Sahar Sabahi; Seyed Ali Mortazavi; Mohammad Reza Nassiri; Arash Ghazvini; Fakhri Shahidi
Abstract
Introduction: Immunoglobin Y (IgY) or egg yolk antibodies have captured attention as substitutes for antibiotics against the presence of disease-causing organisms (Thomsen et al., 2016; Li et al., 2015; Pereira et al., 2019). Compared to mammalian IgG, IgY has several advantages including high yield, ...
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Introduction: Immunoglobin Y (IgY) or egg yolk antibodies have captured attention as substitutes for antibiotics against the presence of disease-causing organisms (Thomsen et al., 2016; Li et al., 2015; Pereira et al., 2019). Compared to mammalian IgG, IgY has several advantages including high yield, cost-effectiveness, and convenience (Li et al., 2015). It has been reported that the use of IgY against Helicobacter pylori can be particularly useful for prevention and treatment of diseases (Najdi et al., 2016). The growth inhibitory effect of IgY antibody is one of its crucial features (Alfarouk et al., 2019), meaning that IgY antibody could be consumed daily with no health risks. Therefore, daily consumption of IgY requires a suitable host. The ice cream is the best choice for this proposed, because this product is maintained at -18 C and ice cream composition has positive effect on the function of IgY. Therefore, we develop a new product for the prevention and treatment of Helicobacter pylori. Material and Methods: The Mueller-Hinton media was used for H. pylori growth. Then, 109 colonies of bacterium were inactivated by formaldehyde methods. The 3 white leghorn chickens (Animal Science Department of Ferdowsi University of Mashhad) of 16 weeks old were immunized by H. pylori and Freund’s complete adjuvant with an equal volume. The reminder immunization was done two weeks later and using of Freund’s incomplete adjuvant for stage 2. One week after the chickens received their last injection, their eggs were collected on a daily basis for eight weeks, labeled and kept at 4 °C (Pauly et al., 2011). Extraction and purification of IgY antibody from egg yolk was done using polyethylene glycol 6000 (PEG 6000) powder (Merck, Germany) according to Polson’s method (Pauly et al., 2011). The IgY production wasverified by the SDS-PAGE and Bradford methods. The specific activity of IgY antibody against H. pylori was examined by ELISA. The absorbance of each well was measured at 492 nm (Nasiri et al., 2016). The egg yolk ice cream preparation contains IgY was prepared according to the method of Herald et al (2008). For estimation of IgY shelf life in ice cream, the residual activity of IgY after freezing was measured by ELISA. The Sensory evaluation of the egg yolk ice creams was conducted by 5-point hedonic scale through surveys to assess consumers’ level of acceptance. Results and Discussion: The purification of IgY using the SDS-PAGE method was verified correctly and the size of heavy and light chains of IgY was estimated about 65 and 27 kDa. The IgY yield in samples was 11.46 mg/ml, therefore, the high purity and yield of IgY, it could be effectively used as an ideal antibody in the food industry. The purification of IgY from egg yolk by PEG method is remarkably cost effective. The ELISA assay indicated binding of IgY to H. pylori antigen that was fixed on plates. Also, the results revealed significant differences between the control and treatment groups (P-value <0.001). The shelf life of IgY in ice cream showed that this product could be most certainly used as bio-food. ELISA analysis confirmed that IgY could remain effective in ice cream for 3 months. The activity of IgY during this period decreased about 6%, which is insignificant. The sensory features of ice cream using hedonic scale revealed that this product could be suitable for marketing, as no significant difference was found between egg yolk and vanilla ice creams. Ice cream is considered as the best candidate for production of IgY bio-foods, as it possesses the features needed to protect IgY and to increase efficiency of IgY activity. The optimum temperature for protection of ice cream was -18 °C that is utterly suitable for IgY protection. Furthermore, ice cream is consumed while cold and studies showed that IgY can remain active at temperatures of up to 80 °C and pasteurization process had no effect on IgY activity (Horie et al., 2004). Jaradat et al (2000) reported that, IgY activity was inhibited in presence of carbohydrates (sucrose, lactose and trehalose). Therefore, here, we used freeze dried egg yolk to protect IgY. Consequently, ice-cream is an ideal candidate for transferring antibodies into human body, as the particular composition of ice cream can properly protect IgY antibodies.
Elham Eshaghabadi; Farajollah Shahriari; Mohammadreza Nassiry; Mohammad Reza Edalatian Dovom; Amin Mirshamsi
Abstract
Introduction: Bread is considered as one of the most important commodity which has provided human nutritional needs for several centuries. In recent decades, bread consumption has witnessed tremendous increase due to rising of other foods expenses. Cereal and bread industries has been always concerned ...
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Introduction: Bread is considered as one of the most important commodity which has provided human nutritional needs for several centuries. In recent decades, bread consumption has witnessed tremendous increase due to rising of other foods expenses. Cereal and bread industries has been always concerned about bread production in an industrial scale along with suitable shelf life, acceptable organoleptic properties and free from any chemical additives and preservatives. Among different methods such as addition of synthetic enzymes, gums, emulsifiers and improvers, sour dough has gained special attention besides shelf life enhancement and improvement of sensorial and nutritional features. The main role of sour dough has been prominent for fermentation and gas production in dough as a result of inclusion of Lactic Acid Bacteria (LAB) as a technology. Fluctuation of process parameters including temperature, dough yield and starter culture composition determine the sour dough quality. The main reason of sour dough application in wheat bread is flavor enhancement. Culture-based microbiological examinations have accompanied with some difficulties in accurate and precise identification of Lactobacilli. So, in recent years, in order to identify precisely, different molecular and PCR assays and gene sequencing have been applied. Thus, in present study, fragment of 253bp from 16 S rRNA gene in Lactobacillus plantarum was amplified with the help of PCR and sequenced in the next step. Also, comparison of nucleotide sequence of this region of traditional Lb. plantarum with nucleotide sequence of industrial starter (Germany) and sequences deposited in Gene Bank was evaluated.
Material and Methods: Sampling: According to sensory properties, sour dough samples were collected from Torbat e jam city and industrial starter sample (sour dough) was prepared from Buker Company, Germany.
Isolation and bio chemical analysis: Ten- fold serial dilutions of samples were prepared and cultured on MRS agar. Gram positive, catalase negative isolates were considered as presumptive LAB.
Molecular Identification: DNA of isolates were extracted according to the procedure of DNA extraction kit (Thermo, K721, USA). In order to amplify the 253 bp fragment of 16S rRNA in Lb. plantarum, specific primers were designed using Primer Premier 5. PCR was performed in Biometra Thermocycler (T- personal, Germany) in 36 cycles. In the following step, PCR products were sent to Macrogen Company for sequencing.
Bioinformatics analysis: Analysis of sequencing results was conducted with the aid of bioinformatics soft wares including CLC Main work bench, Chromas Lite 2.01, MEGA 5 and BioEdit. Phylogenetic tree was designed among different isolates using Neighbor-Joining method (bootstrap=1000) in MEGA 5 software. For determination of genetic distance, Create Pairwise comparison method was used in CLC Main workbench.
Results and Discussion: Quality and quantity of extracted DNA samples were confirmed by Gel electrophoresis. PCR and electrophoresis of PCR products confirmed and proved the accuracy and validity of amplified 253 bp fragment with intended primers. Comparison of Torbat and Germany nucleotide sequences demonstrated that two replacement mutations have been occurred including (A/C) and (T/C) in positions 182 and 205, respectively. These changes resulted in replacement of Histidine by Proline, and Glutamine by stop codon. Nucleotide content of 16S rRNA gene sequence in Lb. plantarum isolated from Torbat e Jam and Germany sour dough samples displayed both sequences contained 49.4% (G+C) and 50.6% (A+T). Phylogenetic results demonstrated that Lb. plantarum isolated from Torbat e Jam and Germany samples were classified in the same sub group. This implies the close phylogenetic relationship between these two lactobacilli. In order to identify Lb. plantarum, 16S rRNA gene was sequenced in sour dough samples of Germany and Belgium and phylogenetic tree showed that Lb. plantarum had the lowest genetic distance with Lb. pontis and highest one with Lb. sanfranciscensis and Lb. salivarius
Mahboubeh Sarabi; Fakhri Shahidi; Ahmad Reza Bahrami; Seyed Ali Mortazavi; Mohammadreza Nassiry; Massoumeh Mehraban Sangatash; Marzieh Hosseininejad
Abstract
Raisins are dried grapes that may be eaten raw or used in cooking, baking and brewing as an ingredient. Iran is one of the major exporters of raisins in recent years and Iranian raisins are exported to many countries. It is important to ensure of microbiological quality and safety of raisins. The aim ...
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Raisins are dried grapes that may be eaten raw or used in cooking, baking and brewing as an ingredient. Iran is one of the major exporters of raisins in recent years and Iranian raisins are exported to many countries. It is important to ensure of microbiological quality and safety of raisins. The aim of this study was isolation and identification of mycoflora from main raisin’s varieties (Poloie, Pikami & Teiphi) produced in Khorasan Razavi Province. Four types of culture medium (Yeast Extract Glucose Chloramphenicol Agar (YGC), Potato Dextrose Agar (PDA), Czapek Dox Agar (CA) and Dichloran Glycerol Agar (DG18)) were used for isolation. The highest contamination was found in Teiphi raisin samples. The drying method was not effect on fungi contamination. For Fungi Identification, Macroscopic and Microscopic features were investigated. The Result showed that frequently isolated fungi were Aspergillus and Penicillium species. The fragment (600 bp) of Internal Transcribed Spacer 5.8SrRNA of isolated colonies was amplified using polymerase chain reaction for identification down to the species level. According to the results, the raisins had varius species of Aspergillus, Penicillium, Rhizomucor, Mucor, Alternaria and Cladosporium. Aspergillus niger was the most abundant fungi.
