Allium cepa

At baseline, registered dietitians completed a 14-item Mediterranean Diet adherence screener (Table 1) in a face-to-face interview with the participant [21] (link), [27] –[29] (link). The dietitians had been previously trained and certified to implement the PREDIMED protocol and had been hired to work full-time for the trial. The 14-item tool was developed in a Spanish case-control study of myocardial infarction [30] (link), where the best cut-off points for discriminating between cases and controls were selected for each food or food group. With this first step, 9 of the 14 items were obtained [31] (link). Five additional items that were felt to be especially relevant to assess adherence to the traditional Mediterranean diet were subsequently added. Two of these items used short questions to inquire on food habits: Do you use olive oil as the principal source of fat for cooking? and Do you prefer to eat chicken, turkey or rabbit instead of beef, pork, hamburgers or sausages? The other 3 items inquired on frequency of consumption of nuts, soda drinks and a typical Mediterranean sauce (“sofrito”): How many times do you consume nuts per week? How many carbonated and/or sugar-sweetened beverages do you consume per day? How many times per week do you consume boiled vegetables, pasta, rice, or other dishes with a sauce (“sofrito”) of tomato, garlic, onion, or leeks sauteed in olive oil?[26] (link).
The baseline 14-item questionnaire (Table 1) was the primary measure used in this study to appraise adherence of participants to the Mediterranean diet. In addition, a full-length 137-item validated FFQ [32] (link) was also administered to all participants. We obtained information about total energy intake and alcohol intake (only with descriptive purposes) from this FFQ. In the validation study, the score obtained with brief 14-item questionnaire correlated significantly with that obtained from the full-length FFQ score (Pearson correlation coefficient (r) = 0.52; intraclass correlation coefficient = 0.51). Associations in the anticipated directions for the different dietary intakes reported on the FFQ were found [26] (link). Significant inverse correlations of the 14-item tool with fasting glucose, total:HDL cholesterol ratio, triglycerides and the 10-y estimated coronary artery disease risk also supported the validity of this brief Mediterranean diet adherence screener [26] (link).
Also a general medical questionnaire, and the validated Spanish version of the Minnesota Leisure-Time Physical Activity Questionnaire [33] (link)–[34] (link) were collected by the dietitians in the personal interview with each participant [21] (link). Weight, height and WC were directly measured by registered nurses who had been previously trained and certified to implement the PREDIMED protocol and were hired to work full-time for this trial, as previously described [21] (link), [27] –[29] (link). The WHtR was calculated as WC divided by height, both in centimeters.

The vectors, pCM1205-RFP, pLPGM113, pLPGM202 and pLPGM413, were transformed into A. tumefaciens strain GV3101 for further transient transformation in onion epidermis. Positive Agrobacterium harboring pCM1205-RFP was selected and cultivated in YEB media supplemented with 100 mg/L rifampicin and 25 mg/L chloramphenicol. Positive Agrobacterium harboring pLPGM113, pLPGM202 and pLPGM413 were selected and cultivated in YEB media supplemented with 100 mg/L rifampicin and 100 mg/L Kan. Positive Agrobacterium cultivated overnight at 28°C were harvested at OD₆₀₀ of 1.5 to 2.0, centrifuged at 5000 rpm for 10 min and re-suspended in 50 ml of infiltration liquid, and the centrifugation and resuspension procedure was repeated three to five times. Finally, Agrobacterium cells were diluted in agroinfiltration liquid to appropriate concentration for agroinfiltration. Different agroinfiltration durations (24 h, 48 h, 72 h and 96 h) and Agrobacterium concentrations (OD₆₀₀ 0.05, 0.10 and 0.15) were evaluated to determine conditions to obtain high transformation efficiency (Table 1).
The complete infiltration liquid was made as following: 41.65 mM D-glucose, 100 mM CaCl₂, 100 mM MES-KOH (pH 5.6) stock solution, 0.011 µM BAP, 0.01% Silwet L-77, 0.05 mM MgCl₂ and 12.5 mM AS (made with DMF, dimethylformamide) stock solution, and suitable amount of ddH₂O to make final volume to 20 ml. About 200 µl infiltration liquid with Agrobacterium carrying constructed vectors were injected into the interface of adaxial epidermis and mesophyll of onion scales to make a bubble for agroinfiltration. In order to investigate whether all the components are necessary, different infiltration liquid without one of the components were used to evaluated the effect of different components of the infiltration liquid (Table 2).

Saussurea costus peel specimens were bought from a local market in Riyadh, Saudi Arabia (Fig 1) [28 ]. The dry S. costus root peel was fermented and dried with water. A milling machine was used to grind dehydrated specimens into powder form. Extracts were produced from 20 g of plant powder in 100 ml of 50% methanol and stirred with a magnetic stirrer until a homogeneous solution was formed. To prevent enzyme activity, the aqueous extract was heated for 30 min at 70°C in a water bath. Furthermore, the extract was filtered over the Whatman mentioned above. In a simple method, iron oxide NPs were formed by adding 100 ml of onion peel extract into (50 mL of 1 mM FeCl₂+ 100 mL of 1 mM FeCl₃). To concentrate the extract, 5 ml of the prepared peel extract was blended with 45 ml (2: 1, FeCl₂: FeCl₃) solution in a flask and left at room temperature for 24 hours. Then, a magnetic stirrer was used to agitate the solution for 30 minutes at 80°C. The first indicator of γ-Fe₂O₃ NPs production as a result of reducing iron ions to iron nanoparticles is the exchange of the binder color from yellow to dark brown. The use was kept at 4°C for further IONP description and apps [29 (link)].