Prussian blue assay

The most commonly used methods for determination of total phenolics in foodstuffs are the Prussian Blue assay and the Folin-Ciocalteau method. Although these procedures have undergone numerous modiiicalions, the version adopted by the American Oil Chemists Society (AOCS) is most commonly used nowadays. These assays are not specific for phenohc compounds, although they are widely used to get a global concentration of phenolic compounds present in foods. 

Total phenolic compounds content was assayed spectrophotometri-cally using a modified Prussian blue method as described by Graham (28) employing an LKB Biochrom Ultrospec II spectrophotometer (Cambridge, England). Tannic acid was used as calibration standard. 

Ferric-to-Ferrous Reduction— Polyphenols are able to reduce Fe to Fe " " with varying efficiencies. The production of Fe " ions can be monitored by the formation of Prussian blue, Fe4[Fe(CN)6]j. In one assay format (19), 0.2 mL of 8 mM K3Fe(CN)g was added to 3 mL of a 50-pM tannin solution, followed by the addition of 0.2 mL of 0.1 M FeCl3 in 0.1 Af HCl. The optical density was read after 5 min at 700 nm. In a cruder format, the presence of tannins can be determined by the addition of 5% FeCl3/methanol (w/v) solution to an equal volume of sample a brown coloration indicates the presence of polyphenolics. 

C. J. Geoffroy mentioned as a volatile salt the crystals deposited from essential oils. He published on antimony, tartar emetic, and kermes mineral. He showed that soap is readily soluble in hot alcohol, the liquid on coohng setting to a clear jelly, and that the oil set free from the soap (really oleic acid) by the action of acids is more soluble in alcohol than the original oil. In his paper on Prussian blue he argued that the soufre animal of the organic material sets free iron in a fine state of division which, when mixed with the sulphurous principle and absorbent earth, exhibits a blue colour. (Until recently, the blue colour of ultramarine was ascribed to finely-divided sulphur.) He translated the work on assaying of Christian Carl Schindler. ... 

It is a popular method to quantify total phenolics because it is simple, rapid, and with little interferences by nonphenolic compounds. Critical for the successful use of this assay are temperature, pH, and order in which reagents are added, since this affects the formation of Fe4[Fe(CN)e]3 (Prussian blue) [86, 87]. 

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