Mahmoud Yolmeh; Mohammad Bagher Habibi Najafi; Reza Farhoosh; Fereshteh Hosseini
Abstract
IntroductionFood consumers tend to use natural products without any synthetic additives. Therefore, many studies have been conducted to investigate the possibility of replacing synthetic additives with natural substances in various food products.Annatto dye is a natural carotenoid pigment extracted from ...
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IntroductionFood consumers tend to use natural products without any synthetic additives. Therefore, many studies have been conducted to investigate the possibility of replacing synthetic additives with natural substances in various food products.Annatto dye is a natural carotenoid pigment extracted from the pericarp of BixaorellanaL. seeds. The major fraction of the annatto extract is 9´-cis-bixin that is soluble in oil and 9´-cis-norbixin is the major dye fraction of the alkaline extract that is soluble in water. Annatto dye creates orange to red color in food and to be used as a natural pigment in a variety of food materials including cheese, butter, margarine, confectionary and bakery products, different kinds of drinks, snacks and jams. In addition, annatto dye has antioxidant and antimicrobial activity.Nowadays, the extraction of natural dye from plant resources has become a common technology. However, complementary information using new methods and optimization of the extraction conditions seems to be necessary in order to accomplish the highest yield of extraction. Response surface method (RSM) is effective and efficient in optimizing color extraction conditions.In this study, the different conditions of extraction process were optimized through RSM in order to obtain maximum yield and best quality of annatto dye.Materials and methodsMaterialsAnnatto seeds were purchased from Hyderabad, India. All solvents were analytical grade, Merck, Germany.Extraction of annatto dyeA certain amount of annatto seeds was soaked in n-hexane for 6 hours in order to remove oils. After filtration, the defatted seeds were used for dye extraction. Since chloroform and acetone showed the highest yields of extraction during preliminary experiments, these two solvents and their mixtures were exploited for the final experiments assigning 0 for pure acetone and 100 for pure chloroform. The extracts were filtered through Whatman filter paper NO.1 and then vacuum-dried in the 1410D-2E vacuum oven (Shel Lab, USA) to produce dye powder. Low temperatures (40°C) were applied to prevent thermal dissociation of conjugated double bonds during drying. Dye measurementThe coloring strength was measured according to Vasu et al. method; model UV-160A spectrophotometer Shimadzu, Japan, at 502 nm in which bixin has the maximum absorbance value when it is dissolved in chloroform.Determination of extraction efficiencyThe obtained powder was weighed and the mass ratio of the powder to the weight of the seeds was taken into account as the extraction yield.Experimental designIn this study, Minitab® software version 16.1.1 (Minitab Inc. USA. 2010), was used and a five level four factor central composition design was created to investigate the effect of the independent variables such as temperature, extraction time, seed to solvent ratio and chloroform concentration on the dependent variables namely the extraction efficiency and absorbance values.Results and DiscussionThe values of R2, R2-adj and R2-pred revealed that the full quadratic models were the most adequate for the extraction efficiency and absorbance values.The all of the linear terms show a significant effect except the extraction time (P< 0.05). The quadric term of extraction time and the seed to solvent ratio also had a significant effect (P< 0.05) on the extraction efficiency, however, the effect of other two quadric terms was insignificant (P> 0.05). The interactive terms of extraction temperature* seed to solvent ratio(X1X3) and the seed to solvent ratio*Chloroform concentration (X3X4) had a significant effect on the extraction efficiency (P< 0.05); however, the other two interactive terms was insignificant (P> 0.05).For the absorbance values, the all of the linear terms show a significant effect (P < 0.05); the quadric term of extraction temperature (X12) and the seed to solvent ratio (X32) also had a significant effect (P< 0.05) on absorbance values, but, the effect of other two quadric terms (X22 and X42) did not show a significant effect (P> 0.05). The all of interactive terms was insignificant (P> 0.05).An increase in the extraction efficiency was observed with the increasing temperature. Banik and Pandey while extracting oleanolic acid from Lantana camararoots demonstrated that as temperature increases extraction efficiency improves too. However, at temperatures higher than 70 °C, the annatto seed pigments were degraded and the response was reduced so the quadratic effect of temperature was negative. The absorbance value was increased by increasing the temperature; however, the absorbance value decreased at higher temperature by thermal decomposition and damage of the conjugate double bond.The absorbance value increased by increasing the chloroform concentration and seed to solvent ratio initially, however, subsequently decreased due to the damage of the conjugate double bond in higher chloroform concentration and saturation of solvent in higher seed to solvent ratio.Temperature of 48.33 ˚C, extraction time of 2 hr, the ratio of seed to the solvent of 12.88 and chloroform concentration of 100% were found to be as the optimum conditions of the process. The extraction efficiency of 3.95 percent of annatto seed and absorbance value of 0.597 were acquired as the predicted results.
Mahmoud Yolmeh; Mohammad Bagher Habibi Najafi; Mahmoud Najafzadeh
Abstract
Introduction: One of the most important aspects of food preservation is controlling the growth of microorganisms, which if overlooked it leads to uncontrolled growth of microorganisms associated with food spoilage and food poisoning. Microbial contamination of foods is important because of pathogens ...
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Introduction: One of the most important aspects of food preservation is controlling the growth of microorganisms, which if overlooked it leads to uncontrolled growth of microorganisms associated with food spoilage and food poisoning. Microbial contamination of foods is important because of pathogens are capable to transfer to foods during the processing, distribution, and storage. Escherichia coli and Bacillus cereus can cause spoilage in food; intake food contains plenty of bacteria and toxic. Therefore it is important to eliminate or control these bacteria safely. Ultraviolet (UV) radiation is considered as non-ionizing radiation and the first time in 1940 was used as a method for infection elimination in air. This approach nowadays is widely used for controlling microbial growth and as disinfection in food industry. Wavelength range of ultraviolet radiation is approximately 328-210 nm. The beam is naturally present in the sunlight. The bactericidal effect of UV irradiation depends on the type of bacteria, the distance, and dose of radiation. The most cytotoxic effect of UV irradiation is obtained at the wavelength of 260 nm, which corresponds to the intense absorption of energy by organic bases in the nucleic acid. UV irradiation causes radicals generation, which subsequently attack the nucleic acid and develop mutations in their genomes and gene transcription and translation processes. In this study, the antibacterial effect of different exposure times of ultraviolet radiation on the growth of E. coli and B. cereus was evaluated.Materials and methods: All media used in this study was procured from Merck Company. Ultraviolet device (Camag, USA) was used at wavelength of 254 nm, Nr= 29000, Amp= 0.25. B. cereus isolation: 1 mL of different dilutions of rice (0.1, 0.01, and 0.001) was transferred to Brain-heart infusion (BHI) and it was incubated at 32 °C for 24 h. A loop containing the bacteria was then transferred to Mannitol Egg Yolk Polymyxin (MYP) agar and it was incubated at 35 °C for 24 h. B. cereus produce big and round colonies, with a halo around the colonies. Starch test was carried out as confirming test for B. cereus colonies. Briefly, some colonies of B. cereus were added to test tube with sterile distilled water containing starch and a few drops of lugol. Development of blue color indicates the presence of B. cereus due to starch hydrolysis.E. coli isolation: E. coli was isolated from raw milk following the method described by Kargar et al. (2005). Briefly, raw milk was first homogenized; 0.1 ml of each dilution of homogenized raw milk was inoculated on Escherichia coli broth medium containing 20 mg novobiocin. E. coli was then isolated after transferring the former media on EMB specific culture and incubation at 36 °C for 24 h. After confirming colonies by Durham tube and complementary tests, pure cultures were obtained from them by streak-plate method.UV irradiation: A loop of E. coli colonies was transferred to nutrient broth and it was treated with UV beam (254 nm) at three times (40, 60, and 80 s). After preparing dilution of 0.0001 for each of the treatments and incubating for 24 h, survival curve was plotted. These operations were also carried out on B. ceruse colonies. A control sample also was considered for each examined bacterium.Results and Discussion: Rate of Bacillus cereus growth was reduced under UV radiation. As it is shown, Death curve of E. coli, E. coli count was decreased by increasing the time of UV radiation, so that count of this bacteria reached to about zero after UV radiation for 80 s. However, reduction of B. cereus count was less than E. coli count at same wavelength (254 nm) and time of irradiation. This revealed that B. cereus have more resistance to UV radiation compared to E. coli. These results were consistent to observation of Sharp (1940) who evaluate the effects of UV light on bacteria suspended in air and reported that required energy for air sterilization containing B. cereus is more than twice the energy is needed to eliminate E. coli. UV light more penetrates to cell wall of Gram-negative bacteria compared to Gram-positive bacteria due to having a small amount of peptidoglycan in the cell wall and caused mutations in regulating genes of transcription and translation. Conclusion: The efficiency of the two main processes of cell is reduced in the presence of UV irradiation and leads to growth reduction and death. The more resistance of B. cereus can be for several reasons, such as having a thicker cell wall compared to E. coli, and the capability to produce spore, and the capability to proofing mutations.
