Total Phenolics and Proanthocyanidins:
To each test tube add 30mg of dry finely ground plant material. Add 10ml of 50% aqueous ethanol and extract for two hours shaking every 15 minutes (vortex).
Tannic acid; use 10mg, and 100 to 600µl
for a curve.
Metrosideros extract (SH) for proanthocyanidins; dissolve about 5mg into 10ml of extracting solution for a stock standard. Dilute 100/200/400 and 800µl of stock to 1ml of extracting solution for a curve.
For total phenolics
Add 200µl of initial extract to a 50ml volumetric flask containing about
25ml of water.
At time 0 add 2.5ml of Folin-Denise reagent and mix well.
After 3 minutes add 5ml of saturated sodium carbonate and bring to volume with water; mix well.
After exactly 20 minutes, measure the absorbance at 760nm
Add 1.0ml initial extract to a test tube.
Add 10ml of the n-butanol-HCL reagent and mix well (vortex).
Place on dry bath at 100°C for 1 hour.
Let cool and measure absorption at 550nm
To 750ml water add 100g sodium tungstate and 20g phosphomolybdic acid to a 2L flask. Add 50ml orthophosphoric acid. Reflux for 2h. Allow to cool and make upto 1L. Store in dark.
Dissolve .7g ferrous sulfate heptahydrate in 25ml HCL and a little n-butanol. Then dilute to 1L with n-butanol.
Saturated Sodium carbonate
Dissolve 350g sodium carbonate/1l water at 70-80°C
reference for Folin-Denise reagent and Saturated Sodium carbonate; Official Methods of Analysis of the AOAC, Seventh edition, 1950
reference for n-butanol-HCL reagent; S. Mole and PG Waterman, A critical analysis of techniques for measuring tannins in ecological studies., Oecologia 1987 72:137-147