Alkaloid extraction was performed as described by Goossens et al. [65 (link)]. Briefly, five-week-old wild-type and transgenic plantlets were sprayed with 0.1% DMSO (control) or 100 µM MeJA. Leaves were then collected and lyophilized 48 h after the treatment. 50 mg lyophilized tissue were homogenized in liquid nitrogen and basified with 10% NH4OH. 100 µg quinaldine was added as an internal standard. Total alkaloids were extracted with CH2Cl2, vacuum concentrated, and resuspended into 400 µL CH2Cl2. For the TLC assay, alkaloid extracts from three individuals of each line were mixed together and then equal amounts of the extracts from different lines were loaded onto a silica gel TLC plate (UV254, Analtech). Separation was done with the mobile phase composed of dichloromethane: methanol: 10% NH4OH (125:15:2). Spots were visualized by the spray with Dragendorff reagent (Sigma-Aldrich, St. Louis, MO, USA).
Dragendorff reagent
Manufactured by Merck Group
Sourced in United States, China, Germany
Dragendorff reagent is a chemical solution used in analytical chemistry for the detection and identification of certain organic compounds, particularly alkaloids. It is a mix of bismuth subnitrate, potassium iodide, and acetic acid in water.
Automatically generated - may contain errors
Lab products found in correlation
Chloroform, by Merck Group (3 mentions)
Hydrochloric acid (hcl), by Merck Group (3 mentions)
Lead acetate, by Merck Group (2 mentions)
Acetic anhydride, by Merck Group (2 mentions)
Mayer reagent, by Merck Group (2 mentions)
Folin ciocalteu reagent, by Merck Group (2 mentions)
Uv 3600, by Shimadzu (1 mentions)
Sulphuric acid, by Merck Group (1 mentions)
Galactose, by Thermo Fisher Scientific (1 mentions)
Ammoniacal alcohol, by Merck Group (1 mentions)
Ferric chloride, by Merck Group (1 mentions)
Mayer s reagent, by Merck Group (1 mentions)
Sodium hydroxide (naoh), by Merck Group (1 mentions)
Tropicamide ophthalmic solution, by Alcon (1 mentions)
Blank disc, by Thermo Fisher Scientific (1 mentions)
Nano2, by Merck Group (1 mentions)
Acetic acid anhydride, by Merck Group (1 mentions)
Alcl3, by Merck Group (1 mentions)
Wagner reagent, by Merck Group (1 mentions)
Crystal violet, by Merck Group (1 mentions)
8 protocols using dragendorff reagent
1
Alkaloid Extraction and TLC Analysis
2
Nicotine Vaccine Development and Characterization
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The vaccine was prepared by the conjugation of 3′-aminomethylnicotine with the AFPL1 component in the presence of peptides [37] . The peptide structure and the proportions of nicotine hapten, short peptide and adjuvant in addition to the coupling technique are undisclosed as they are being submitted for patent review. AFPL1 from N. meningitidis serogroup B (meningococcal strain Cu385 (B4:P1.19,15)) was manufactured under Good Manufacturing Practice conditions and provided by the manufacturing plant of Finlay Vaccine Institute, Cuba. Nicotine was also quantified by using UV adsorption of derived nicotine as a standard at 265 nm on UV-3600 (Shimadzu) and by TLC on Silica gel 60 F254, EM Science with Dragendorff reagent (Sigma, Mississauga, ON) for detection. Particle size as a parameter for stability analysis was analyzed on Malvern Zetasizer ZS. The nicotine vaccine was stored at room temperature for one month and then stored at 4 °C before in vitro testing. We performed in vivo tests using only the vaccine stored at 4 °C.
3
Evaluation of Antioxidant Potential in Rat Eyes
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Galactose was purchased from ACROS, New Jersey, USA; tropicamide ophthalmic solution (1%) was purchased from Alcon Laboratories South Africa (Pty) Ltd; rat enzyme-linked Immunosorbent Assay (ELISA) kits (Glutathione (GSH), Bicinchoninic acid (BCA), alpha-crystallin A chain (CRYAA) and aquaporin 0 (AQPO)) were purchased from Shanghai chemical limited, Shanghai, China; sodium selenite, ammoniacal alcohol, sulphuric acid, chloroform, dragendorff reagent, Mayer's reagent, sodium hydroxide, Fehling's solution A & B, lead acetate, ferric chloride, ammonia (liquid), hydrochloric acid, acetic anhydride, and 2,2-dipheny-1-picrylhydrazyl (DPPH) were purchased from Sigma Aldrich, Germany; quercetin was purchased from Double Wood LLC, USA; gallic acid was purchased from Reg-LABOGENS, India; and ascorbic acid was purchased from Brenntag, Ghana.
4
Antimicrobial Potential of Plant Extracts
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Nutrient Agar (NA), Sabouroud Dextrose Agar (SDA), Aquadest (Brataco), 70% Ethanol (Brataco), FeCl3 (Merck), Wagner reagent, Mayer reagent, Dragendorff reagent, Ammoniak (Merck), Acetic acid anhydride (Merck), NaNO2 (Merck), AlCl3 (Merck), HCl (Merck), Chloroform (Merck), H2SO 4 (Merck), DMSO, immersion oil, Crystal violet (Merck), Safranin, Lugol's iodine, 0.9% NaCl, Blank disc (Oxoid), the antibiotic disk of Nystatin and Amoxicillin, analytical balance (Excellent), oven (Memmert), blender (Phillips), aluminium foil (Klin Pak), autoclave, incubator, vacuum rotary evaporator, Hot plate, and Laminar Air Flow.
5
Phytochemical Screening of Herbal Medicines
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All chemicals used were of analytical grade, and they were used without further purification. All solutions were made using distilled water. Allopurinol was obtained from Nanjing Pharma Chemical Plant. 2-Naphthol, ammonium hydroxide 25%, hydrochloric acid 37%, bismuth subnitrate, ethanol 95%, ferric chloride, potassium hydroxide, potassium iodide, sodium hydroxide, potassium chlorate crystals, sodium nitrite, sodium nitroprusside, sodium sulfite anhydrous, silver nitrate, Dragendorff reagent, p-DAB, and Folin-Ciocalteu reagent were purchased from Merck. Alkaline fuchsin was purchased from Sigma Aldrich, and sample packaged herbs were purchased from traditional drug store around Jatinangor, Sumedang, West Java. Whatman No. 1, No. 4, and No. 6 and Whatman 1 chromatography qualitative-grade filter paper were purchased from GE Healthcare. The absorbance measurement was recorded by UV-Visible spectrophotometer Analytik Jena SPECORD 200 using a 1.0 cm quartz cell.
6
Phytochemical Screening of Ficus moluccana
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F. moluccana twig were obtained from sengon plantation in Cisempur Village, Jatinangor Sub-District, Sumedang Regency, West Java, Indonesia. All chemical reagents for phytochemical screening i.e. methanol, folin-ciocalteau, phenol, ammonia, toluene, chloroform, hydrochloric acid, dragendorff reagent, mayer reagent were purchased from Merck. Gallic acid, quercetin, 2,2-diphenyl-1-picrylhydrazyl (DPPH) were purchased from Sigma-Aldrich (MO, USA).
7
Phytochemical Extraction and Analysis
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Aluminum chloride, ascorbic acid, chloroform, ethanol, ethyl acetate, ferric chloride, ferrous sulfate, gallic acid, glacial acetic acid, hexane, hydrogen peroxide, α-naphthol, petroleum ether, potassium dihydrogen phosphate, potassium hydrogen phosphate, potassium chloride, potassium ferricyanide, sodium bicarbonate, trichloroacetic acid (TCA), and vanillin were procured from SD Fine Chemicals Limited, Mumbai, India. Acetone, hydrochloric acid, methanol, sodium hydroxide, and sulfuric acid were acquired from Spectrum Chemical Private Limited, Cochin. Acetic anhydride, Dragendorff reagent, Folin–Ciocalteu reagent, lead acetate, Mayer's reagent Sodium nitrate, and Wagner's reagent were purchased from MERCK Specialties Private Limited, Mumbai. 2,2-diphenyl-1-picrylhydrazyl (DPPH) was procured from SISCO Research Laboratories, Maharashtra, India. Tris base was from NR Chemicals Private Limited, Mumbai, India. Thiobarbituric acid (TBA) was bought from Loba Chemie Private Limited, Mumbai, India. All chemicals and reagents were of analytical grade.
8
Alkaloid Analysis by TLC and HPLC-MS
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Fractions from CCC separation were analyzed by TLC. Plates were cut in half (10 × 20 cm) and carefully spotted in evenly spaced dashes using Camag Autosampler System (Muttenz, Switzerland). Samples were applied to the plates as 5 mm spots or bands, 10 mm apart, 15 mm from the left and lower edges. The development was performed in Camag unsaturated glass TLC chambers, using the solvent system methanol–dichloromethane (10:90) (v/v) with the addition of 5 % ammonia, in 64 % humidity. Dried TLC were examined under UV light at 254 and 365 nm and derivatized with Dragendorff reagent (Merck, Darmstadt, Germany) until vivid orange spots of alkaloids became visible. Similar fractions were mixed and evaporated to dryness. The combined fractions were developed on HP-TLC plates in the same solvent system. Four were prepared simultaneously to be sprayed with Dragendorff and vanillin reagents, to be analyzed by TLC–HPLC–MS, and to be subjected to the anticholinesterase test.
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