A microtechnique for the analysis of free and conjugated indole-3-acetic acid in milligram amounts of plant tissue using a benchtop gas chromatograph-mass spectrometer

David M. Ribnicky; Todd J. Cooke; Jerry D. Cohen

doi:10.1007/s004250050223

A microtechnique for the analysis of free and conjugated indole-3-acetic acid in milligram amounts of plant tissue using a benchtop gas chromatograph-mass spectrometer

David M. Ribnicky, Todd J. Cooke, Jerry D. Cohen

Horticultural Science

Research output: Contribution to journal › Article › peer-review

31 Scopus citations

Abstract

A microtechnique was developed for the quantification of indole-3-acetic acid (IAA) in plant samples of one milligram fresh weight or less. The method permitted quantification of both free and conjugated IAA using a benchtop gas chromatograph-mass spectrometer. New methods for sample purification with high recovery at microscale levels, together with simple changes that result in enhanced sensitivity of the instrumentation, allowed for a significant reduction in the amount of plant material required for analysis. Single oat (Avena sativa L.) coleoptile tips could be studied with this method and were found to contain free and total IAA levels of 137 and 399 pg · mg^-1 fresh weight, respectively. A single 5-d-old Arabidopsis thaliana (L.) Heynh. seeding was shown to contain 61 pg · mg^-1 fresh weight free IAA and 7850 pg · mg^-1 fresh weight of total IAA following basic hydrolysis. This microtechnique provides a way to accurately measure IAA levels in very small structures and individual seedlings, thus making it a valuable research tool for elucidating the role and distribution of auxin in relation to growth and development.

Original language	English (US)
Pages (from-to)	1-7
Number of pages	7
Journal	Planta
Volume	204
Issue number	1
DOIs	https://doi.org/10.1007/s004250050223
State	Published - Dec 1997

Bibliographical note

Funding Information:
We thank Dr. Kai-Hsien Chen (Hewlett Packard, Taiwan) for helpful suggestions which facilitated the development of this technique, Eric Schoettker (Hewlett Packard Co.) for help with repairs and modification of the mass spectrometer necessary for increased sensitivity and Drs. Jennifer Normanly (University of Massachusetts) and Martha Koester (Weyerhaeuser Co.) for helping with the development of this technique for use as a generally available method. We also thank Drs. Normanly and Koester as well as Drs. Harry J. Swartz (University of Maryland) and Janet P. Slovin (USDA-ARS Climate Stress Laboratory, Beltsville, Md., USA) for their helpful comments on the manuscript. Supported by U.S. Department of Energy grant DE-AI02-94-ER20153 and by the Maryland Agricultural Experiment Station.

Keywords

Arabidopsis (auxin analysis)
Auxin analysis
Avena (auxin analysis)
Mass spectrometry
Stable isotopes

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10.1007/s004250050223

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title = "A microtechnique for the analysis of free and conjugated indole-3-acetic acid in milligram amounts of plant tissue using a benchtop gas chromatograph-mass spectrometer",

abstract = "A microtechnique was developed for the quantification of indole-3-acetic acid (IAA) in plant samples of one milligram fresh weight or less. The method permitted quantification of both free and conjugated IAA using a benchtop gas chromatograph-mass spectrometer. New methods for sample purification with high recovery at microscale levels, together with simple changes that result in enhanced sensitivity of the instrumentation, allowed for a significant reduction in the amount of plant material required for analysis. Single oat (Avena sativa L.) coleoptile tips could be studied with this method and were found to contain free and total IAA levels of 137 and 399 pg · mg-1 fresh weight, respectively. A single 5-d-old Arabidopsis thaliana (L.) Heynh. seeding was shown to contain 61 pg · mg-1 fresh weight free IAA and 7850 pg · mg-1 fresh weight of total IAA following basic hydrolysis. This microtechnique provides a way to accurately measure IAA levels in very small structures and individual seedlings, thus making it a valuable research tool for elucidating the role and distribution of auxin in relation to growth and development.",

keywords = "Arabidopsis (auxin analysis), Auxin analysis, Avena (auxin analysis), Mass spectrometry, Stable isotopes",

author = "Ribnicky, {David M.} and Cooke, {Todd J.} and Cohen, {Jerry D.}",

note = "Funding Information: We thank Dr. Kai-Hsien Chen (Hewlett Packard, Taiwan) for helpful suggestions which facilitated the development of this technique, Eric Schoettker (Hewlett Packard Co.) for help with repairs and modification of the mass spectrometer necessary for increased sensitivity and Drs. Jennifer Normanly (University of Massachusetts) and Martha Koester (Weyerhaeuser Co.) for helping with the development of this technique for use as a generally available method. We also thank Drs. Normanly and Koester as well as Drs. Harry J. Swartz (University of Maryland) and Janet P. Slovin (USDA-ARS Climate Stress Laboratory, Beltsville, Md., USA) for their helpful comments on the manuscript. Supported by U.S. Department of Energy grant DE-AI02-94-ER20153 and by the Maryland Agricultural Experiment Station.",

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AU - Cohen, Jerry D.

N1 - Funding Information: We thank Dr. Kai-Hsien Chen (Hewlett Packard, Taiwan) for helpful suggestions which facilitated the development of this technique, Eric Schoettker (Hewlett Packard Co.) for help with repairs and modification of the mass spectrometer necessary for increased sensitivity and Drs. Jennifer Normanly (University of Massachusetts) and Martha Koester (Weyerhaeuser Co.) for helping with the development of this technique for use as a generally available method. We also thank Drs. Normanly and Koester as well as Drs. Harry J. Swartz (University of Maryland) and Janet P. Slovin (USDA-ARS Climate Stress Laboratory, Beltsville, Md., USA) for their helpful comments on the manuscript. Supported by U.S. Department of Energy grant DE-AI02-94-ER20153 and by the Maryland Agricultural Experiment Station.

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N2 - A microtechnique was developed for the quantification of indole-3-acetic acid (IAA) in plant samples of one milligram fresh weight or less. The method permitted quantification of both free and conjugated IAA using a benchtop gas chromatograph-mass spectrometer. New methods for sample purification with high recovery at microscale levels, together with simple changes that result in enhanced sensitivity of the instrumentation, allowed for a significant reduction in the amount of plant material required for analysis. Single oat (Avena sativa L.) coleoptile tips could be studied with this method and were found to contain free and total IAA levels of 137 and 399 pg · mg-1 fresh weight, respectively. A single 5-d-old Arabidopsis thaliana (L.) Heynh. seeding was shown to contain 61 pg · mg-1 fresh weight free IAA and 7850 pg · mg-1 fresh weight of total IAA following basic hydrolysis. This microtechnique provides a way to accurately measure IAA levels in very small structures and individual seedlings, thus making it a valuable research tool for elucidating the role and distribution of auxin in relation to growth and development.

AB - A microtechnique was developed for the quantification of indole-3-acetic acid (IAA) in plant samples of one milligram fresh weight or less. The method permitted quantification of both free and conjugated IAA using a benchtop gas chromatograph-mass spectrometer. New methods for sample purification with high recovery at microscale levels, together with simple changes that result in enhanced sensitivity of the instrumentation, allowed for a significant reduction in the amount of plant material required for analysis. Single oat (Avena sativa L.) coleoptile tips could be studied with this method and were found to contain free and total IAA levels of 137 and 399 pg · mg-1 fresh weight, respectively. A single 5-d-old Arabidopsis thaliana (L.) Heynh. seeding was shown to contain 61 pg · mg-1 fresh weight free IAA and 7850 pg · mg-1 fresh weight of total IAA following basic hydrolysis. This microtechnique provides a way to accurately measure IAA levels in very small structures and individual seedlings, thus making it a valuable research tool for elucidating the role and distribution of auxin in relation to growth and development.

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KW - Avena (auxin analysis)

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A microtechnique for the analysis of free and conjugated indole-3-acetic acid in milligram amounts of plant tissue using a benchtop gas chromatograph-mass spectrometer

Abstract

Bibliographical note

Keywords

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