International Journal of Basic & Applied Sciences IJBAS-IJENS Vol:14 No:02 4 Physical and Chemical Characteristics of Olive Oils from Cooperatives for Olive Growers in the North of Morocco I. Ibrahim Hassan Abdalla1, M. khaddor1, A.Boussab1, D. El Garrouj2 and B. Souhial3 1. Laboratory of physical chemistery of material , natural substance and environment, faculty of Science and Technology.University of Abdelmalik Essaadi Tanger Tetouan 2. Regional Directorate of Agriculture Tangier - Tetouan (Department of local product) 3. Faculty of science Tetouan- University of Abdelmalik Essaadi Tanger Tetouan correosponding auther E- mail: [email protected] Abstract-- The purpose of this study was to evaluate the physical and chemical characteristics of olive oils from cooperatives for olive growers and allowed us to identify and classify the oils. The samples which were studied collected from the North of Morocco are specific to the region of Ouazzane in Loukkos were examined for physical and chemical properties (acidity, pH, peroxide values, saponification number, iodine value, ultraviolet spectrophotometric analysis (k232 and k270), refractive index), fatty acids composition and total phenol content. Olive oil was analyzed for fatty acids commonly present in olive oils which are palmitic, palmitoleic, stearic, oleic, linoleic, linolenic, and arachidic. Oliec acid was found in high percentage ranged from (64.80 to 72.80), followed by palmitic, Linoleic, palmitoleic, stearic and linolenic. Arachidic acid was detected in all olive oil samples but in low percentage. Total phenol contents expressed as gallic acid of olive oils values ranged from (112 to 313 mg/kg). All olive oil samples were compared with International Olive Oil Council (IOOC), were not exceed the limits and exhibited remarkable physicochemical properties and could be useful as edible oils and industrial applications. Index Term-- olive oils, physical and chemical properties, fatty on many factors related to olive tree cultivation and to the harvesting, storage and olive processing steps and time of particular importance for olive oil quality are the olive cultivar, the pedoclimatic conditions of cultivation, as well as the pruning, fertilization and irrigation of olive trees and this illustrates the need to determine the quality of olive oil from a range of harvest times and cultivars to establish an optimum harvest time [3]. Plant density and number of processing steps required; mainly crushing, malaxation and centrifugation effect on the quality of the oil [4]. The objective of this study was to evaluate physical and chemical characteristics of local olive oils in north of Morocco from cooperatives for olive growers, and samples were subjected to comparison to the International Olive Oil Council which have designated the quality of olive oil, based on physicochemical analysis especially the quantity of free fatty acids (FFA) is an important factor for classifying olive oil into commercial grades and this certification increases the oil value [5]. Indeed, the varietal characteristics appreciated by the chemical composition analysis which is extremely influenced by the agronomic and technological conditions [6]. acids, total phenol, North of Morocco I. INTRODUCTION Olive oil, an important component in the diet of Mediterranean people, is obtained by mechanical extraction from the fruit of the Olea europaea L. tree, which belongs to the Olive family, comprises some 400 species, and thrives in temperate and tropical climates [1]. In Morocco especially in Ouazzane region (northern Morocco), the Picholine cultivar is a population variety which dominates the olive zone (90% of the cultivated olive patrimony). Nonetheless, there are other olive varieties which are not widely spread and which are distinguished by their own biometric characteristics, especially, autochthon varieties are protected by the population. Ouzzani et al. [2], have demonstrated that these varieties present genotypic and phenotypic variations. Olive oil quality depends II. MATERIAL AND METHODS Olive oils samples Ten olive oils samples were collected from Loukkos areas from cooperatives for olive growers in North of Morocco are specific to the region of Ouazzane such as Azohour Harara (ZH), Bulouaan Asjenne (BS), Beni Hamed algarbiah (BH), Aljauhara (JA), Alraihan Beni Kalaa (RB), Alwifaag Beni Darkol (BD), Alamaal Mounasara (AM), Masmouda (MS), Alnajaah Brikcha (NB) and Cefchaouen (CF). Samples were collected during the period when olives are usually harvested for oil production (crop season, November 2010). The olive oils samples were placed into sterilized bottles. Chemical and physical analysis Chemical analysis (free oil acidity, peroxide value and iodine value) were performed according to AOAC [7]. Saponification number was determined using the method 141402-6969- IJBAS-IJENS @ April 2014 IJENS IJENS International Journal of Basic & Applied Sciences IJBAS-IJENS Vol:14 No:02 described by Lecoq [8]; the pH measurement of olive oils were obtained with a pH meter (Consort) was calibrated with standard solutions buffered. Refractive index was determined by means of Abbe refractometer according to AOAC [7]. Absorbance at 232 and 270 nm was determined in a spectrophotometer (Shimadzu UV-2450, Japan), using the pure cyclohexane as the blank. The Wijs method was used for the determination of the iodine value (IV). For the free oil acidity, a known weight of olive oil was dissolved in a mixture of diethyl ether/ethanol (1:1 v/v). The mixture was titrated with potassium hydroxide in methanol (0.05M) in the presence of phenolphthalein as indicator. For peroxide value, about 5g of olive oil was dissolved in a mixture of acetic acid/ chloroform (3:2 v/v), and saturated solution of KI (1ml) was then added. The liberation iodine was titrated with sodium thiosulphate solution (0.05M) in the presence of starch as indicator. For saponification number, a known weight of olive oil (1g) was dissolved in alcoholic potassium hydroxide (25 ml) then evaporated for 30mn. The sample was titrated with chlorydric acid (0.5N) in the presence of phenolphthalein as indicator. 5 pure standards. Results were expressed as percent of relative area [9]. Determination of total phenol content The total phenol content was determined according to the methods described by Ollivier [10]. Total phenol was isolated from a solution of oil in hexane by tripleextraction with water – methanol (40: 60 v/v). The total phenols in the oil extracts were measured by the FolinCiocalteu assay. The measurement was carried out at 765nm via UV-spectrophotometer. Results were expressed as mg of gallic acid equivalent in one kg oil. Statistical analysis Analyses were performed in triplicate and statistical analysis of the data was done by analysis of variance (ANOVA) using a SPSS (the 10th version) was used. Duncan’s Multiple Range Test was used to compare mean variance. Significance was accepted at 5% level probability following Steel procedures [11]. Data were expressed as mean values ± standard deviation derived from triplicate determination. III. RESULTS AND DISCUSSION Analysis of fatty acids In order to determined fatty acid composition (%), the methyl-esters were prepared by vigorous shaking of a solution of oil in hexane (0.1 g in 2 ml) with 0.2 ml of 2 N methanolic potassium hydroxide solution and analyzed by GC gas-chromatographic where fatty acid methyl esters were performed on a Perkin Elmer gas chromatograph, equipped with a flame ionization detector (Shimadzu QP2010). The column was a fused silica capillary SE30 length 25meters, diameter 0.25 µm. Helium was the carrier gas (6ml/min). The column temperature program was initially isotherm at 140°C for 10min, an initial programmed rate of 1°C/min up to 160°C, then a second rate of 2°C/min up to 220°C and a final isotherm for 15min. Samples were injected into the split mode. The apparatus itself carried out recording and integration. The gaschromatographic peaks were identified as corresponding fatty acid methyl esters by check of the elution order on the column and compared the retention times with those of Chemical and physical properties The Chemical and Physical characteristics of olive oils sample were shown in Table 1 and table 2. There were little differences found in chemical values (free fatty acid and peroxide value) among olive oils samples. These properties especially depend on the initial quality of the olives samples. In table. 1 Acidity (% oleic acid) was in the range (0.25 – 0.57 %). According to the International olive oils council ( I.O.O.C, 2007), olive oils should have acidity (%) ≤ 3.3%, and the acidity contents of all samples were not higher than limited. There were no big significant differences among samples except (BH) sample have somewhat high acidity 0.57 % compare to the rest of samples shown in figure1. Also the results obtained, indicate that peroxide values were not higher and ranged between 6.58 meq O2/kg and 10.14 meq O2/kg. Table I Physicochemical properties (free fatty acid, peroxide value, K232 and K270) of olive oils samples (crop season, November 2010). Sample No. 1 2 3 4 5 6 7 8 9 10 Sample code FFA (% Oliec acid) ZH BS BH JA RB BD AM MS NB CF 0.25a ± 0.003 0.27a ± 0.006 0.57d ± 0.00.3 0.25a ± 0.004 0.29b ± 0.006 0.30b ± 0.004 0.33c ± 0.008 0.29b ± 0.005 0.29b ± 0.003 0.30b ±. 0.004 Peroxide value (meq O2 /kg) 8.51d ± 0.4 8.51d ± 0.07 10.14e ± 0.04 10.10e ± 0.08 7.50b ± 0.02 7.47b ± 0.07 6.58a ± 0.03 8.38c ± 0.02 8.39c ± 0.03 10.09e ± 0.07 K232 K270 3.18b ± 0.03 3.17b ± 0.01 3.45d ± 0.04 3.22bc ± 0.05 3.16b ± 0.02 3.22bc ± 0.06 3.16b ± 0.03 3.25c ± 0.02 2.86a ± 0.02 3.45d ± 0.03 0.35a ± 0.05 0.37a ± 0.08 0.47ab ± 0.07 0.55bc ± 0.06 0.34a ± 0.11 0.38a ± 0.11 0.62c ± 0.08 0.46ab ± 0.13 0.32a ± 0.06 0.34a ± 0.08 Different letters within a column indicate significant differences (p < 0.05) 141402-6969- IJBAS-IJENS @ April 2014 IJENS IJENS International Journal of Basic & Applied Sciences IJBAS-IJENS Vol:14 No:02 It is clear that peroxide values of all olive oils samples were under the value of 20 meq O2/kg of olive oil, which is the maximum established by the Council for International Olive Oil. Figure 2 shows peroxide values, where high peroxide value was found in samples coded BH, JA and CF; and when we compared these values with others 6 studies there were no differences due to the same pedoclimatic factors and the same olive trees species, which had been approved by Boukachabine et al. in the north of morocco [12]. Fig. 1. Shows free fatty acid values ((% oleic acid) of olive oils samples (crop season, November 2010). Fig. 2. Shows peroxide values (meq O2 /kg) of olive oils samples (crop season, November 2010). Table I shows specific extinction at 232 nm range from (2.86 – 3.45) and 270 nm range from (0.32 – 0.62), were not exceeding permitted limits according to IOOC [5]. The values shown somewhat significant difference at E270 of all olive oils samples and it was clear in figure 4 they were fluctuated. In figure 3 all samples have close values of E232. Fig. 3. Shows specific extinctions at (232 nm) of olive oils samples (crop season, November 2010). 141402-6969- IJBAS-IJENS @ April 2014 IJENS IJENS International Journal of Basic & Applied Sciences IJBAS-IJENS Vol:14 No:02 7 Fig. 4. Shows specific extinctions at (270 nm) of olive oils samples (crop season, November 2010). Table II shows the results of pH, saponification numbers, iodine values and refractive index. Samples coded BH had lowest pH (4.61), where approved; it had a high acidity value. Saponification number of all olive oils samples ranged from 187.05 mg KOH/ g to 194.85 mg KOH/ g, were not below acceptable limits according to IOOC [5]. Table II Physicochemical properties (pH, Saponification number, iodine value and refractive index) (crop season, November 2010). Sample No. 1 2 3 4 5 6 7 8 9 10 Sample code ZH BS BH JA RB BD AM MS NB CF pH 5.05d ± 0.04 4.95c ± 0.005 4.61a ± 0.01 5.08d ± 0.01 4.96c ± 0.01 4.98c ± 0.01 4.81b ± 0.02 4.96c ± 0.01 4.96c ± 0.01 4.82b ± 0.01 Saponification number (mg KOH / g) 188.71ab ± 1.63 192.48c ± 0.97 187.05a ± 1.81 189.55b ± 0.65 187.15a ± 0.78 187.59a ± 0.70 187.93ab ± 0.21 192.48 c ± 0.52 194.85d ± 0.42 194.82d ± 0.59 Iodine value (g /100 g) 79.44a ± 0.68 83.12ab ± 3.16 81.21ab ± 4.12 81.50ab ± 0.20 82.59ab ± 4.58 85.95bc ± 0.25 84.86b ± 3.63 90.43cd ± 1.30 90.66cd ± 1.75 91.38e ± 2.82 Refractive Index 1.4701e ± 0.00005 1.4702e ± 0.0002 1.4688a ± 0.0001 1.4700e ± 0.00005 1.4693bc ± 0.0002 1.4692bc ± 0.0002 1.4689ab ± 0.00005 1.4697d ± 0.0002 1.4693c ± 0.0003 1.4691abc ± 0.0002 Different letters within a column indicate significant differences (p < 0.05). Figure 5 and figure 6 shown sample coded BH had lowest pH and saponification number. The saponification values indicate oxidation and decrease suggest the unset of oxidation [13]. Fig. 5. Shows pH of olive oils samples (crop season, November 2010). 141402-6969- IJBAS-IJENS @ April 2014 IJENS IJENS International Journal of Basic & Applied Sciences IJBAS-IJENS Vol:14 No:02 8 Fig. 6. Shows saponification number (mg KOH/g) of olive oils samples (crop season, November 2010). Iodine value of all olive oils samples ranged from (79.44 - 91.38 g of iodine/100 g oil), they were not exceeding limits that give very strong indication of degree of instauration of a molecule [14]. Refractive index ranged from (1.4688 – 1.4702), were within permitted limits for olive oils [5]. These parameters have important roles which were influence on the quality of olive oils in the North of Morocco [15]. Figure 7 showed sample coded ZH had lowest iodine value. Whereas figure 8, showed sample coded BH had lowest refractive index compared to the rest of samples. Fig. 7. Shows Iodine value (g/100g) of olive oils samples (crop season, November 2010). Fig. 8. Shows refractive index of olive oils samples (crop season, November 2010). Fatty acid composition The fatty acid compositions of the ten olive oils samples were determined by gas chromatography and the results are shown in table 3. Olive oils contain fatty acid present in olive oils samples, such as palmitic, palmitoleic, stearic, oleic, linoleic, linolenic and arachidic acid had specific carbon number as C16:0, C16:1, C18:0, C18:1, C18:2, C18:3 and C20:0 respectively. When examining the fatty acid composition, there were differences among the samples were observed. Oleic acid was present in the highest concentration; the values were ranged between ( 64.80 % and 72.80 %). It was followed by palmitic acid (11.34 % – 17.33 %), linoleic acid (6.08 % – 11.68 %), stearic acid (1.04 % – 2.06 %), palmitoleic acid (1.00 % – 1.64 %), linolenic acid (0.16 % – 0.96 %) and arachidic acid (0.28 % – 0.92). Sample coded JA contained the highest concentration of oleic acid (72.80 %) but sample coded BS has a lowest percentage of the same fatty acid (64.80 %). Palmitic fatty acid were not exceeding 18 %, this results were agreement with those obtained by Boukachabine et al. [12] in the same region. 141402-6969- IJBAS-IJENS @ April 2014 IJENS IJENS International Journal of Basic & Applied Sciences IJBAS-IJENS Vol:14 No:02 9 Table III Fatty acid contents of olive oils samples (crop season, November 2010). Sample No. 1 2 3 4 5 6 7 8 9 10 Sample code Oleic Palmitic Linoleic Stearic palmitoleic linolenic Arachidic. ZH AS BH JA RB BD AM MS NB CF 70.96 64.80 71.97 72.80 65.31 65.55 65.78 68.99 65.53 72.32 16.88 16.72 11.34 11.95 13.16 11.77 17.33 11.94 15.94 13.64 6.08 11.68 8.26 7.94 9.63 9.35 7.43 8.85 7.49 7.06 2.06 1.14 1.56 1.04 1.80 1.94 1.80 2.03 1.74 1.39 0.80 0.72 0.70 0.75 1.01 0.46 0.71 1.64 1.00 0.98 0.16 0.64 0.67 0.96 0.45 0.52 0.90 0.64 0.49 0.84 0.52 0.84 0.92 0.85 0.76 0.50 0.59 0.28 0.55 0.66 El Antari et al. [16] have demonstrated that the level of linolenic fatty acids decrease from south to the north of Morocco, the current study confirms the same finding concerning the cooperative olive oils samples, which normally register proportions generally lower than 1 % in North region. Among the factors that influence the oils composition of fatty acids and especially oleic acids, altitude, climate, variety and olive maturity are mentioned [17]. The fatty acid compositions of the olive oils samples were obvious shown in figure 9 that oleic acid had a highest percentage, whereas arachidic was the lowest in all olive oil samples. Fig. 9. Shows fatty acid content of olive oils samples from cooperatives growers (crop season, November 2010). Palmitoleic fatty acid and linolenic fatty acid in low percentage, whereas palmitic fatty acid somewhat high and was not exceeding (18 %). When these compared to study, which conducted by Aparicio and Luna [18], the main fatty acids of the virgin olive oils in Spain was range between 9.17 – 11.6 % palmitic, 2.2 – 2.4 % stearic, 78.1 – 80.3 % oleic, 4.8 – 5.7 % linolenic acid respectively. It was found that oleic, stearic and linolenic fatty acids have low percentage, but palmitic fatty acid was high compared to Aparicio and Luna study. Oleic acid is monounsaturated and therefore contributes to that stability. However, linoleic and linolenic acids, despite their perceived nutritional benefits, are both susceptible to oxidation as they are polyunsaturated [19]. Total phenol contents The results of total phenol contents of olive oils samples are shown in table 4. Total phenol contents expressed as gallic acid of olive oils values ranged from 112 and 313 mg /kg. Total phenol content as gallic acid equivalent in sample coded RB was the highest (313 mg/kg) but sample coded BH had a lowest percentage of these components (112 mg/kg) and rest of samples have closed values, shown in fig.10. These results showed a significant difference in total phenol contents of olive oils samples. These differences may be due to v a r i e t y and total phenol contents of the samples can be considered medium-high levels in accordance with previous and found within reported by Boukachabine et al. [12]. 141402-6969- IJBAS-IJENS @ April 2014 IJENS IJENS International Journal of Basic & Applied Sciences IJBAS-IJENS Vol:14 No:02 10 Table IV Total phenol content of olive oils samples (crop season, November 2010). Sample No. 1 2 3 4 5 6 7 8 9 10 Sample code Gallic acid equivalent ZH BS BH JA RB BD AM MS NB CF 216 cd ± 3.51 233 e ± 10.11 112 a ± 4.16 309 f ± 4.36 313 f ± 3.05 208 c ± 2.52 220 d ± 7.63 189 b ± 8.54 185 b ± 6.02 218 cd ± 4.00 Different letters within a column indicate significant differences (p < 0.05). The proportions and speed of phenolic compounds decrease depend on variety [20]. Study done in the same region shown that the dominating local Moroccan olive oil cultivar had appropriate amount of polyphenol contents but the quality was a affected by storage time, it was observed a decrease in the amounts of polyphenols during storage time [21]. Figure 5 shows sample coded BH had low percentage of total polyphenols; whereas sample coded JA and RB had a highest total polyphenols percentage. Fig. 10. Concentrations of total polyphenols contents (crop season, November 2010). VI. CONCLUSION In this study, the obtained results revealed that the olive oils samples from cooperatives for olive growers were free from any defects and were classified as virgin olive oil according to international olive oils council, acidity and peroxide values were within the limits for virgin olive oil. The physical and chemical characteristics of olive oils samples showed somewhat considerable differences, but have good properties as they contain low percentages of acidity, therefore could be utilized successfully as a source of edible oil for human consumption and it contains high monounsaturated to saturated fatty acids ratio that oleic acid is monounsaturated and therefore contributes to that stability and however, linoleic and linolenic acids, despite their perceived nutritional benefits are both susceptible to oxidation as they are polyunsaturated [22]. V. ACKNOWLEDGEMENT We are thankful to Regional Directorate of Agriculture Tanger - Tetouan (Department of local product) for providing olive oils samples from cooperatives for olive growers in the North of Morocco for research work and great thankful to department of chemistry, faculty of science and technology, University of Abdelmalik Essaadi, Morocco for conducting this research. [1] [2] [3] REFERENCES N. Liphschitz in M. 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The second international symposium on Analytical Chemistry for a Sustainable Development (ACSD - 2013), the 4th Federation of African Societies of Chemistry (FASC) Congress, du 7 au 9 May 2013Marrakech –Morocco. Brodnjak-Voncina, D., Kodba, Z.C., Novic, M. Multivariate data analysis in classification of vegetable oils characterized by the content of fatty acids. Chemometrics and Intelligent Laboratory Systems 75 (2005) 31-43. 11 Isam Eldeen Ibrahim Hassan Abdalla Graduated with first class degree in Bachelor of Science in agriculture (Plant Protection) from Sudan University of Science and Technology, Sudan in October 2000. Upon graduation, he has been employed at Nile Valley University as teaching assistant and lecturer in the department of plant protection till now and he work among farmers community and help them and he is still actively involved in various consultation in Plant Protection and identification and analysis of Pesticide Residues on Foods. He decided to further his study and received M.Sc. degree of Agriculture (Plant Protection) from Gadjah Mada University, Indonesia in January 2008. He’s currently pursuing his PhD in Morocco, University of Abdelmalik Essaadi. He communicated and shared in The Second International Symposium on “Analytical Chemistry for a Sustainable Development”–ACSD 2013- The 4th Federation of African Societies of Chemistry (FASC) Congress May 7 – 9 – 2013, Marrakech – Morocco. Under the title: "The effect of storage time on the quality of dominating local Moroccan olive oil cultivar". Also he shared and communicated in The Third International collogue of Microbial Biotechnology, from 15 to 17 March 2012, Tangier – Morocco, under the title“Comparative Study in Olive oils in The North of Morocco. Khaddor Mohammed Prof. Dr. Mohamed khaddor was awarded a PhD degree in (geochemical and resource recovery, Geochemistry and valorization of the resources). He is currently a Professor in Faculty of Science and Technology of Tangier, University of Abdelmalek Essaâdi, Tangier-Tetouan, Morocco. His research interests focuses on Organic Geochemistry (soils, sediments and liquids) and valorization of natural substances like oils olives. Additionally, his research interest is moreover directed towards the field of the photo-catalysis of persistent organic pollutants (water treatment). He has published more than 15 papers. He is Director of the laboratory LAMSE and he framed 6 PhD students. He is member of the Scientific Committee of journal “Physical and Chemical News” ISSN 1114-3800, and he has reviewed several articles of scientific journals (Physical and Chemical News; J. Mater. Environ. Sci., ISSN : 2528-2028; International Journal of Environmental Analytical Chemistry; Phosphorus, Sulfur, and Silicon and the Related Elements and scientific congress). 141402-6969- IJBAS-IJENS @ April 2014 IJENS IJENS
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