory abilities of ND4, ND4L, ND6 genes and related intergenic spacers for the phylogenetic study of Iranian honey bees. Sampling was conducted throughout Iran (30 locations). ND4–ND4L–ND6 (with their ITS regions) demonstrated the most accurate analysis for the phylogenetic study of Iranian honey bees compared with ND4–ND4L (with ITS1) and ND4L–ND6 (with ITS2, ITS3, ITS4 and ITS5 regions). Additionally, the findings illustrated that all the collected honey bee samples were A. m. meda except those collected from the Kolachay area which were classified as A. m. carnica. Ten haplotypes were identified in the Iranian honey bee populations using the mitochondrial sequences of ND4–ND4L–ND6. Two main haplogroups were identified in the distribution range of the sampled areas. The samples of haplotype 1 containing the shortest sequence length (1580 bp), found in the Qazvin, Marand, Najafabad, Firuzkuh, Gonabad, Ilam, Birjand, Dehdasht, Sarpole-Zahab, Sanandaj, Borojerd, Sari, Zahedan and Shahrekurd samples, were grouped with A. m. meda KY464957. Moreover, the samples of haplotype 3 containing the longest sequence length (1588 bp) and found in the Kerman samples, was grouped with Bileh Savar and Soltaniyeh.

Genetic characteristics of the Iranian honey bee, Apis mellifera meda, based on mitochondrial genes of ND4, ND4L and ND6 and their internal transcribed spacers

Genetic characteristics of the Iranian honey bee, Apis mellifera meda, based on mitochondrial genes of ND4, ND4L and ND6 and their internal transcribed spacers

Physicochemical and Antioxidant Properties of Iranian Commercial Honeys

Background: All over the world, the quality of commercial honeys varies greatly depending on many factors. Hence, this study was conducted to evaluate the physicochemical and antioxidant properties of Iranian commercial honeys to confirm their nutritional and economical quality. Methods: Fifty three honey samples from six different types, including alfalfa, milkvetch, lotus, thyme, coriander and multifloral honeys, were analyzed for pH, free acidity, electrical conductivity, Ash, moisture, hydroxymethylfurfural, reducing sugars, glucose, fructose, proline, color, total phenolic content, ferric reducing antioxidant power and DPPH radical-scavenging activity. Results: : In total, physicochemical properties of different honey types were in the range of 4.3 ± 0.49 to 5.2 ± 0.49 for pH, 13.81 ± 2.5 to 26.22 ± 2.8 for free acidity (meq/kg), 15.95 ± 0.45 to 17.98 ± 0.97 for moisture (%), 282.2 ± 50.3 to 578 ± 47 for electrical conductivity (µS/cm), 0.14 ± 0.04 to 0.38 ± 0.03 for ash (g/100 g), 68.66 ± 4.3 to 72.09 ± 6.4 for reducing sugars (g/100 g), 34.67 ± 2.6 to 36.7 ± 2.2 for glucose (g/100 g), 31.9 ± 4.75 to 37.04 ± 4.0 for fructose (g/100 g), 6.1 ± 5.8 to 22.85 ± 10.9 for hydroxymethylfurfural (mg/kg), 426.7 ± 53.91 to 593.9 ± 108.6 for proline (mg/kg), 46.27 ± 4.3 to 96.04 ± 14.4 for color (mm) and 245.4 ± 48.45 to 573.9 ± 69.3 for total phenolic (mg/kg) content. In terms of antioxidant capacity, lotus and thyme honeys were preferable, while the highest level of DPPH radical scavenging activity and ferric reducing antioxidant power were observed in lotus and thyme honeys. Conclusion: These results confirmed that the Iranian commercial honeys meet the quality standards established by national and international regulatory agencies.

Detection and replication of deformed wing virus and black queen cell virus in parasitic mites, Varroa destructor, from Iranian honey bee (Apis mellifera) colonies

The parasitic mite Varroa destructor and the viruses it transmits to honey bees (Apis mellifera) are often mentioned as culprits of colony losses. Therefore, surveys are conducted in different countries to determine the presence and distribution of honey bee viruses. In Iran, the information about honey bee viruses is scarce and limited to their presence in honey bees, but has never been reported from V. destructor. We conducted a study for detecting five viruses in samples of V. destructor mites that were collected from 12 apiaries located in the Iranian provinces of Alborz, Ardabil, and Gilan. RT-PCR was used to diagnose the viruses, including deformed wing virus (DWV), black queen cell virus (BQCV), acute bee paralysis virus (ABPV), Israeli acute paralysis virus (IAPV), and Kashmir bee virus (KBV). Several mite samples resulted positive for DWV and BQCV, but all were negative for ABPV, IAPV, and KBV. Minus RNA strands were detected for DWV and BQCV with tagged RT-PCR, indicating viral replication in the mites, which could have implications for honey bee health. This is the first report of BQCV from Iranian honey bee colonies.

Physicochemical properties of Iranian ziziphus honey and emerging approach for predicting them using electronic nose

The aim of this paper is development and evaluation of an electronic nose as a non-destructive system for predicting the physicochemical parameters of honey including ash content (AC), free acidity (FA), moisture content (MC) and pH based on its aroma. To verify the authenticity of the honey samples the HPLC analysis carried out. Artificial neural network (ANN) and Support Vector Machine (SVM) models were applied to the features extracted from the sensors responses to predict AC, FA, MC and pH. The developed models for prediction of the physicochemical parameters showed high performance based on the honey headspace gases and the best results for predicting the AC, FA, MC and pH obtained from the ANN model which the R2 values were 0.838, 0.918, 0.926 and 0.933, respectively. The results proved feasibility and applicability of using the developed e-nose as a non-destructive system for predicting the physicochemical properties of honey.

Kinetics of temperature effect on antioxidant activity, phenolic compounds and color of Iranian jujube honey

In the present study, the Iranian jujube honey was evaluated for its total antioxidant activity by DPPH assay, total phenolic content (TPC) by using the Folin–Ciocalteu reagent, and brown pigment formation (BPF). The kinetics of changes in jujube honey samples heated at various temperatures (45, 55 and 65 °C) over 10 days were studied. Increasing treatment temperature and time caused an increase in all three parameters including, antioxidant activity, BPF and TPC. Increases in BPF and TPC followed zero-order kinetics, and the rise in antioxidant activity varied depending on heating temperatures, following second-order, first-order and zero-order kinetics when samples were heated at 45, 55 and 65 °C, respectively. At 45–65 °C, activation energy values of 68 and 64.7 kJ/mol−1 were obtained for BPF and TPC, respectively. Linear relationships were observed between antioxidant activity and BPF, TPC and antioxidant activity, and BPF and TPC, such that the highest phenol content was related to the darkest honey sample. For all three parameters, heating honey to 65 °C was found to be more effective than heating to 45 or 55 °C.

Potential antimicrobial activity of honey phenolic compounds against Gram positive and Gram negative bacteria

This study is focused on the potential antimicrobial activity of honey phenolic compounds against Gram positive and Gram negative bacteria. For this purpose phenolic compounds were isolated from 33 Iranian honeys obtained from different botanical and geographical origins using solid-phase extraction (SPE). Characterization of honey extracts was carried out by HPLC-DAD-ESI-QTOF/MS. Minimum inhibitory concentration (MIC) and inhibition zone were used to determine the antimicrobial capacity of honey phenolic extracts against Gram positive (Staphylococcus aureus and Enterococcus faecalis) and Gram negative bacteria (Escherichia coli and Pseudomonas aeruginosa). The results shown that E. coli was the most sensitive bacteria and P. aeruginosa the most resistant strain. Moreover, five phenolic extracts shown lower MIC values than the whole honey pointing out that honey phenolic fraction may exert antimicrobial activity by itself and be a source of bioactive compounds to develop functional ingredients.

Genetic Variation in Iranian Honey bees, Apis mellifera meda Skorikow, 1829, (Hymenoptera: Apidae) Inferred from PCR-RFLP Analysis of two mtDNA Gene Segments (COI and 16S rDNA)

In this study, the genetic structure of Iranian honey bee (Apis mellifera meda) populations, mainly obtained from all of regions, were investigated at two different mitochondrial regions. A total of 300 worker bees were collected from 20 different populations in 20 different locations. Portions of the mitochondrial 16S ribosomal RNA (16S rDNA) and cytochrome C oxidase I (COI) genes were amplified by PCR and then subjected to RFLP pattern analysis using 8 restriction enzymes. Nucleotide polymorphisms were revealed using restriction enzyme Sau3A I, Ssp I and Taq I in COI and Bsp143I, Ssp I and Dra I in the 16S rDNA gene segment. In this study, 3 novel composite genotypes (haplotypes) were found in Iranian honey bee populations. The average haplotype diversity (h) within populations was 0.0405. Heterozygosity values, Shannon index and the number of alleles of Iranian honey bee populations were low that could be caused by low definite geographic structure of Iranian honey bee populations. Genetic distance (D) values were found to be low (0.0–0.0011) within Iranian honey bee populations. Cluster analysis based on UPGMA method revealed that all populations and samples groups be in one cluster. Also, the phylogenetic tree based on Neighbor-joining method divided 29 subspecies of honey bee to 5 distinct clusters. The Iranian subspecies honey bee composed of a shared clade with subspecies of Eastern Mediterranean, Near East and Eastern parts of Middle East (O branch). This result is very useful for the control of conservation of local honey bees, as the movement of colonies across the border line of these neighboring countries, may affect the genetic structure of honey bee populations.

Characterization and Classification of Iranian Honey Based on Physicochemical Properties and Antioxidant Activities, with Chemometrics Approach.

In the present study, the physicochemical properties and antioxidant activities of different Iranian honey samples are investigated using various multivariate techniques in order to develop a quality control model. Forty-eight Iranian honey samples were tested for 15 physicochemical and antioxidant parameters. The parameters for which the samples were tested included color intensity, moisture, electrical conductivity, pH, free acidity, diastase activity, hydroxymethylfurfural content, proline level, total phenolic content, antioxidant activity, and radical scavenging activity. The study attempted to differentiate honey samples based on origin and composition. In the study, the Iranian honey samples were classified according to their respective physiochemical properties and antioxidant activities using principal component analysis and hierarchical cluster analysis. Furthermore, the relationships between the geographical and botanical origins were determined for the samples used in the study.

Apoptotic Effect of Melittin Purified from Iranian Honey Bee Venom on Human Cervical Cancer HeLa Cell Line

Melittin, an amphipathic 26-residue peptide, is the main component of honey bee venom. Studies have been demonstrated that melittin has an inhibitory effect on proliferation of cancer cells. However, the precise mechanism of action is not completely understood. In the present study we have shown that purified melittin from Iranian honey bee venom shows anti-cancer effects on human cervical cancer cell line through induction of apoptosis. The venom was collected from Iranian honey bee (Apis mellifera meda) and melittin isolated using reversed phase HPLC. Biological activity of melittin was analyzed by hemolytic test on human red blood cells. In order to investigate whether melittin inhibits proliferation of cervical cancer cells, the viability of the melittin treated HeLa cell line was measured via MTT assay. Finally, cell death analysis was performed using Propidum iodide and Annexin V-FITC dual staining. The results showed that the half hemolytic concentration (HD50) induced by mellitin was 0.5 µg/ml in free FBS solution. IC50 obtained after 12 h at 1.8 µg/ml by MTT assay. According to flow cytometric analysis, melittin induced apoptosis at concentrations more than 1 µg/ml. These results suggest that melittin induces apoptotic cell death in cervical cancerous cells as observed by flow cytometric assay. It is concluded that melittin could be regarded as a potential candidate in future studies to discovery of new anticancer agents.

Morphometric diversity and phylogenetic relationships among Iranian honey bee (Apis mellifera meda Skorikow, 1829) populations using morphological characters

In this study, the morphometric diversity and phylogenetic relationships of Iranian honey bee populations, were investigated using 14 morphometric characteristics. A total of 2250 young adult worker bees from 20 different populations in 20 different provinces of Iran were collected during June to October 2014. The results of nested analysis of variance showed that there were significant differences (P<0.01) between the provinces for all analyzed morphometric traits indicating the existence of a diversity among them. Correlation coefficient analysis showed a high degree of association among the most of the traits. This correlation coefficient should be a putative mean to improve of certain characters in breeding of honey bee. Principal component analysis revealed three principal components explained 81.5% of the total variation. Cluster analysis using WARD method classified honey bee populations into two main groups. The first group includes the honey bees collected from North, Northwest and West portions of Iran. The second group was represented by the honey bees from Eastern North, Central and Southern regions of Iran. The phylogenetic tree based on UPGMA method divided 29 subspecies of honey bee to 5 distinct clusters. The Iranian subspecies honey bee composed of a shared clade with subspecies of Eastern Mediterranean, Near East and Eastern parts of Middle East (O branch).

The first comprehensive molecular detection of six honey bee viruses in Iran in 2015-2016

At least 18 viruses have been reported in the honey bee (Apis mellifera L.). However, severe diseases in honey bees are mainly caused by six viruses, and these are the most important in beekeeping. These viruses include: deformed wing virus (DWV), acute bee paralysis virus (ABPV), chronic bee paralysis virus (CBPV), sacbrood virus (SBV), kashmir bee virus (KBV), and black queen cell virus (BQCV). In this study, we evaluated 89 Iranian honey bee apiaries (during the period 2015-2016) suffering from symptoms of depopulation, sudden collapse, paralysis, or dark coloring, by employing reverse transcription-PCR. Samples were collected from four regions (Mazandaran, Hormozgan, Kurdistan, and Khorasan Razavi) of Iran. Of the 89 apiaries examined, 16 (17.97%), three (3.37%), and three (3.37%) were infected by DWV, ABPV, and CBPV, respectively. The study results for the other viruses (SBV, KBV, and BQCV) were negative. The present study evaluated the presence of the six most important honey bee viruses in bee colonies with suspected infections, and identified remarkable differences in the distribution patterns of the viruses in different geographic regions of Iran.

The synergistic effect of honey and cinnamon against Streptococcus mutans bacteria

ObjectiveTo investigate the effect of Iranian honey, cinnamon and their combination against Streptococcus mutans bacteria. MethodsNine experimental solutions were examined in this study, including two types of honey (pasteurized and sterilized), two types of cinnamon extract (dissolved in distilled water or dimethyl sulfoxide) and five different mixtures of cinnamon in honey (prepared by admixing 1%–5% w/w of cinnamon extract into 99%–95% w/w of honey, respectively). Meanwhile, each of mentioned agent was considered as the first solution while it was diluted into seven serially two-fold dilutions (from 1:2 to 1:128 v/v). Therefore, eight different concentrations of each agent were tested. The antibacterial tests were performed through blood agar well diffusion method, and the minimum inhibitory concentration (MIC) was determined. Ultimately, the data were subjected to statistical analysis incorporating Two-way ANOVA and Bonferroni post hoc tests (α = 0.01). ResultsThe highest zone of inhibition was recorded for the mixtures of honey and cinnamon while all the subgroups containing 95%–99% v/v of honey were in the same range (P < 0.01). The MIC for both honey solutions were obtained as 500 mg/mL whereas it was 50 mg/mL for both cinnamon solutions. Moreover, the MIC related to all honey/cinnamon mixtures were 200 mg/mL. ConclusionsA profound synergistic effect of honey and cinnamon was observed against Streptococcus mutans while there was no significant difference among extracts containing 99%–95% v/v of honey admixing with 1%–5% v/v of cinnamon, respectively.

Physicochemical properties and biological activities of honeys from different geographical and botanical origins in Iran

In recent years, the pharmaceutical interest in honey due to its well-known health properties has increased. In the present study, 33 Iranian honey samples from different geographical and floral origins were characterized for their physicochemical, antioxidant, and antimicrobial properties. Variations of the quality parameters, total phenolic content (TPC), antioxidant properties, and antimicrobial effects were correlated with the geographical and botanical origins to establish the best source of honeys analyzed. Honey samples collected in south Iran showed the highest TPC. Antioxidant activities varied markedly among the honey samples, even in honeys within the same floral origin. Comparative assessments of honeys indicated the marked correlation between antioxidant activity and total phenolic content. Color intensity (ABS450) of honeys was also found to be correlated with total phenolic content and the overall antioxidant activity. Antibacterial activity against both Gram-positive and Gram-negative bacteria revealed minimum inhibitory concentration values from 6.25 to 100% (w/v). Bacteria sensitivity to the antimicrobial activity of honeys followed the order E. coli > S. aureus > P. aeruginosa > E. faecalis. Remarkable differences were pointed out in the ability of each honey sample to elicit antimicrobial responses. Besides, bacterial sensitivity to honey samples was up to 16 times higher in some samples than the expected results based on previous studies in the literature. Consequently, Iranian honeys may be sources of antioxidants and antimicrobial agents that could be used as food antioxidants or as ingredients in nutraceutical products due to their interesting pharmaceutical properties.

First report on the isolation of melittin from Iranian honey bee venom and evaluation of its toxicity on gastric cancer AGS cells

BackgroundIt has been previously reported that melittin, the main ingredient of honey bee venom, has anticancer properties. However, there appears to be no earlier study focusing on the isolation of melittin from Iranian honey bee venom (Apis mellifera meda), and evaluation of its effect on cancerous cells. MethodsWe isolated melittin using reversed-phase high performance liquid chromatography, and its potential toxicity on gastric cancer AGS cells was determined with an MTT [3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide] assay. Furthermore, to ascertain whether melittin induces apoptosis or necrosis in these cells, morphological evaluation, DNA fragmentation assay, propidium podide and annexin-V-FITC dual staining, and flow cytometric analysis were also conducted. ResultsThe results of our study suggested that melittin inhibited the proliferation of AGS cells in a dose and time-dependent trend. All of the above four distinct assays indicated that melittin induces necrosis in AGS cells at concentrations of ≥ 1 μg/mL. ConclusionThe present study indicated that melittin has an anticancer effect on gastric cancer AGS cells and stimulates necrotic cell death in these cells.

Assessment of the physicochemical quality of Iranian honey

Honey samples supplied by Iranian beekeepers in different regions of Iran were analysed for moisture, acidity, diastase activity, hydroxymethylfurfural (HMF), invert sugar, sucrose, ash and proline content. The mean values found were 18.35±5.6% for moisture, 24.45±12 meq/kg for acidity, 199.87±96.55 DN for diastase activity, 17.66±15.35 mg/kg for HMF, 37.31±17.13% for invert sugar, 2.62±1.9% for sucrose, 0.53±0.21% for ash, and 20.96±11.6 mg/kg for proline. The honey sample from Markazi province had the highest proline content and the highest diastase activity was observed in the honey sample from Isfahan. Honey samples from Mazandaran, Tabriz, and Ardebil had moisture contents that were greater than the EU standard (20%). The highest quality honey samples were from Kurdistan followed by Kermanshah, which showed higher quality in terms of moisture content, proline, HMF, and diastase activity.

Characterization and Classification of Several Monofloral Iranian Honeys Based on Physicochemical Properties and Antioxidant Activity

The aim of this study was to characterize and classify 10 honey samples collected from different regions of Iran based on their physicochemical properties. Moisture, ash and hydroxymethylfurfural content, pH, electrical conductivity, diastase activity, color, and fructose to glucose ratio were determined. Phenolic and flavonoid compounds, antioxidant activity, and the ability to inhibit enzymatic browning were also evaluated in the samples. Based on diastase activity and hydroxymethylfurfural content, Tamarisk honey showed the best quality. The highest ratio of fructose to glucose (1.5) belonged to Ziziphus. Coriander honey with the lowest L* (18.9) was considered darker than other samples. High correlation coefficients between phenolic, flavonoid compounds, and antioxidant activity indicated that these compounds are mainly responsible for the antioxidant capacity of honey. Based on the first principal component, honey samples were classified into four main groups. The first group included coriander and ziziphus, the second group was dill, the third group contained thyme, parsley, and qanqal, and the fourth group included astragal, alfalfa, tamarisk, and orange blossom.