Premium
Method of Detection of Auxin‐Indole‐3‐Acetic Acid in Soils by High Performance Liquid Chromatography
Author(s) -
Frankenberger W. T.,
Brunner W.
Publication year - 1983
Publication title -
soil science society of america journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.836
H-Index - 168
eISSN - 1435-0661
pISSN - 0361-5995
DOI - 10.2136/sssaj1983.03615995004700020012x
Subject(s) - chemistry , chromatography , high performance liquid chromatography , reagent , indole 3 acetic acid , detection limit , indole test , acetic acid , absorbance , column chromatography , tryptophan , mass spectrometry , auxin , organic chemistry , biochemistry , amino acid , gene
A sensitive, specific, and precise method for detection of indole‐3‐acetic acid (IAA) in soils is described. This method involves the use of reverse phase, ion‐pair, high performance liquid chromatography (HPLC) to detect IAA produced from L‐tryptophan (L‐TRP) added to soils. A 20‐µL aliquot of the soil extract was injected in the eluent stream consisting of a 60:40 CH 3 OH:H 2 O solution containing 0.01 M l‐heptanesulfonic acid (ion‐pair reagent) which was acified to pH 2.5 with H 3 PO 4 and separated with an analytical ultrasphere‐ODS (C 18 ) column. Ultraviolet‐absorbance (254 nm) was used as the mode of detection. Separation of indole‐3‐acetamide (IAM) indole‐3‐pyruvic acid (IPyA) and IAA (identified by mass spectrometry) was achieved only in the presence of the ion‐pair reagent. Detection of these 3‐substituted indoles in soils incubated with L‐TRP suggests that IAA is synthesized by two distinct pathways: (i) TRP is converted to IAA by deaminating to IPyA followed by decarboxylation to indole‐3‐acetaldehyde (IAAld) which is further oxidized to IAA, and (ii) TRP is decarboxylated to IAM which is hydrolyzed to IAA. The detection limit for IAA by the method described is 50 pg/mL of methanol under standardized conditions. The concentration of IAA can be determined in soil extracts by HPLC in < 5 min with the precision of 1.3 to 3.4% C.V.