Detection of primary and secondary amines using ninhydrin
ID: 198784 | No.: 660 | Alfred Krause
School experiment from Year 5 onwardsCategory: Evidence
Five different aqueous solutions are prepared in test tubes (TG) and mixed with ninhydrin solution.
1. Blank sample: 5 mL of demineralised water is poured into test tube 1 and 0.5 mL of ninhydrin solution is added. The mixture is heated to approximately 50 °C using a hot-air blower.
2. Glycine: 5 mL of a 0.05 mol/L glycine solution is mixed with 0.5 mL of ninhydrin solution. The mixture is heated with a hot-air blower until a violet colour develops. When left to stand, the colour deepens to a deep violet.
3. Proline: 5 mL of a 0.05 mol/L proline solution is mixed with 0.5 mL of ninhydrin solution. The mixture is heated with a hot-air blower until a yellow colour becomes visible. This deepens when left to stand.
4. Optional: Dimethylglycine. Approximately 1 mL of the reaction mixture from the ‘Preparation of dimethylglycine’ experiment is diluted with 2 mL of water and 0.5 mL of ninhydrin solution is added. The mixture is heated to approximately 50 °C using a hot-air blower. No significant colour change is observed.
5. Optional: Methylproline. Approximately 1 mL of the reaction mixture from the ‘Preparation of methylproline’ experiment is diluted with 2 mL of water and 0.5 mL of ninhydrin solution is added. The mixture is heated to approximately 50 °C using a hot-air blower. No significant colour change is observed.
Optional supplement: Thin-layer chromatography detection
- TLC plates: silica gel on glass (2.5 × 7.5 cm)
- Mobile phase: acetone / isopropanol / acetic acid (25%) in a ratio of 7 mL : 7 mL : 6 mL
- Running time: approx. 30 min
- Application: glycine or proline on the left; nicotinic acid solution on the right as a control (see degintu No. 198730)
After blow-drying the HPLC plates (in a fume cupboard), they are first treated with ninhydrin spray reagent. Once the spots have dried and been marked, the tertiary amine in the nicotine can be visualised using Dragendorff’s reagent.
Fume cupboard, 5 test tubes, test tube rack, hot-air blower, DC plates (silica gel), DC chamber, plastic pipettes
| Name | Formula | CAS no. | H-statements | P-statements | Pictograms |
|---|---|---|---|---|---|
| Glycin | NH₂CH₂COOH | 56-40-6 | |||
| Prolin | C₅H₉NO₂ | 609-36-9 | |||
| Ninhydrin-Sprühlösung 0,5% in Ethanol | C₉H₆O₄ | 485-47-2 |
H225
– Liquid and vapour are highly flammable.
German original: Flüssigkeit und Dampf leicht entzündbar.
H319
– Causes serious eye irritation.
German original: Verursacht schwere Augenreizung. |
P210
– Keep away from heat, hot surfaces, sparks, open flames and other sources of ignition. No smoking.
German original: Von Hitze, heißen Oberflächen, Funken, offenen Flammen und anderen Zündquellen fernhalten. Nicht rauchen.
P240
– Earth the container and the equipment to be filled.
German original: Behälter und zu befüllende Anlage erden.
P403+P233
– Store the container tightly sealed in a well-ventilated place.
German original: Behälter dicht verschlossen an einem gut belüfteten Ort aufbewahren.
P305+P351+P338
– IF IN EYES: Rinse gently with water for several minutes. Remove contact lenses, if possible. Continue rinsing.
German original: BEI KONTAKT MIT DEN AUGEN: Einige Minuten lang behutsam mit Wasser spülen. Vorhandene Kontaktlinsennach Möglichkeit entfernen. Weiter spülen. |
GHS02, GHS07 |
| Ninhydrin | C₉H₆O₄ | 485-47-2 |
H302
– Harmful if swallowed.
German original: Gesundheitsschädlich bei Verschlucken.
H315
– Causes skin irritation.
German original: Verursacht Hautreizungen.
H319
– Causes severe eye irritation.
German original: Verursacht schwere Augenreizung. |
P301+P312+P330
– IF SWALLOWED: If you feel unwell, call the POISON INFORMATION CENTRE or a doctor. Rinse your mouth.
German original: BEI VERSCHLUCKEN: Bei Unwohlsein GIFTINFORMATIONSZENTRUM oder Arzt anrufen. Mund ausspülen.
P305+P351+P338
– IF IN EYES: Rinse gently with water for several minutes. Remove any contact lenses if possible. Continue rinsing.
German original: BEI KONTAKT MIT DEN AUGEN: Einige Minuten lang behutsam mit Wasser spülen. Vorhandene Kontaktlinsennach Möglichkeit entfernen. Weiter spülen. |
GHS07 |
| N,N-Dimethylglycin | C₄H₉NO₂ | 1118-68-9 |
H302
– Harmful if swallowed.
German original: Gesundheitsschädlich bei Verschlucken. |
P264
– After use … wash thoroughly.
German original: Nach Gebrauch ... gründlich waschen.
P270
– Do not eat, drink or smoke whilst using this product.
German original: Bei Gebrauch nicht essen, trinken oder rauchen.
P280
– Wear protective gloves / protective clothing / eye protection / face protection.
German original: Schutzhandschuhe / Schutzkleidung / Augenschutz / Gesichtsschutz tragen.
P301+P312
– IF SWALLOWED: If you feel unwell, call the POISON INFORMATION CENTRE / a doctor ...
German original: BEI VERSCHLUCKEN: Bei Unwohlsein GIFTINFORMATIONSZENTRUM / Arzt ... anrufen. |
GHS07 |
| N-Methylprolin | C₆H₁₁NO₂ | 475-11-6 |
H/P statements: machine translation, with the German original below (authoritative). Name, formula, CAS no. and pictograms are not translated.
-
Safety goggles must be worn.German originalEine Gestellschutzbrille ist zu tragen.
-
Disposable nitrile gloves are used as splash protection.German originalAls Spritzschutz dienen Nitril-Einmalhandschuhe.
-
A long, full-length laboratory coat (protective coat) must be worn.German originalEin langer, geschlossener Labormantel (Schutzkittel) ist zu tragen.
-
through skin contactGerman originaldurch Hautkontakt
-
through eye contactGerman originaldurch Augenkontakt
-
Fire hazardGerman originalBrandgefahr
The DC plates must be dried with a hairdryer under a fume cupboard. The same applies to spraying them with ninhydrin or Dragendorff’s reagent.
Das Trockenföhnen der DC-Platten ist im Abzug durchzuführen. Ebenso das Besprühen mit Ninhydrin, bzw. Dragendorff.
There are no known hazardous properties of the substances or other risks that would, in principle, preclude the experiment from being carried out by pupils or teachers. The list of substances in DGUV Information 213-098 at degintu.dguv.de has been taken into account.
Experimental procedure: Detection of primary and secondary amines using ninhydrin (2026) Krause, A. and Klemeyer, H. DOI: 10.5281/zenodo.22831346