Antibiotics Laboratory: Chemical Reactions and Color-Based Identification of Antibiotics

Antibiotics are chemical substances produced by living microorganisms that have bacteriostatic or bactericidal properties against other living microorganisms. Classifying reactions to antibiotics is difficult because antibiotics have different structures. Color reaction 1. Concentrated acids 2. FeCl3 3. Marquis: 2 drops of formalin + H2SO4concentrated 4.Frohde reaction: (Amm. Molybdate 0.5% in water) + H2SO4concentrated Fehling reaction B. Precipitation/Crystal Reaction 1. Acetone -water 2. Complex Fe 3. Dragendorf a. Tools and materials Antibiotic compound reagents, test tubes, dropper pipettes, tube racks, glass objects, cover glasses, microscopes, spirit lamps, sublimation rings b. Work procedures 1. Amoxicillin a. Dissolve Substance in water + Fehling → ↓red with brown solution b. Substance + FeCl3→ brown c. Substance + H2SO4Dark → yellow d. Substance + HNO3Dark → yellow e. Substance + Diazo A & B reagent → red f. Dissolve the substance in water, heat in a water bath for 2 minutes while adding 5 drops of mercury nitrite in an acidic atmosphere → red g. Dissolve Substance in water + CuSO4in NaOH → purple h. Substance + Pb. Acetate → ↓ black i. Crystal reactions : water acetone, Mayer, Dragendorf. 2. Ampicillin a. Substance + H2SO4Dark → yellow b. Substance + HNO3Dark → yellow c. Substances in alcohol + H2SO4+ resorcin, heated→ yellow green d. Substances in alcohol + lart. Ammoniacal copper nitrate, heated→ greendark yellow e. Substance + Diazo A & B reagent → red f. suspension of substance in water + 2 ml Fehling + water → violet/ red violet g. suspension of substance in water + 2 ml Potassium Copper (II) Tartrate → violet h. suspension of substances in water + CuSO4in NaOH→ purple i. Substance + FeCl3→ yellow brown/brown j. Substance + Pb. Acetate → ↓ black k. Substance + potassium CuSO4-water (2:6) → reddish purple l.Crystal reactions : water acetone, Mayer, Dragendorf. 3. Chloramphenicol 1. Substances in water + H2SO4→ negative 2. Zat + Marquis → green 3. Substance + concentrated HCl + heated Zn powder → after cooling + DAB HCl 2 drops → orange/yellow 4. Substance + NaOH is then heated → orange red 5. Substance + Cuprifil→ dark blue, when heated → ↓ brick red 6. Zat + Carletti → purple 7. Substance + FeCl3→ Yellow Orange 8. Substance + Azo→ bright red 9. Substance + Nessler→ ↓grey/Yellow 10. Substance + Deniges → ↓ white 11. Zat + Mayer→ ↓ white 12. Zat + Dragendorf→ light orange 13. Substance in water + 5 drops of Cu(NH3)2(NO3)2,let stand 5 minutes, heat 2 minutes → gray brown 14. Substance in water + AgNO3→ no precipitate is formed 15. Substance + p-DAB-HCl→ colorless, heat + Zn +HCl + HCl→ yellow 16. Substances in water red pyridine, yellow water layer ppt 17.Crystal reactions : water acetone. p-DAB- + 2 ml NaOH 40% Pyridine,heat slowly→ layer 4. Thiamphenicol a. Zat + Marquis → weak green b. Substance + Cuprifil→ blue c. Zat + Carletti → weak purple d. Substance + Deniges → ↓ white e. Substance + FeCl3→ Bright yellow f. Substance + Azo→ red g. Substance + Nessler→ Orange yellow h. Substance + Mayer→ ↓ white i. Substance + Dragendorf→ yellow green j. Substance in water + 5 drops of Cu(NH3)2(NO3)2,let stand 5 minutes, heat 2 minutes → gray brown k. Substance in water + AgNO3→ no precipitate is formed l. Substance + p-DAB-HCl→ colorless, heat + Zn +HCl + HCl→ yellow m. Substances in water Bright yellow pyridine, yellow water layer ppt n.Crystal reactions : water acetone. p-DAB- + 2 ml NaOH 40% Pyridine,heat slowly→ layer 5. Cephalexin a. Substance in water + Hydroxylamine HCl + NaOH, leave for 5 minutes + HCl + FeCl3→ purple/red b. Substance in water + Potassium Cupril tartrate solution → purple/green which then turns yellow/brown when left to stand c. Substances in water + FeCl3→ colorless d. Substance + parapormaldehyde solution in H2SO4→ yellow, then when heated for 2 minutes and immediately cooled it remains yellow 6. Tetracycline a. Zat + Marquis → burgundy b. Substance + H2SO4Dark → purple red diluted with water → dark yellow c. Zat + Frohde → burgundy/purple blue d. Substance + HNO3solid → negative (orange) e. Substance + FeCl3→ light brown f. Zat + Vitalli → yellow/brown g. Substance + Diazo A+ Diazo B+Na2CO3→ orange red h. Zat + Marquis → orange brown/burgundy i. Substance + Nessler (+ NaOH) → brown-black j. Zat + Nessler → brown-black k. Zat + Millon → rosa, stir → chocolate l. Substance + AgNO3→ reduction m. Zat + Benedict → purple green n. Substance + vanillin-H2SO4→ green solution, heated → moss green o. Zat + Amm. molybdate → blue black p. Substance + H2SO4thick → purple green, if you add water → dark yellow 7. Oxytetracycline a. Substance + H2SO4intense → violet red (color appears after 2 minutes). After 5 minutes add water ± 1 ml → yellow b. Heat 2 ml of ZnCl2 solution in a steam cup + Substance →purple blue c. Substance + FeCl3+ ethanol 96 % → dark brown d. Substance + H2SO4dark → bright red, if you add water → dark reddish brown e. Substance + NaHCO31 % + diazo benzene sulfonate → orange red 8. Erythromycin a. Substance + water + H2SO4 concentrated, shake→ red brown b. Zat + Nessler → gray black c. Substance + HNO3solid → yellow + water → green d. Substance in water + 5 tts Cu(NO3)2ammoniacal, leave for 5 minutes, then heat → gray brown e. Dissolve 3 mg of the substance in 2 ml of acetone and concentrated HCl→ orange→ red → purple red. When chloroform is added and shaken, the chloroform solution turns purple.