Understanding tests for flavonoids is essential for B.Pharm students focused on phytochemistry and natural product analysis. This concise overview explains qualitative and confirmatory tests for flavonoids, covering the Shinoda test (Mg/HCl), ferric chloride reaction, alkaline reagent test, AlCl3 and lead acetate responses, hydrolysis of flavonoid glycosides, and UV-visible spectral shift reagents (NaOMe, AlCl3). Emphasis is placed on characteristic color changes, differentiation of flavone, flavonol, flavanone and anthocyanin classes, sample preparation, chromatographic and spectroscopic confirmation, and common interferences. Practical laboratory interpretation and safety considerations are highlighted for reliable identification. Now let’s test your knowledge with 30 MCQs on this topic.
Q1. Which reagent combination is used in the Shinoda test for flavonoids?
- Magnesium and hydrochloric acid
- Ferric chloride and water
- Aluminum chloride and sodium hydroxide
- Lead acetate and ethanol
Correct Answer: Magnesium and hydrochloric acid
Q2. What characteristic color change indicates a positive Shinoda test for many flavonoids?
- Deep blue
- Red or pink
- Violet-black
- Colorless
Correct Answer: Red or pink
Q3. The ferric chloride test primarily detects which functional group common to flavonoids?
- Ether linkages
- Phenolic hydroxyl groups
- Alkene double bonds
- Glycosidic oxygen
Correct Answer: Phenolic hydroxyl groups
Q4. What is the typical observation for the alkaline reagent (NaOH) test with flavonoids?
- Formation of a blue precipitate
- Intense yellow coloration that disappears on acidification
- Permanent green coloration
- No change in color
Correct Answer: Intense yellow coloration that disappears on acidification
Q5. What is the principle of the AlCl3 test used in flavonoid analysis?
- Oxidation of flavonoids to quinones
- Formation of chelate complexes with hydroxyl groups causing bathochromic shift and fluorescence
- Precipitation of flavonoids as insoluble salts
- Hydrolysis of glycosides to liberate sugars
Correct Answer: Formation of chelate complexes with hydroxyl groups causing bathochromic shift and fluorescence
Q6. In UV-visible spectroscopy of flavonoids, what are the typical wavelength regions for Band I and Band II?
- Band I: 300–380 nm (cinnamoyl); Band II: 240–280 nm (benzoyl)
- Band I: 200–220 nm; Band II: 400–450 nm
- Band I: 150–200 nm; Band II: 210–230 nm
- Band I: 420–480 nm; Band II: 300–320 nm
Correct Answer: Band I: 300–380 nm (cinnamoyl); Band II: 240–280 nm (benzoyl)
Q7. Which reagent produces a characteristic large bathochromic shift for flavonols due to the free 3-hydroxyl group?
- Sodium methoxide (NaOMe)
- Concentrated sulfuric acid
- Lead acetate solution
- Silver nitrate
Correct Answer: Sodium methoxide (NaOMe)
Q8. The lead acetate test for flavonoids typically gives which result?
- Blue fluorescence under UV
- Yellow precipitate indicating phenolic compounds
- Immediate gas evolution
- No visible reaction
Correct Answer: Yellow precipitate indicating phenolic compounds
Q9. What is the outcome of acid hydrolysis of flavonoid glycosides during qualitative analysis?
- Conversion of aglycone to glycoside
- Formation of aglycone (flavonoid) and free sugars
- Polymerization into tannins
- Complete volatilization of compounds
Correct Answer: Formation of aglycone (flavonoid) and free sugars
Q10. Which structural difference distinguishes flavonols from flavones?
- Flavonols have a saturated C2–C3 bond; flavones have a double bond
- Flavonols possess a free 3-hydroxyl group; flavones lack the 3-OH
- Flavonols contain a sugar moiety; flavones are always aglycones
- Flavonols are anthocyanins; flavones are chalcones
Correct Answer: Flavonols possess a free 3-hydroxyl group; flavones lack the 3-OH
Q11. How do anthocyanin colors change with pH during simple spot tests?
- They are colorless at all pH values
- Red in acidic medium and blue/green in alkaline medium
- Blue in acidic medium and red in alkaline medium
- Always give a persistent yellow color
Correct Answer: Red in acidic medium and blue/green in alkaline medium
Q12. Which of the following tests is NOT used for detecting flavonoids?
- Shinoda test
- Ferric chloride test
- Bornträger’s test
- Alkaline reagent test
Correct Answer: Bornträger’s test
Q13. Which mobile phase is commonly used for TLC separation of flavonoids in phytochemical analysis?
- Toluene : Ethyl acetate : Formic acid (5 : 4 : 1)
- Hexane only
- Water only
- Methanol : Chloroform (1 : 1)
Correct Answer: Toluene : Ethyl acetate : Formic acid (5 : 4 : 1)
Q14. Which combined reagent sequence helps differentiate flavonols from flavones by UV shift behavior?
- AlCl3, then HCl — shift persists for flavonols but is lost for flavones
- NaOH then AgNO3 — only flavones react
- Lead acetate followed by NaBH4 — only flavanones show changes
- Ferric chloride then sodium chloride — specific for anthocyanins
Correct Answer: AlCl3, then HCl — shift persists for flavonols but is lost for flavones
Q15. The intense yellow color seen with NaOH in flavonoid tests arises from which chemical phenomenon?
- Formation of a phenolate ion through deprotonation of phenolic OH
- Oxidative cleavage of the B-ring
- Precipitation of metallic salts
- Esterification of hydroxyl groups
Correct Answer: Formation of a phenolate ion through deprotonation of phenolic OH
Q16. Which solvent is most suitable for extracting flavonoid glycosides from plant material?
- Non-polar solvent like hexane
- Polar solvents such as methanol or 70% ethanol
- Pure benzene
- Liquid nitrogen only
Correct Answer: Polar solvents such as methanol or 70% ethanol
Q17. Mechanistically, what does Mg/HCl do to flavonoid aglycones in the Shinoda test?
- Oxidizes the molecule to a quinone with blue color
- Reduces the C-ring and forms colored flavilium species (red/pink)
- Hydrolyzes glycosidic bonds selectively
- Forms insoluble metallic complexes
Correct Answer: Reduces the C-ring and forms colored flavilium species (red/pink)
Q18. Which simple test can indicate the presence of chalcones among flavonoid-related compounds?
- Chalcones give intense yellow color with NaOH due to enolate formation
- Chalcones give a purple precipitate with lead acetate
- Chalcones are colorless with ferric chloride
- Chalcones produce red fluorescence with NP/PEG
Correct Answer: Chalcones give intense yellow color with NaOH due to enolate formation
Q19. Which reagent specifically forms chelates involving the 3-hydroxyl and 4-carbonyl group in flavonoids, aiding spectral interpretation?
- Aluminum chloride (AlCl3)
- Sodium chloride
- Silver nitrate
- Potassium permanganate
Correct Answer: Aluminum chloride (AlCl3)
Q20. Which class of flavonoids is characterized by a saturated C2–C3 bond (no C2=C3 double bond)?
- Flavones
- Flavanones
- Flavonols
- Anthoocyanins
Correct Answer: Flavanones
Q21. For confirmatory identification of flavonoids by UV-visible spectroscopy, which approach is most informative?
- Recording spectra before and after addition of shift reagents such as NaOMe and AlCl3
- Measuring only at 254 nm
- Using visible light only (400–700 nm)
- Assessing turbidity changes
Correct Answer: Recording spectra before and after addition of shift reagents such as NaOMe and AlCl3
Q22. Which substances commonly cause false positives in the ferric chloride test for flavonoids?
- Tannins and other phenolic compounds
- Aliphatic hydrocarbons
- Proteins exclusively
- Inorganic salts like NaCl
Correct Answer: Tannins and other phenolic compounds
Q23. A catechol-type B-ring in a flavonoid typically gives what color with ferric chloride?
- Bright yellow only
- Green to blue coloration
- No reaction
- White precipitate
Correct Answer: Green to blue coloration
Q24. After acid hydrolysis of a flavonoid glycoside, which test can confirm the presence of released sugars?
- Molisch’s test producing a purple ring
- Shinoda test producing red color
- AlCl3 test producing fluorescence
- Ferric chloride giving green color
Correct Answer: Molisch’s test producing a purple ring
Q25. Which colorimetric assay is commonly used to quantify total flavonoids in extracts?
- AlCl3 colorimetric assay measured by UV spectrophotometry (around 415 nm)
- Biuret test measured at 540 nm
- Gravimetric precipitation with NaCl
- Flame photometry of dried residue
Correct Answer: AlCl3 colorimetric assay measured by UV spectrophotometry (around 415 nm)
Q26. Which TLC visualization reagent produces characteristic fluorescence for many flavonoids under UV light?
- NP/PEG (diphenylboric acid 2-aminoethyl ester followed by PEG)
- ninhydrin
- Dragendorff’s reagent
- Wagner’s reagent
Correct Answer: NP/PEG (diphenylboric acid 2-aminoethyl ester followed by PEG)
Q27. The vanillin-HCl test is especially useful for detecting which flavonoid-related compounds?
- Flavan-3-ols (catechins) and condensed tannins producing a red coloration
- Flavones producing blue color
- Anthraquinones producing pink color
- Terpenes producing white precipitate
Correct Answer: Flavan-3-ols (catechins) and condensed tannins producing a red coloration
Q28. Which analytical technique provides definitive structural differentiation between isomeric flavones and flavonols?
- NMR spectroscopy (1H and 13C NMR)
- Simple spot test with NaOH
- Paper chromatography only
- Color comparison by eye
Correct Answer: NMR spectroscopy (1H and 13C NMR)
Q29. What important safety consideration applies when using lead acetate in qualitative flavonoid tests?
- Lead acetate is non-toxic and requires no precautions
- Lead acetate is toxic and must be handled with appropriate PPE and disposal procedures
- Lead acetate reacts explosively with methanol
- Lead acetate neutralizes all phenolics
Correct Answer: Lead acetate is toxic and must be handled with appropriate PPE and disposal procedures
Q30. Upon hydrolysis of a flavonoid glycoside, how does the Rf value of the aglycone compare to the glycoside on TLC?
- The aglycone is more polar and has a lower Rf
- The aglycone is less polar and typically shows a higher Rf
- Both have identical Rf values always
- Glycoside evaporates leaving no spot
Correct Answer: The aglycone is less polar and typically shows a higher Rf
I am a Registered Pharmacist under the Pharmacy Act, 1948, and the founder of PharmacyFreak.com. I hold a Bachelor of Pharmacy degree from Rungta College of Pharmaceutical Science and Research. With a strong academic foundation and practical knowledge, I am committed to providing accurate, easy-to-understand content to support pharmacy students and professionals. My aim is to make complex pharmaceutical concepts accessible and useful for real-world application.
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