A basic storage design was performed to study shelf life [10 (link)] in pasta developed throughout the determination of fatty acid profiles and chemical parameters (TBARS and acidity index). Fatty acid profiles were determined according to Bligh and Dyer, 1959 [29 (link)]. Firstly, each sample was homogenized with an Ultraturrax device (IKA-WERKE, T-25 basic) using different solvents such as chloroform, methanol, potassium chloride, and water. This mixture was centrifuged for 10 min at 4000 rpm, and the fat was extracted from the top, and afterwards, after incorporating BHT, butylated hydroxytoluene, as an antioxidant substance, solvents were evaporated with nitrogen gas. Afterwards, methylation was performed using 0.03 g of this previous fat. This fat was mixed with an intern pattern, C23:0, which does not caused interferences in the matrix. Then, 2 ml of hexane and 1 ml of potassium hydroxide saturated solution in methanol was added. The upper phase was extracted to measure. To analyze the fatty acid profile, a gas chromatograph (HP-6890 II Hewlett-Packard) was employed with a column SP-2380 (100 m × 0.25 mm × 0.20 µm). The temperature program was 140–165 °C at 3 °C/min during 10 min and 165–220 °C at 5 °C/min during 50 min. Fatty acid content was quantified as total area (%) of identified fatty acids.
Ultraturrax device
Manufactured by IKA Group
Sourced in Germany
The Ultraturrax device is a high-speed dispersing tool designed for the homogenization and emulsification of liquids, pastes, and suspensions. It operates at speeds up to 24,000 rpm, allowing for the effective mixing and dispersing of various materials.
Automatically generated - may contain errors
Lab products found in correlation
J221 m centrifuge, by Beckman Coulter (2 mentions)
Hp 6890 2, by Hewlett-Packard (1 mentions)
T25 basic, by IKA Group (1 mentions)
Dc protein assay, by Bio-Rad (1 mentions)
Ripa buffer, by Cell Signaling Technology (1 mentions)
R 300, by Büchi (1 mentions)
Whatman 4 filter paper, by Cytiva (1 mentions)
Bca method, by Merck Group (1 mentions)
6 protocols using ultraturrax device
1
Pasta Shelf Life Fatty Acid Analysis
2
Parvalbumin Extraction from Muscle
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The parvalbumin extraction
was performed following an extraction protocol reported by Carrera
et al.63 (link) Sarcoplasmic protein extraction
was carried out by homogenizing 5 g of white muscle in 10 mL of 10
mM Tris–HCl pH 7.2, supplemented with 5 mM PMFS, for 30 s in
an Ultra-Turrax device (IKA-Werke, Staufen, Germany). The sarcoplasmic
protein extracts were then centrifuged at 40,000 g for 20 min at 4 °C (J221-M centrifuge; Beckman, Palo Alto,
CA). Parvalbumins were purified by taking advantage of their thermostability,
heating the sarcoplasmic extracts at 70 °C for 5 min. After centrifugation
at 40,000 g for 20 min (J221-M centrifuge, Beckman,
Palo Alto, CA), supernatants composed mainly of parvalbumins were
quantified by the bicinchoninic acid (BCA) method (Sigma-Chemical
Co., USA).
was performed following an extraction protocol reported by Carrera
et al.63 (link) Sarcoplasmic protein extraction
was carried out by homogenizing 5 g of white muscle in 10 mL of 10
mM Tris–HCl pH 7.2, supplemented with 5 mM PMFS, for 30 s in
an Ultra-Turrax device (IKA-Werke, Staufen, Germany). The sarcoplasmic
protein extracts were then centrifuged at 40,000 g for 20 min at 4 °C (J221-M centrifuge; Beckman, Palo Alto,
CA). Parvalbumins were purified by taking advantage of their thermostability,
heating the sarcoplasmic extracts at 70 °C for 5 min. After centrifugation
at 40,000 g for 20 min (J221-M centrifuge, Beckman,
Palo Alto, CA), supernatants composed mainly of parvalbumins were
quantified by the bicinchoninic acid (BCA) method (Sigma-Chemical
Co., USA).
3
Fatty Acid Profiling in Enriched Pasta
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Fatty acids profile was determined according to Bligh and Dyer method (1959) [19 (link)], taking into account modifications made in the analysis protocol in agreement with the procedure described by Ainsa et al. (2021) [20 (link)] to get a better adjustment to assayed enriched pasta. Each sample was homogenized with different solvents (chloroform, methanol, potassium chloride and water) using an Ultraturrax device (IKA-WERKE, T-25 basic). Subsequently, it was centrifuged at 4000 rpm at 10 min, and the fat was extracted. Solvents were evaporated with BHT (butylated hydroxytoluene) as an antioxidant. Then, 2 mL of hexane and 1 mL of potassium hydroxide saturated were incorporated. Fatty acid profile was analyzed using a gas chromatograph (HP-6890II). Fatty acids were measured as the total area (%) of identified fatty acids.
4
Protein Extraction and Analysis in Colon Cells
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HCT116 cells were seeded in 6 cm2 Petri dishes at a density of 1 × 106 cells and allowed to adhere for 24 h. Cells were then incubated with DMW/WF (6% or 10% v/v) for 0, 3, 6, 12, and 24 h. At each endpoint, cells were washed twice with ice-cold 1 × PBS, collected by scraping and lysed in RIPA buffer (Cell Signaling Technology, USA). After 30 s of sonication, lysates were centrifuged at 14,000 g (4 °C for 15 min), the supernatant was collected, and protein concentration was determined using the Bio-Rad DC protein assay (Bio-Rad, Munich, Germany). Total protein lysates from colon tissues were extracted as previously described [38 (link)]. In brief, colon tissues were washed with chilled 1 × PBS to remove all blood contaminants. The colon tissues were then dissected into small pieces, lysed in RIPA buffer using an Ultra Turrax™ device (Ika-Werke GmbH, Staufen, Germany) and sonicated for 30 s. Finally, the lysates were centrifuged at 14,000 g (4 °C for 15 min), the supernatant was collected and the protein concentration was determined using the Bio-Rad DC protein assay (Bio-Rad).
5
Phytochemical Extraction from Tropical Fruits
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Fruits were purchased at the local market in Bogotá (Colombia). At least 10 fruits were combined for each of the three replicated samples (one per month). Samples were examined and cleaned by using distilled water. Pulp, peel and seed were manually separated wherever it is possible in order to submit each portion to extraction procedure. The freeze-dried vegetable material (2.5 g of clean seeds or 5 g of peel or 5 g of pulp or seed + pulp) were finely ground with an Ultraturrax device (IKA®-Werke GmbH & Co. KG, Königswinter, Germany) and undergone to cold extraction with absolute ethanol (40 mL) for 24 h at room temperature (20 °C). The top phase was filtered through Whatman filter paper #4 (9.0 cm diameter) and the residue was re-extracted with 30 mL of absolute ethanol. The filtered top phases were combined and dried in a rotary evaporator (Buchi R-300, Milan, Italy) with operating temperature of 35 °C. Extraction yield ranged from 7.2 to 36.5 g ethanolic extracts on 100 g dry product in Solanaceae family and from 2.2 to 49.1 g/100 g dry product in Passifloraceae family. Extracts were then stored at −20 °C until further analysis.
6
Extraction and Characterization of Fish Allergens
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Fifteen different fish species were used in this work (Table 1). These species were acquired from marketplaces in order to comprise the most consumed fish species in Europe containing the main species that cause fish allergy [24] . These species were genetically identified using the fishID Kit (Bionostra, Madrid, Spain).
Extraction of sarcoplasmic proteins was prepared as described [35] . Briefly, 0.5 g of fish white muscle were homogenized in 4 mL of 10 mM Tris-HCl buffer, pH 7.2, with 5 mM of PMSF, for 2 min using an Ultra-Turrax device (IKA Werke, Staufen, Germany). β-PRVBs were purified by taking advantage of their thermostability [7] . After centrifugation at 40000 g for 20 min (J221-M centrifuge, Beckman, Palo Alto, CA, USA), supernatants containing principally β-PRVBs were quantified by the BCA method (Sigma-Chemical Co., USA).
Samples were analyzed in triplicate.
Extraction of sarcoplasmic proteins was prepared as described [35] . Briefly, 0.5 g of fish white muscle were homogenized in 4 mL of 10 mM Tris-HCl buffer, pH 7.2, with 5 mM of PMSF, for 2 min using an Ultra-Turrax device (IKA Werke, Staufen, Germany). β-PRVBs were purified by taking advantage of their thermostability [7] . After centrifugation at 40000 g for 20 min (J221-M centrifuge, Beckman, Palo Alto, CA, USA), supernatants containing principally β-PRVBs were quantified by the BCA method (Sigma-Chemical Co., USA).
Samples were analyzed in triplicate.
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