An improved butanol-HCl assay for quantification of water-soluble, acetone:methanol-soluble, and insoluble proanthocyanidins (condensed tannins).
Identifieur interne : 001526 ( Main/Exploration ); précédent : 001525; suivant : 001527An improved butanol-HCl assay for quantification of water-soluble, acetone:methanol-soluble, and insoluble proanthocyanidins (condensed tannins).
Auteurs : Philip-Edouard Shay ; J A Trofymow ; C Peter ConstabelSource :
- Plant methods [ 1746-4811 ] ; 2017.
Abstract
BACKGROUND
Condensed tannins (CT) are the most abundant secondary metabolite of land plants and can vary in abundance and structure according to tissue type, species, genotype, age, and environmental conditions. Recent improvements to the butanol-HCl assay have separately helped quantification of soluble and insoluble CTs, but have not yet been applied jointly. Our objectives were to combine previous assay improvements to allow for quantitative comparisons of different condensed tannin forms and to test protocols for analyses of condensed tannins in vegetative plant tissues. We also tested if the improved butanol-HCl assay can be used to quantify water-soluble forms of condensed tannins.
RESULTS
Including ~50% acetone in both extraction solvents and final assay reagents greatly improved the extraction and quantification of soluble, insoluble and total condensed tannins. The acetone-based method also extended the linear portion of standard integration curves allowing for more accurate quantification of samples with a broader range of condensed tannin concentrations. Estimates of tannin concentrations determined using the protocol without acetone were lower, but correlated with values from acetone-based methods. With the improved assay, quantification of condensed tannins in water-soluble forms was highly replicable. The relative abundance of condensed tannins in soluble and insoluble forms differed substantially between tissue types.
CONCLUSIONS
The quantification of condensed tannins using the butanol-HCl assay was improved by adding acetone to both extraction and reagent solutions. These improvements will facilitate the quantification of total condensed tannin in tissues containing a range of concentrations, as well as to determine the amount in water-soluble, acetone:MeOH-soluble and insoluble forms. Accurate determination of these three condensed tannin forms is essential for careful investigations of their potentially different physiological and ecological functions.
DOI: 10.1186/s13007-017-0213-3
PubMed: 28775761
PubMed Central: PMC5539752
Affiliations:
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Box 3020, STN CSC, Victoria, BC V8W 3N5 Canada.</nlm:affiliation><wicri:noCountry code="subField">BC V8W 3N5 Canada</wicri:noCountry></affiliation><affiliation><nlm:affiliation>Pacific Forestry Centre, Canadian Forest Service, Natural Resources Canada, Victoria, BC Canada.</nlm:affiliation><wicri:noCountry code="subField">BC Canada</wicri:noCountry></affiliation></author><author><name sortKey="Constabel, C Peter" sort="Constabel, C Peter" uniqKey="Constabel C" first="C Peter" last="Constabel">C Peter Constabel</name><affiliation><nlm:affiliation>Department of Biology & Centre for Forest Biology, University of Victoria, P.O. Box 3020, STN CSC, Victoria, BC V8W 3N5 Canada.</nlm:affiliation><wicri:noCountry code="subField">BC V8W 3N5 Canada</wicri:noCountry></affiliation></author></titleStmt><publicationStmt><idno type="wicri:source">PubMed</idno><date when="2017">2017</date><idno type="RBID">pubmed:28775761</idno><idno type="pmid">28775761</idno><idno type="doi">10.1186/s13007-017-0213-3</idno><idno type="pmc">PMC5539752</idno><idno type="wicri:Area/Main/Corpus">001214</idno><idno type="wicri:explorRef" wicri:stream="Main" wicri:step="Corpus" wicri:corpus="PubMed">001214</idno><idno type="wicri:Area/Main/Curation">001214</idno><idno type="wicri:explorRef" wicri:stream="Main" wicri:step="Curation">001214</idno><idno type="wicri:Area/Main/Exploration">001214</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en">An improved butanol-HCl assay for quantification of water-soluble, acetone:methanol-soluble, and insoluble proanthocyanidins (condensed tannins).</title><author><name sortKey="Shay, Philip Edouard" sort="Shay, Philip Edouard" uniqKey="Shay P" first="Philip-Edouard" last="Shay">Philip-Edouard Shay</name><affiliation><nlm:affiliation>Department of Biology & Centre for Forest Biology, University of Victoria, P.O. Box 3020, STN CSC, Victoria, BC V8W 3N5 Canada.</nlm:affiliation><wicri:noCountry code="subField">BC V8W 3N5 Canada</wicri:noCountry></affiliation></author><author><name sortKey="Trofymow, J A" sort="Trofymow, J A" uniqKey="Trofymow J" first="J A" last="Trofymow">J A Trofymow</name><affiliation><nlm:affiliation>Department of Biology & Centre for Forest Biology, University of Victoria, P.O. Box 3020, STN CSC, Victoria, BC V8W 3N5 Canada.</nlm:affiliation><wicri:noCountry code="subField">BC V8W 3N5 Canada</wicri:noCountry></affiliation><affiliation><nlm:affiliation>Pacific Forestry Centre, Canadian Forest Service, Natural Resources Canada, Victoria, BC Canada.</nlm:affiliation><wicri:noCountry code="subField">BC Canada</wicri:noCountry></affiliation></author><author><name sortKey="Constabel, C Peter" sort="Constabel, C Peter" uniqKey="Constabel C" first="C Peter" last="Constabel">C Peter Constabel</name><affiliation><nlm:affiliation>Department of Biology & Centre for Forest Biology, University of Victoria, P.O. Box 3020, STN CSC, Victoria, BC V8W 3N5 Canada.</nlm:affiliation><wicri:noCountry code="subField">BC V8W 3N5 Canada</wicri:noCountry></affiliation></author></analytic><series><title level="j">Plant methods</title><idno type="ISSN">1746-4811</idno><imprint><date when="2017" type="published">2017</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><p><b>BACKGROUND</b></p><p>Condensed tannins (CT) are the most abundant secondary metabolite of land plants and can vary in abundance and structure according to tissue type, species, genotype, age, and environmental conditions. Recent improvements to the butanol-HCl assay have separately helped quantification of soluble and insoluble CTs, but have not yet been applied jointly. Our objectives were to combine previous assay improvements to allow for quantitative comparisons of different condensed tannin forms and to test protocols for analyses of condensed tannins in vegetative plant tissues. We also tested if the improved butanol-HCl assay can be used to quantify water-soluble forms of condensed tannins.</p></div><div type="abstract" xml:lang="en"><p><b>RESULTS</b></p><p>Including ~50% acetone in both extraction solvents and final assay reagents greatly improved the extraction and quantification of soluble, insoluble and total condensed tannins. The acetone-based method also extended the linear portion of standard integration curves allowing for more accurate quantification of samples with a broader range of condensed tannin concentrations. Estimates of tannin concentrations determined using the protocol without acetone were lower, but correlated with values from acetone-based methods. With the improved assay, quantification of condensed tannins in water-soluble forms was highly replicable. The relative abundance of condensed tannins in soluble and insoluble forms differed substantially between tissue types.</p></div><div type="abstract" xml:lang="en"><p><b>CONCLUSIONS</b></p><p>The quantification of condensed tannins using the butanol-HCl assay was improved by adding acetone to both extraction and reagent solutions. These improvements will facilitate the quantification of total condensed tannin in tissues containing a range of concentrations, as well as to determine the amount in water-soluble, acetone:MeOH-soluble and insoluble forms. Accurate determination of these three condensed tannin forms is essential for careful investigations of their potentially different physiological and ecological functions.</p></div></front></TEI><pubmed><MedlineCitation Status="PubMed-not-MEDLINE" Owner="NLM"><PMID Version="1">28775761</PMID><DateRevised><Year>2020</Year><Month>10</Month><Day>01</Day></DateRevised><Article PubModel="Electronic-eCollection"><Journal><ISSN IssnType="Print">1746-4811</ISSN><JournalIssue CitedMedium="Print"><Volume>13</Volume><PubDate><Year>2017</Year></PubDate></JournalIssue><Title>Plant methods</Title><ISOAbbreviation>Plant Methods</ISOAbbreviation></Journal><ArticleTitle>An improved butanol-HCl assay for quantification of water-soluble, acetone:methanol-soluble, and insoluble proanthocyanidins (condensed tannins).</ArticleTitle><Pagination><MedlinePgn>63</MedlinePgn></Pagination><ELocationID EIdType="doi" ValidYN="Y">10.1186/s13007-017-0213-3</ELocationID><Abstract><AbstractText Label="BACKGROUND" NlmCategory="BACKGROUND">Condensed tannins (CT) are the most abundant secondary metabolite of land plants and can vary in abundance and structure according to tissue type, species, genotype, age, and environmental conditions. Recent improvements to the butanol-HCl assay have separately helped quantification of soluble and insoluble CTs, but have not yet been applied jointly. Our objectives were to combine previous assay improvements to allow for quantitative comparisons of different condensed tannin forms and to test protocols for analyses of condensed tannins in vegetative plant tissues. We also tested if the improved butanol-HCl assay can be used to quantify water-soluble forms of condensed tannins.</AbstractText><AbstractText Label="RESULTS" NlmCategory="RESULTS">Including ~50% acetone in both extraction solvents and final assay reagents greatly improved the extraction and quantification of soluble, insoluble and total condensed tannins. The acetone-based method also extended the linear portion of standard integration curves allowing for more accurate quantification of samples with a broader range of condensed tannin concentrations. Estimates of tannin concentrations determined using the protocol without acetone were lower, but correlated with values from acetone-based methods. With the improved assay, quantification of condensed tannins in water-soluble forms was highly replicable. The relative abundance of condensed tannins in soluble and insoluble forms differed substantially between tissue types.</AbstractText><AbstractText Label="CONCLUSIONS" NlmCategory="CONCLUSIONS">The quantification of condensed tannins using the butanol-HCl assay was improved by adding acetone to both extraction and reagent solutions. These improvements will facilitate the quantification of total condensed tannin in tissues containing a range of concentrations, as well as to determine the amount in water-soluble, acetone:MeOH-soluble and insoluble forms. Accurate determination of these three condensed tannin forms is essential for careful investigations of their potentially different physiological and ecological functions.</AbstractText></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Shay</LastName><ForeName>Philip-Edouard</ForeName><Initials>PE</Initials><AffiliationInfo><Affiliation>Department of Biology & Centre for Forest Biology, University of Victoria, P.O. Box 3020, STN CSC, Victoria, BC V8W 3N5 Canada.</Affiliation><Identifier Source="ISNI">0000 0004 1936 9465</Identifier><Identifier Source="GRID">grid.143640.4</Identifier></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Trofymow</LastName><ForeName>J A</ForeName><Initials>JA</Initials><AffiliationInfo><Affiliation>Department of Biology & Centre for Forest Biology, University of Victoria, P.O. Box 3020, STN CSC, Victoria, BC V8W 3N5 Canada.</Affiliation><Identifier Source="ISNI">0000 0004 1936 9465</Identifier><Identifier Source="GRID">grid.143640.4</Identifier></AffiliationInfo><AffiliationInfo><Affiliation>Pacific Forestry Centre, Canadian Forest Service, Natural Resources Canada, Victoria, BC Canada.</Affiliation><Identifier Source="ISNI">0000 0001 2295 5236</Identifier><Identifier Source="GRID">grid.202033.0</Identifier></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Constabel</LastName><ForeName>C Peter</ForeName><Initials>CP</Initials><AffiliationInfo><Affiliation>Department of Biology & Centre for Forest Biology, University of Victoria, P.O. Box 3020, STN CSC, Victoria, BC V8W 3N5 Canada.</Affiliation><Identifier Source="ISNI">0000 0004 1936 9465</Identifier><Identifier Source="GRID">grid.143640.4</Identifier></AffiliationInfo></Author></AuthorList><Language>eng</Language><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType></PublicationTypeList><ArticleDate DateType="Electronic"><Year>2017</Year><Month>08</Month><Day>02</Day></ArticleDate></Article><MedlineJournalInfo><Country>England</Country><MedlineTA>Plant Methods</MedlineTA><NlmUniqueID>101245798</NlmUniqueID><ISSNLinking>1746-4811</ISSNLinking></MedlineJournalInfo><KeywordList Owner="NOTNLM"><Keyword MajorTopicYN="N">Douglas-fir</Keyword><Keyword MajorTopicYN="N">Flavonoid</Keyword><Keyword MajorTopicYN="N">Leaf litter</Keyword><Keyword MajorTopicYN="N">Nitrogen stress</Keyword><Keyword MajorTopicYN="N">Polyphenol</Keyword><Keyword MajorTopicYN="N">Poplar</Keyword><Keyword MajorTopicYN="N">Proanthocyanidins</Keyword><Keyword 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