Ninhydrin reagent was first discovered in by a German-English chemist called Siegfried Ruhemann in 1910 (Ruhemann, 1910). Ruhemann outlined the synthesis of this reagent, where he indicated that the colorless prisms of ninhydrin turn red at the temperature of 125oC. In 1913 Abderhalden observed that ninhydrin could be a valuable reagent for the detention of dialyzable amino acid in various tissues such as urine, saliva, blood plasma, and serum. Before the discovery of ninhydrin being used as a reagent for detecting fingerprints, it found the usage in various biochemical and medical test methods. The idea of using ninhydrin reagent for detecting latent fingerprints was suggested by Oden and Von Hofsten in 1954 (Oden & Hofsten, 1954). Many countries and crime laboratories have used the Ninhydrin technique for the development of latent fingerprints to be used in criminal procedure.
The ninhydrin reagents detect ammonia or primary and secondary amines, and for many years ninhydrin has been used as a reagent for visualization of latent fingerprints on absorbent surfaces such as paper. Acetone was the best-suited solvent for the development of ninhydrin. The use of acetone was due to the reason that it is highly soluble with ninhydrin where one can easily dissolve more than enough of ninhydrin to make a useful working solution. Ninhydrin reagent is commonly dissolved in a nonflammable hydro-carbon solvent with other addictives to help it to dissolve since it is only soluble in polar solvent easily. More research leads to an alternative ninhydrin formulation based on petroleum with a solution in the place of acetone with minor additions of methanol. Diethyl ether could be used as a solvent, but it was regarded as too volatile for spraying on the documents because of the flammable atmosphere. Adaptation of the ninhydrin reaction product began to be considered within most laboratory research on the fingerprints, which lead to more innovation and discovery. More research to synthesize analogs of ninhydrin was done with the motive of either to change the color of principle reaction product with amino acid.
Ninhydrin Reaction With Amino Acids
Ninhydrin reacts with the amino group of primary amino acids to produce a purple color refers to us Ruhemann’s purple. The production of Ruhemann’s purple color serves as the primary detection and quantitative detention of amino acids. The chromophore formed is the same in all primary acids where the intensity of the color created depends on the amount and chemical nature of the amino group used. The color produced gives the basis for the quantitative determination of amino acids. The color reaction with ninhydrin is used immensely in the manual and automated procedures. The chemical reaction between ninhydrin and amino acids contained in the fingerprint residue is relatively slow at room temperature. The reaction process is accelerated by steaming or heating the substance with an iron, which has a disadvantage of producing fingerprints, which may not be as highly visible.
The best catalyst to use is Acetic acid and water, where the acetic acid content should be kept as low as possible to minimize any ink diffusion on the material being treated. The reaction formed between ninhydrin and different amino acid are all not purple, and the color of the fingerprint created can vary from nearly red to deep violet depending on the composition of the fingerprint. Different in color has been borne out by the observation of the developed fingerprint of the various individuals. Another factor that may contribute to having different colors after the reaction is due to the reason that the reaction did not proceed to completion. The color of the intermediate amine compound depends on the group of primary amino acids used. The familiar violet color during the reaction of amino acid with ninhydrin is attributed to the anion of this compound. Under specific temperature, condition ammonia condenses to a greater or lesser extent with the reagent to form diketohydrindamine (DYDA). The inconsistence with this reaction is that the ammonia reacts more slowly and less thoroughly than do amino acid under the same condition.
Laboratory research has shown that the use of ninhydrin to detect amino acid with high sensitivity is much less sensitive towards a variety of standard proteins. The reason behind this is protein has only one free amino group at the N-terminal tail.
Introducing the New Contact Ninhydrin Method
The primary amino acid may be quantitatively dominated by ninhydrin with the formation of carbon dioxide and aldehyde. The new contact ninhydrin method recently developed has seen the reaction of amino acid into a rapid and convenient photometric for easy estimation. The introduction of new contact reagent into the system materially increases the amount of color and has made it possible to obtain consistent and reproducible results. A modified procedure is described for the photometric determination of amino acids with ninhydrin. The method is applied to protein hydrolysates and to urine, where a solution of a mixture of sixteen amino acids is used. The improved ninhydrin method was introduced for the quantitative determination of amino acids. Sodium acetate buffer in the presented contact ninhydrin method is used to replace lithium acetic acid. The result of using this improved method showed that a shorter heating time of about ten minutes could be achieved compared to the traditional degree of the color development that was previously used.
Application of Ninhydrin Reagent
Ninhydrin is commonly used during a forensic investigation by the criminal department to analyze the latent of fingerprint on a porous surface. The amino acid containing finger marks formed by minute sweat secretion is treated using a ninhydrin solution, which will result in turning of amino acid finger ridge pattern purple. The effects of the change in color make the finger marks to be visible and used to detect the owner. Ninhydrin reagent can also be used in the amino acid analysis of protein and to monitor deprotection in the solid phase peptide synthesis.
Oden, S. & Hofsten, V. (1954). Background of ninhydrin reagent for detecting latent fingerprints. Journal of Chemical Education. 42 (7): 386–388. DOI:10.1021/ed042p386 . http://www.bvda.com/EN/download/NFN_MoGo_full.pdf
Ruhemann, S. (1910). The discovery of ninhydrin and detention of fingerprint. Journal of chemical society. London. Transactions. 97: 1438–1449. DOI:10.1039/ct9109701438. https://www.jbc.org/content/200/2/803.full.pdf
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