الفهرس 
INTRODUCTIONOnly 14 pages are availabe for public view
Abstract
Honey bees are among the most important insects that humans have benefited from for thousands of years in food and medicine. Among the products of honey bee colonies, propolis, which has received a lot of attention in recent years because of its wide range of desirable biological activities, which includes antimicrobial activity, so there was a need for more exploratory studies on its chemical composition, and potential biological activities, in addition to factors that affect the efficiency and quality of the produced propolis, including the identification of appropriate extraction methods aimed at utilizing its biologically active components and producing an effective and high-quality extract, and for the development of appropriate production methods that must be applied to produce the largest quantity of high-quality propolis.
Laboratory experiments were conducted for the current study in the desert research center (D.R.C) laboratories, while field experiments were conducted to evaluate the propolis traps in three apiaries, two of them were in Dakhla district, new valley governorate (El-Hindaw and El-Mouhoub villages) and third location was in Qalyubia governorate (Qalyub center) during the experimental period extended from July, 2017 to June, 2018.
The current work mainly aims to evaluate three bee propolis-collection traps in addition the traditional method (hand collection) in terms of the quantity and quality of the produced propolis, in addition to evaluating the effect of three different extraction methods (maceration, microwave-assisted, ultrasonic-assisted) on The yield of the extract of the used propolis samples, contents of total phenolic and flavonoid compounds, as well as the antimicrobial properties of extracts on strains bacteria tested.
1. Evaluation of some bee propolis–collection traps in terms of the quantity and quality of propolis produced in New Valley and Qalyubia Governorates during 2017/2018
1.1. Evaluation of bee propolis–collection traps in terms of the quantity of propolis produced
The results represented that, bee propolis – collection trap Type (2) (Hand-cut fiber screens with circular holes in a diameter of 1 mm.) significantly outperformed all other types of bee propolis collecting traps in terms of the amount of propolis produced, followed by the type (1) trap (Glass slides, with 48 cm length, 6 cm width and 6 mm thickness), then the type (3) trap (Hand-cut polypropylene plastic screens with round holes in diameter of 2 m.), while the lowest amount of bee propolis that Collected in the traditional method (hand collection). The results also revealed that the highest amount of bee propolis collected was in the month of July, with an average of 11.19, 11.20 and 5.96 g/colony, in the El-Hindaw, El-Mouhoub and Kalama respectively.
Regarding the season, summer season significantly had the highest amounts of bee propolis produced with an average of 8.46, 8.44 and 4.87 g/colony, in the El-Hindaw, El Mouhoub and Kalama respectively, while the autumn season had the lowest amounts of bee propolis produced with an average of 1.61, 1.58 and 0.83 g/colony in the El-Hindaw, El Mouhoub and Kalama respectively.
It can be concluded that, the highest annual average of the collected bee propolis /colony was obtained with (Type, 2) trap 68.13, 67.99 and 42.81 g, while the lowest annual average was recorded in traditional method with an average of 45.05, 44.67 and 23.15 g/colony in the El-Hindaw, El Mouhoub and Kalama respectively.
1.2. Evaluation of bee propolis–collection traps in terms of the quality of propolis produced
Bee propolis collection traps were evaluated in this study in addition to the traditional method in terms of yield ethanolic propolis extract (EEP) by weight of crude propolis (5 g) collected in August 2017 and extracted by maceration by using 70% ethyl alcohol. Three replicates of each group were tested to estimate the yield ethanolic propolis extract (EEP).
The results concluded that there are no differences in the quality of the bee propolis produced from the tested bee propolis- collection traps, while the quality of the propolis produced from the traditional method (hand collection) of collection was significantly lows noticeable than any of the tested bee propolis- collection traps In all three experimental sites (El-Hindaw, El Mouhoub and Kalama). where yield % of ethanolic extract propolis (EEP) was recorded in bee propolis – collection traps Types, 1, 2 and 3 with yield % 35.36, 35.17 and 34.39 %, respectively. While recorded 19.95 % with traditional method El-Hindaw. At El-Mouhoub also propolis traps (1, 2 and 3) recorded yield % 36.02, 36.10 and 34.79 % respectively, while traditional method recorded yield % 20.75. Also in Kalama the same trend was obtained as shown in the results obtained in the El-Hindaw and El-Mouhoub, with a decrease in yield of propolis extract where, yield % recorded 27.75, 26.80 and 25.88 %, by propolis trap type 1, 2 and 3 respectively. While recorded 14.14 % with traditional method.
2. Evaluation of the Extraction technique on yield of Ethanolic Extract Propolis (EEP)
The experiment aims to evaluated the techniques of different extraction methods in terms of the percentage of dry extract of raw propolis.
The results showed that a significant difference was recorded in the percentage of propolis extraction yield in all tested techniques, where the highest percentage of extraction yield (%) was recorded in microwave-assisted extraction, followed by ultrasonic-assisted extraction, and the lowest percentage of extraction yield (%) in the extraction by maceration method, where the percentage yield (%) was an average of 22.75, 19.44 and 14.85% for microwave-assisted extraction (MAE), ultrasonic-assisted extraction (UAE) and maceration (ME) respectively.
The obtained results showed a significant superiority of the New Valley propolis sample over the rest of the propolis samples for all extraction techniques in terms of extraction yield (%) with an average of 28.18%, followed by the Gharbia propolis sample with an average of 23.57%, then Qalyubia and Sohag propolis samples without significant differences between them with an average of 17.01 and 15.58%, respectively, while the imported propolis sample recorded the lowest percentage of the extraction yield (%) with an average of 10.72%.
The obtained results showed that the highest percentage of propolis extract was achieved by using the microwave-assisted extraction (MAE) method for the New Valley Propolis sample with an average of 33.33%, followed by the New Valley Propolis sample extracted by ultrasonic-assisted (UAE) and Gharbia propolis sample extracted by microwave-assisted (MAE) without significant differences between them with an average of 29.61 and 28.95, respectively, while the imported propolis sample recorded the lowest percentage of the yield of propolis extract using the maceration technique with an average of 8.41%..
3. Screening for antibacterial activity of ethanolic extract propolis (EEP).
The activity of extracts of different samples (imported propolis, Gharbia, Sohag, Qalyubia, and New Valley) was evaluated by different extraction techniques (maceration, microwave, ultrasonic) on the tested Pathogenic bacterial Strains (S. typhimurium ATCC25566, E. coli ATCC 8739 and P. larvae DW5708) where, determine inhibition zone diameter (mm) of propolis samples extracted with different techniques methods, minimum inhibitory concentration (MIC) of propolis samples and minimum bactericidal concentration.
3.1. Evaluation of antibacterial activity of ethanolic extract propolis (EEP).
Results shown that, the propolis samples taken from New Valley and Gharbia which extracted by ultrasonic extraction were given the highest significant inhibition against P. larvae (with diameter zone of 25.84 and 22.10 mm). E. coli ATCC 8739 (with diameter zone of 23.41 and 20.22 mm) and S. typhimurium ATCC 25566 (with diameter zone of 21.24 and 17.53 mm), respectively.
3.2. Determination of minimum inhibitory concentration (MIC).
Nine sequential concentrations were used (9.25, 7.25, 5.5, 4.0, 2.75, 1.75, 1.0, 0.5 and 0.25 mg/mL), the extracts of the New Valley and Gharbia samples extracted by ultrasonic were used, (they were the best samples with the best extraction technique). The results showed the superiority of New Valley sample extract over Gharbia sample extract, where the MIC value was 1.75 mg/mL, while it was 2.75 mg/mL for E. coli ATCC 8739 and S. typhimurium ATCC25566, respectively. The obtained results conclude that, a clear superiority of New Valley sample extract over Gharbia sample extract, as it reached the lowest inhibitory concentration of P. larvae DW5708 0.5 mg/mL while it was 1.75 mg/mL for Gharbia sample extract.
3.3. Determination of minimum bactericidal concentration (MBC).
Results shown that, G-ve bacterial pathogenic E. coli ATCC 8739 and S. typhimurium ATCC25566 exhibited sensitive to propolis New Valley at MBC of 2.75 mg/mL (1/16 dilutions). While with propolis Gharbia at MBC of 5.50 mg/mL (1/4 dilution). In the instance of G+ve bacterial pathogenicity P. larvae DW5708 was susceptible to propolis New Valley at MBC of 0.50 mg/mL (1/128 dilution) and to propolis Gharbia at MBC of 2.75 mg/mL (1/32 dilution).
3.4. Evaluation of propolis mode of action.
Results revealed that the extracted propolis obtained from New Valley and Gharbia had bactericidal effect with MBC / MIC ratio ≤ 2 against 3 the tested pathogenic bacterial strains E. coli ATCC 8739, S. typhimurium ATCC25566 and P. larvae DW5708.
4. Chemical analyses of propolis.
4.1. Determination of total phenolic content.
Results represented that, the highest yield of total phenolic content was obtained with the New Valley and Gharbia propolis sample which extracted by ultrasonic assisted extraction (UAE) with 110.45 and 104.46 mg/g extract, respectively. While the lowest yield of total phenolic content was obtained with imported propolis sample which extracted by microwave-assisted extraction (MAE) and maceration extraction (ME) with 9.86 and 13.78 mg/g extract respectively. The obtained results refer that, highest content of phenol was detected by ultrasonic-assisted extraction (UAE) with an average 76.67 mg/g extract flowed by maceration extraction (ME) with an average 44.77 mg/g extract. While the lowest yield of total phenolic content was recorded with microwave-assisted extraction (MAE) with 26.89 mg/g extract. Results also represented that, New Valley propolis sample outperformed the rest of the other tested samples in terms of yield of total phenolic content for all techniques extraction with an average 76.70 mg/g extract, flowed by Gharbia Propolis sample with an average 68.20 mg/g extract. while the lowest yield for total phenolic content was obtained using imported propolis sample for all techniques extraction with an average 15.40 mg/g extract.
4.2. Determination of total flavonoids content.
Results shown that, the highest yield of total flavonoids content was obtained with the New Valley and Gharbia Propolis sample which extracted by ultrasonic assisted extraction (UAE) with 20.73 and 17.51 mg/g extract, respectively. While the lowest yield of total flavonoids content was obtained with imported Propolis sample which extracted by microwave-assisted extraction (MAE) and maceration extraction (ME) with 5.43 and 6.15 mg/g extract.
Regard to the effect of extraction methods on the yield of total flavonoids content (regardless of the type of propolis sample).The highest content of phenol was detected by ultrasonic-assisted extraction (UAE) with an average 15.12 mg/g extract flowed by maceration extraction (ME) with an average 11.39 mg/g extract. While the lowest yield of total flavonoids content was recorded with microwave-assisted extraction (MAE) with 9.11 mg/g extract.
The obtained results conclude that, New Valley Propolis sample outperformed the rest of the other tested samples in terms of yield of total flavonoids content for all techniques extraction with an average 16.78 mg/g extract, flowed by Gharbia Propolis sample with an average 14.27 mg/g extract. while the lowest yield for total phenolic content was obtained using imported propolis sample for all techniques extraction with an average 6.59 mg/g extract.
4.3. Identification and quantification of bioactive compounds using gas chromatography–mass spectrometry (GC–MS).
GC-MS chromatogram of the New Valley and Gharbia propolis samples were identification of chemical composition of propolis extracts was assessed by mass spectral analysis and the retention time RT of each compound.
The total number of compounds identified from sample collected from New Valley governorate was 65 and 66 from sample collected from Gharbia governorate
A difference was found in the compounds that were identified in the two samples of propolis for the New Valley and Gharbia, with the exception of nine compounds common, with a different districts ratio. as follows: Glycerol, Oleic acid, Caffeic acid, Eicosane, Erythritol, Thunbergol, Batilol, Betulinaldehyde and 1-Heptacosanol with percentage of the area 1.53, 1.38, 0.48, 0.21, 0.29, 0.44, 0.35, 3.19 and 0.19 %, respectively in New valley and 4.67, 0.43, 0.80, 0.24, 0.34, 0.48. 0.57, 1.59 and 0.28 %, respectively in Gharbia.