Nauclea diderrichii stem bark was harvested at Mbanga (Moungo Division, Littoral region of Cameroon) in March 2019 and authentified at Cameroon National Herbarium (CNH) by comparison with the voucher specimen registered under the reference number 6608/HNC. Nauclea diderrichii stem bark was sliced, dried at room temperature (24-25°C), and ground to a fine powder which was used for aqueous and ethanolic extractions. The aqueous extract was prepared by boiling 500 g of powder in 5000 ml of distilled water for 20 minutes. The resulting mixture was filtered using Whatman paper no. 4. After filtration, the filtrate was dried in an oven at 40°C for 2 days. This process allowed 44.37 g of aqueous extract (AEND) for an extraction yield of 8.874%. Concerning ethanolic extract, 500 g of the same powder was macerated in 5000 ml of ethanol (96%) for 48 hours. The resulting mixture was filtered using Whatman paper no. 4, and filtrate was evaporated using a rotavapor under reduced pressure (79°C), giving 24.35 g of ethanolic extract (EEND) for an extraction yield of 4.87%.
Whatman no 4 paper
Manufactured by Cytiva
Sourced in Germany, Switzerland, United States
Whatman No. 4 paper is a medium-grade qualitative filter paper suitable for general filtration applications. It has a medium speed and retains medium-sized particles. The paper is made from high-quality cotton linters and is suitable for a variety of laboratory filtration tasks.
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Lab products found in correlation
Freezone 4, by Labconco (8 mentions)
Whatman no 4 filter paper, by Cytiva (4 mentions)
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R 210, by Büchi (3 mentions)
Model rw20 n, by IKA Group (2 mentions)
R 215, by Büchi (2 mentions)
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Sphereclone, by Phenomenex (2 mentions)
20a series uflc, by Shimadzu (2 mentions)
Uv 1601 pc spectrophotometer, by Shimadzu (1 mentions)
Whatman no 1 paper, by Cytiva (1 mentions)
Delta 2 24 lsc plus, by Martin Christ (1 mentions)
Rotavapor r 300, by Büchi (1 mentions)
Rotary evaporation, by Büchi (1 mentions)
Rotary evaporator, by Büchi (1 mentions)
N hexane, by Merck Group (1 mentions)
Ethyl acetate, by Merck Group (1 mentions)
Freezone 4.5 model 7750031, by Labconco (1 mentions)
R 210 rotary evaporator, by Büchi (1 mentions)
C18 sphereclone, by Phenomenex (1 mentions)
101 protocols using whatman no 4 paper
1
Nauclea diderrichii Stem Bark Extraction
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Nauclea diderrichii stem bark was harvested at Mbanga (Moungo Division, Littoral region of Cameroon) in March 2019 and authentified at Cameroon National Herbarium (CNH) by comparison with the voucher specimen registered under the reference number 6608/HNC. Nauclea diderrichii stem bark was sliced, dried at room temperature (24-25°C), and ground to a fine powder which was used for aqueous and ethanolic extractions. The aqueous extract was prepared by boiling 500 g of powder in 5000 ml of distilled water for 20 minutes. The resulting mixture was filtered using Whatman paper no. 4. After filtration, the filtrate was dried in an oven at 40°C for 2 days. This process allowed 44.37 g of aqueous extract (AEND) for an extraction yield of 8.874%. Concerning ethanolic extract, 500 g of the same powder was macerated in 5000 ml of ethanol (96%) for 48 hours. The resulting mixture was filtered using Whatman paper no. 4, and filtrate was evaporated using a rotavapor under reduced pressure (79°C), giving 24.35 g of ethanolic extract (EEND) for an extraction yield of 4.87%.
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Nauclea diderrichii stem bark was harvested at Mbanga (Moungo Division, Littoral region of Cameroon) in March 2019 and authentified at Cameroon National Herbarium (CNH) by comparison with the voucher specimen registered under the reference number 6608/HNC. Nauclea diderrichii stem bark was sliced, dried at room temperature (24-25°C), and ground to a fine powder which was used for aqueous and ethanolic extractions. The aqueous extract was prepared by boiling 500 g of powder in 5000 ml of distilled water for 20 minutes. The resulting mixture was filtered using Whatman paper no. 4. After filtration, the filtrate was dried in an oven at 40°C for 2 days. This process allowed 44.37 g of aqueous extract (AEND) for an extraction yield of 8.874%. Concerning ethanolic extract, 500 g of the same powder was macerated in 5000 ml of ethanol (96%) for 48 hours. The resulting mixture was filtered using Whatman paper no. 4, and filtrate was evaporated using a rotavapor under reduced pressure (79°C), giving 24.35 g of ethanolic extract (EEND) for an extraction yield of 4.87%.
2
Preparation of Plant Infusions and Extracts
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C. officinalis (air-dried flowers) and M. cervina (air-dried leaves) samples were purchased from two companies, Soria Natural® from Soria, Spain, and Cantinho das Aromáticas® from Vila Nova de Gaia, Portugal, respectively. Both companies have their own organically grown crops. Each sample was reduced to a fine dried powder (20 mesh) and stored in a desiccator, protected from light, until further analysis.
To prepare the infusions, each sample (1 g) was added to 200 mL of boiled distilled water and kept for resting at room temperature for 5 min followed by subsequent filtration through a Whatman No. 4 paper.
For hydromethanolic extract preparation, each sample (1 g) was extracted by stirring in 30 mL of methanol/water (80 : 20 v/v, at 25 °C at 150 rpm) for 1 h and subsequently filtered through a Whatman paper No. 4. The residue was then extracted with an additional portion of 30 mL of the hydro-methanolic mixture. The combined extracts were evaporated under reduced pressure (rotary evaporator Büchi R-210, Flawil, Switzerland) until the complete removal of methanol, and afterwards the aqueous phase was frozen and lyophilized (FeeeZone 4.5, Labconco, Kansas City, MO, USA).
To prepare the infusions, each sample (1 g) was added to 200 mL of boiled distilled water and kept for resting at room temperature for 5 min followed by subsequent filtration through a Whatman No. 4 paper.
For hydromethanolic extract preparation, each sample (1 g) was extracted by stirring in 30 mL of methanol/water (80 : 20 v/v, at 25 °C at 150 rpm) for 1 h and subsequently filtered through a Whatman paper No. 4. The residue was then extracted with an additional portion of 30 mL of the hydro-methanolic mixture. The combined extracts were evaporated under reduced pressure (rotary evaporator Büchi R-210, Flawil, Switzerland) until the complete removal of methanol, and afterwards the aqueous phase was frozen and lyophilized (FeeeZone 4.5, Labconco, Kansas City, MO, USA).
3
Extraction and Quantification of Phenolic Acids
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Each sample (~1.5 g) was extracted with methanol:water (80:20, v/v; 30 mL) at −20 °C for 6 h. After sonication for 15 min and filtered through Whatman No. 4 paper, the residue was then extracted with two additional 30 mL portions of the methanol:water mixture, and combined extracts were evaporated at 40 °C under reduced pressure to remove methanol. The aqueous phase was submitted to a liquid–liquid extraction with diethyl ether (3 × 30 mL) and ethyl acetate (3 × 30 mL). Anhydrous sodium sulphate was added to the combined organic phases and after were filtrated through Whatman No. 4 paper, evaporated to dryness and re-dissolved in a mixture of water:methanol (80:20, v/v; 1 mL). The obtained extracts were filtered through a 0.22 μm disposable LC filter disk for HPLC analysis. Phenolic acids and related compounds were determined in the UFLC system mentioned above, as previously described by the author [25 (link)]. DAD detection was carried out using 280 as preferred wavelengths. The phenolic acids and related compounds were quantified by comparison of the area of their peaks with calibration curves obtained from commercial standards of each compound. The results were expressed in μg per g dw.
4
Carob Seed Extraction: Maceration and Ultrasound
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Carob seeds were extracted by maceration (ME) and ultrasoundassisted extraction (UAE). For both extraction methods, water and ethanol were chosen as solvents, with four different proportions being used: i) ethanol:water (25:75; v/v); ii) ethanol:water (50:50; v/v); iii) ethanol:water (75:25; v/v); and iv) 100% water.
For ME the dried powdered samples (1 g) were placed in a beaker with 30 mL of each of the four solvents, under magnetic stirring (150 rpm) for 1 h at room temperature. After this period, the extracts were filtered (Whatman No 4 paper) and the extraction procedure was repeated with an additional portion of the solvent. The obtained extracts were combined, the ethanol was removed (rotary evaporator Büchi R-210, Flawil, Switzerland) and the residual aqueous phase was frozen and lyophilized (freeze 4.5 FreeZone model 7750031, Labconco, Kansas City, MO, USA).
The UAE was carried out in an ultrasonic device (QSonica sonicators, model CL-334, Newtown, CT, USA), based on a methodology previously optimized by Rached et al. (2016) (link). The dried powdered samples (3 g) were extracted with 100 mL of each of the four solvents by the ultrasonic device at 375 W for 10 min. The extracts obtained were filtered (Whatman No 4 paper) and, as for the ME, the ethanol was removed, and the residual aqueous phase was frozen and lyophilized.
For ME the dried powdered samples (1 g) were placed in a beaker with 30 mL of each of the four solvents, under magnetic stirring (150 rpm) for 1 h at room temperature. After this period, the extracts were filtered (Whatman No 4 paper) and the extraction procedure was repeated with an additional portion of the solvent. The obtained extracts were combined, the ethanol was removed (rotary evaporator Büchi R-210, Flawil, Switzerland) and the residual aqueous phase was frozen and lyophilized (freeze 4.5 FreeZone model 7750031, Labconco, Kansas City, MO, USA).
The UAE was carried out in an ultrasonic device (QSonica sonicators, model CL-334, Newtown, CT, USA), based on a methodology previously optimized by Rached et al. (2016) (link). The dried powdered samples (3 g) were extracted with 100 mL of each of the four solvents by the ultrasonic device at 375 W for 10 min. The extracts obtained were filtered (Whatman No 4 paper) and, as for the ME, the ethanol was removed, and the residual aqueous phase was frozen and lyophilized.
5
Extraction and Characterization of Herbal Infusions and Decoctions
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To prepare infusions, each sample (5 g) was added to 25 mL of boiling distilled water and left to stand at room temperature for 5 min, and then filtered through Whatman No. 4 paper. To prepare decoctions, each sample (5 g) was added to 25 mL of distilled water, heated on a heating plate (VELP Scientific, Usmate, Italy) and boiled for 30 min. The mixture was left to stand at room temperature for 5 min more, and then filtered through Whatman No. 4 paper, and the extracts were kept at -80 °C for 1 night and lyophilized (Free Zone 4.5, Labconco, Kansas City, MO, USA) for a minimum of 4 h. The extracts taken into bottles were kept in the refrigerator until the activity study was performed. The biological activities were evaluated directly on the decoctions/infusions [21] .
To prepare essential oils, approximately 30 g dried herbs, leaves and seeds were used for the essential oil isolation in dill. Dried herbs, leaves and seeds were put into balloon using a clevenger apparatus with the TS8882 method throughout 3 h. Anhydroussodium sulphate was used to obtained dry essential oil isolation and was kept at 4 °C until use. The yields of essential oil were calculated after dried weight of each sample.
To prepare essential oils, approximately 30 g dried herbs, leaves and seeds were used for the essential oil isolation in dill. Dried herbs, leaves and seeds were put into balloon using a clevenger apparatus with the TS8882 method throughout 3 h. Anhydroussodium sulphate was used to obtained dry essential oil isolation and was kept at 4 °C until use. The yields of essential oil were calculated after dried weight of each sample.
6
Extraction and Purification of Asian Seabass Liver Lipase
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Asian seabass liver lipase (ASL-L) was extracted using an extraction buffer (25 mM Tris–HCl, pH 8.0 containing 1 mM CaCl2) [14 (link)]. Firstly, LP was dispersed in the extraction buffer at a ratio of 1:9 (w/v) and thoroughly stirred for 40 min at 4 °C with an overhead stirrer (Model RW20.n, IKA-Werke Gmb H&CO. KG, Staufen, Germany) at 300 rpm. Afterward, the samples were centrifuged at 10,000× g and 4 °C for 30 min. The supernatant was filtered with Whatman paper No. 4 to remove the remaining fat debris. The filtrate or ASL-L was kept in ice for further experiments.
7
Aflatoxin Extraction from YES Culture
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Extraction of aflatoxins produced in the YES culture was carried out [17 (link)] by harvesting the mycelium of each flask through filtration in Whatman paper (No. 4) then extraction by 100 ml chloroform. The chloroform extract was then dried by adding anhydrous sodium sulfate and evaporated to dryness under nitrogen at temperature below 60ºC. The net dry weight of mycelia was determined. The dry film was then used for the detection of aflatoxins.
8
DPPH Radical Scavenging Assay Protocol
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The DPPH radical scavenging capacity was determined using the method of the Lee et al. (27 (link)) with slight modifications. The soyasaponin, ascorbic acid, α-tocopherol, or BHT was standardized to give a stock solution (25 mg/mL) and filtered through a 20 μm Whatman paper no 4. Aliquots (25 μL) were placed in a cuvette, and an ethanolic solution of DPPH (100 μM) was added to a final volume of 1 mL. The decrease in absorbance at 515 nm was determined continuously with data capturing at 30 s intervals using a UV-1601 PC spectrophotometer (Shimadzu Corporation, Kyoto, Japan). The degree of DPPH radical scavenging activity of the antioxidants was calculated as percentage of inhibition (% inhibition) using the following equation:
where Abscontrol is the absorbance at 0 min and Abssample is the absorbance of the sample at 5 min. An EC50 value was determined as the concentration that elicited a half-maximal response.
where Abscontrol is the absorbance at 0 min and Abssample is the absorbance of the sample at 5 min. An EC50 value was determined as the concentration that elicited a half-maximal response.
9
Extraction of Bioactive Compounds from Medicinal Plants
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Fresh A. indica leafs and pulverized M. oleifera leafs were obtained from a naturist shop. Both were free of impurities and had a sanitary registry. These products were placed in different containers and were treated with absolute methanol (1 : 2 weight/volume) to be later stored at room temperature for 10 days without sunlight exposure. The solutions were filtered through Whatman paper no. 4 and placed in labeled tubes. A rotatory evaporator was then used at 50°C to separate the methanol by distillation to finally obtain a pure extract. The extracts were stored at 4°C until use.
10
Aflatoxin B1 Inhibition by Biosynthesized AgNPs
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A hundred ml of YES medium were put in a 500 ml flasks and then autoclaved at 120°C for 15 min. Inoculation was carried out by adding 1 ml of a suspension of spores (105 spores) of a toxigenic A. flavus ATCC28542 strains without AgNPs (control) or with 0.5, 1.0 and 1.5 mg/100ml YES medium of one of the tested AgNPs (6a, 6b, 6c AgNps HA1N were synthesized by Aspergillus terreus (KR364880),3a, 3b, 3C AgNps EH were synthesized by Egyptian honey, and 1a, 1b, 1C AgNps HA2N were synthesized by Penicillium expansum (KR269857). The flasks were incubated in the dark for 14 days at 28°C. After the incubation period, extraction of AFB1 from in the YES culture according to the method of Munimbazi and Bullerman [23 ]. Where, the mycelium of each flask contained YES medium was harvested by filtration through Whatman paper (No. 4), then extracted with 100 ml chloroform. The chloroform extract was dried by addition of anhydrous sodium sulfate. The residue was transferred to a vial and evaporated off using a stream of nitrogen at a temperature below 60oC. The dry film was used for the detection and determination of AFB1 by (HPLC) according to (Deabes et al., [24 ,6 ] the retention time of AFB1 standard separation is 4.061. The percentage of inhibition of AFB1 is calculated using equation: % inhibition = (control- treatment /control) X100
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