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Научни трудове на Съюза на учените в България-Пловдив. Серия В. Техника и технологии, т. XV, ISSN 1311 -9419 (Print), ISSN 2534-9384 (On- line), 2017. Scientific Works of the Union of Scientists in Bulgaria-Plovdiv, series C. Technics and Technologies, Vol. XV., ISSN 1311 -9419 (Print), ISSN 2534-9384 (On- line), 2017.
HEAVY METAL LEVELS OF PROPOLIS IN R. MACEDONIA Naco Jovcevski*, Borche Makarijoskil, Stefce Presilskil, Biljana Bogdanova Popov2, Mohammad Ali Shariati3, Mohammad Usman Khan4 Mentor: d-r Vesna Karapetkovska - Hristova1
1 Faculty of Biot echnical Sciences, Unive rsity "St. Ediment Ohridski", Bitola, R.Macedonia 2 Faculty "f Natural Sciences, University of Nis, Department of Biology and Ecology, Serbia, 3Research Department, LLC «Science & Education», Russia and Researcher All Russian Research Institute of Phytopathology,
Moscow Region, Russia 4D epartment of Energy Systems Engineering, Urnveasity of AgricultureFnisalabad,3"g00,Faisalabd,oakiita"
Abstract
The aim of this stady is it determine the heavy metal levels in Patptlis samplesfrom iwt selected regions (lowland and mountain "аe") in the RepabliconMacedonia.The research material was obtained from five Uangstaoth - type beehives at each apiary, daring the period May - October lOlS.Qaantitatice analysis of the studied toxic elementswascondacted asing mms- Atomic Absorption Spectaometea after day ashing. All samples met the World Health Organization (WHO) and FAO standards regarding the maximum permitted amount of Pb> Cd > Zn.The seasonal research that was made, give results that define Propolis from this region foa ecological pure paoduct foa consuming. Keywoads/Paopolis, toxic heavy metals, Atomic absorption spectaoscopy.
Introduction
Propolis is a sticky, gummy, resinous substance collected by honeybees from various plant sources.Propolis was used for its healing by the Egyptians, Greeks and Romans etc.Propolis is collected by honeybees from buds and exudates of plants to be used as a protective barrier in the beehive. It is currently incorporated in food and beverages to improve health and to prevent several diseases. The color of propolis depends on its origin. Besides its use in folk medicines, it possesses various pharmacological activities, such as antibacterial, (TrushevaB. et al. 2010) antiviral, (Lotfy M. 2006) antifungal, anesthetic, anti-inflammatory, hypertensive, immunostimulatory, and cytostatic properties.
Nowadays, the negative influence of mankind and the development of industry especially chemical, engineering and mining industry, as well as traffic-related pollution contributed to a huge increase in environmental pollution (Porrini Claudio et al.2003).
The aim of this study was to determine the concentration of selected "heavy metals" in propolis as an indicator of contamination, from different regions, collected insummer -fall season.In Republic of Macedonia some experimental research were made in attempt to provide level of ecological safety and to prevent pollution of surrounding.Monitoring of trace elements in honey products also were investigated to determine ecological safety (Stankovska E.et.al. 2008).
2. Materials and Methodology
2.1.Area description and propolis samples
Propolis samples were collected monthly from four different areason a territory of R. Macedonia,divided in to twogroups: one lowlandarea, and one mountain area.This area and climate in is classified as Mediterranean. The combined quantitative effects of air temperature, relative humidity, and wind speed have been explored. The air temperature varied from 10 °C to 30 °C in 5 °C- intervals. The relative humidity ranged from 0% to 50%. The wind speed was from 0.1 m/s to 0.4 m/s. (http://www.weather.com/weather/wxclimatology/monthly/graph/MKXX0005)
2.2. Instrumental Analysis
Quantitative analysis of the studied elements wereconducted using AAS- Atomic Absorption Spectrometer after dry ashing. (EN 14084:2003) Foodstuffs - Determination of trace elements -lead, cadmium, zinc, copper and iron by atomic absorption spectrometry (AAS) after microwave digestion. We used the official method for the determination of heavy metals in feed and food.
2.2.1.Sample preparation for processing on AAS:
Propolis samples were collected during the period May - October, from two different areas. The research material was obtained from five Langstroth - type beehives at each apiary, from two different areas, which they disposed a"Health certificate of bee colonies and apiary products".Propolis material was collected representative, directly from the hives, with a special sharp instrument, (Ra'ed J. Abu Fareset al. 2008) from the wooden walls and frames was scraped down and collected in dark flacons-containers.Propolis samples were pooled, combining in identical quantities samples (10 gr) from the 5 months of each colony. Propolis samples were classified into 2 different groups.
It was weighted (with precision of 0.1mg)1gr of each sample-propolisin quartz potandsamples areplaced on hitting plot, for about 30min. Next, the samples are passing in another stage of heating in muffinoven at 500° C about 3h, and samples of propolis are glowing to the tarnishing of the ash. We were using 5ml distillate water filing up to the mark with 5ml 5% HNO3 (Trace Pure).Experimental material was filtering throw the filter paper and it's measured in volumetric pumpkins of 25ml, it's filth up with distillate water to the mark. Samples prepared through this typeof procedure wereplacedin cuvette of 1ml and measured on AAS.
Statistical Analyses were estimated by statistical one-way analysis of variance (ANOVA) and t-test, to present statistically significant differences in the mean concentrations of heavy metals in propolis between the groups from the determent regions and their differences. The chosen level of significance for all statistical tests was (p<0.05).
3. Results and Discussions
The analysed samples of propolishave presented a low concentrations in heavy metals. Heavy metal accumulation in propolis was dependable on the region of the sample origin. In samples
coming from a region in the lowland areas the analyzed toxic elements were present in a higher concentrationthan the elements contained in there search material from the mountain areas.
Lead
In the agricultural lowland region (X), the mean values of the Lead element in propolis samples were higher than the mean levels of the lead accumulation in the propolis from the highland area (Y), (0.40 mg/kg and 0.34 mg/kg). Statistically significant differences (p<0.05) in Lead levels between study areas were observed (Tab. № 1).
Table№ 1. Determination of the concentration of chosen element (Pb) in bee propolis (n=20), surrounding of four different areas Xand Y
X Y
Concentration 600m (ASL) 802m (ASL)
Pb (m g/kg)
Min 0.033 0.022
Max 0.049 0.042
Average 0.040 a 0.034 b
SD 0.005 0.007
CV(%) 12.25 20.58
* a. bDifferences between average values with the different letters are significant on level p< 0.05
Cadmium
Cadmium concentration in the assesed propolis samples was an average of 0.030 mg/kg in samples originated from lowland region (X) and 0.014 mg/kg in the highland region (Y) (Tab. № 2). Statistically significant differences (p<0.05) in Cadmium levels between regions of the study were demonstrated. A considerably very low concentrations of Cadmium were observed by Caroli et al.(2000).
Table№ 2. Determination of the concentration of chosen element (Cd) in bee propolis (n=20), surrounding of four different areas X and Y
X Y
Concentration 600m (ASL) 802m (ASL)
Cd (mg/kg)
Min 0,027 0,011
Max 0,038 0,018
Average 0,030 a 0,014 b
SD 0.003 0.002
CV(%) 10 14.28
*a. bDifferences between average values with the different letters are significant on level p< 0.05
Zinc
In the present study a very low concentration of Zinc in propolis was observed without exceeding the acceptable limits. Statistically significant differences (p<0.05) in Zinc levels between lowland
and highland regions of the study were demonstrated (Tab. № 3) . Comparable concentrations of zink were noted by Conti et al. (2001). Cvek et al. (2008) observed a much higher concentration of zinc in propolis from diferent regions of Turkey.
Table№ 3. Determination of the concentration of chosen element (Zn) in bee propolis (n=20), surrounding of four different areas X and Y.
Concentration X Y
600m (ASL) 802m (ASL)
Zn (mg/kg)
Min 0,021 0,013
Max 0,038 0,031
Average 0,030 a 0,020 b
SD 0,006 0,005
CV(%) 20 25
* ^ bDifferences between average values with the different letters are significant on level p< 0.05
Conclusion
Many factors influence the heavy metal contents of propolis, such as geographical and botanical origin, soil, atmosphere, beekeeping equipments/practices, elemental composition of nectar, season of the year, rainfall and anthropogenic activities among the others. Our present results indicate that, concentration of the selected heavy metals in the samples of propolis from the selected areas are generally not considered contaminated and are well below the standard regulations.
References
Caroli, S., et al. (2000). A pilot study for the production of a certified reference material for trace elements in honey. In Microchemical Journal, vol. 67, 2000, p. 227-233.
Cvek, J., et al.(2008). The content of essential and toxic elements in Croatian propolis samples and their tinctures. Journal of Apicultural Research, 47, p.p.35-45.
Conti M.E., Botre F. (2001) Honeybees and their products as potential bioindicators of heavy metals contamination, Environ. Monit. Assess. 69, 267-282.
Trusheva Boryana., et al. (2010). Antibacterial mono- and sesquiterpene esters of benzoic acids from Iranian propolis,Chemistry Central Journal, p.p.4-8.
Mahmoud Lotfy., (2006). Biological Activity of Bee Propolis in Health and Disease, Asian Pac J Cancer prev, Vol 7, p.p.22-31.
Porrini Claudio., et al. (2003). Honey bees and bee products as monitors of the enviromental contamination, Apiacta, p.p.63-70.
BiljanaBogdanova Popov., et al. (2017)."Assesment of heavy metals in propolis and soil from Pelagonia region, Republic of Macedonia",Macedonian Journal of Chemistry and Chemical Engineering, Vol 36 (No. 1), pp. xx-xx.
Stankovska E., et al.(2008). Monitoring of trace elements in honey from the Republic of Macedonia by atomic absorption spectrometry, Environmental monitoring and assessment, 142, p.p. 117-126 Ra'ed J. Abu Fares., et al. (2008). Honey Bee Hive Modification for PropolisCollectionJordan Journal of Agricultural Sciences, Volume 4, No.2.
FAO-Food and Agriculture Organizationavailable at: htm:// www.fao.org. WHO-World health organization
