Development and Validation of a Reversed-Phase Liquid Chromatography Method for the Simultaneous Determination of Indole-3-Acetic Acid, Indole-3-Pyruvic Acid, and Abscisic Acid in Barley (<i>Hordeum vulgare</i>L.)

Drought and salinity stress highly affect the plant growth and production around the world. Secondary metabolites play a main role in adaptation to the environment and in overcoming stress conditions. In order to investigate the effect of drought and salinity, alone or in combination, on secondary metabolism‐related enzyme activities, plant hormones and yield parameters, a greenhouse pot experiment was conducted using two cotton genotypes Zhongmian 23 (salt tolerant) and Zhongmian 41 (salt sensitive). Results showed that single and combined drought and salinity stresses caused remarkable decrease in plant height, bolls and lint yield in the order as follows: D + S > salinity > drought, and Zhongmian 41 > Zhongmian 23. Lower H2O2 and superoxide but higher proline content and secondary metabolism‐related enzyme activities were observed in Zhongmian 23 under drought and salinity, both alone and combined, compared with control in Zhongmian 41. Our findings suggest that controlling reactive oxygen species (ROS) levels and increasing activities of secondary metabolism‐related enzymes in Zhongmian 23 might be an effective mechanism to reduce the negative effects of drought and salinity stress. However, cinnamyl alcohol dehydrogenase (CAD), and shikimate dehydrogenase (SKDH) activities were markedly decreased in Zhongmian 41 under salinity stress alone as compared with control. Meanwhile, Zhongmian 23 had higher expression levels of genes related to secondary metabolism (c.f. phenylalanine ammonia‐lyase, PAL; polyphenol oxidase, PPO and CAD) under the three stresses compared to Zhongmian 41. The content of flavonoids and phenols were significantly enhanced under drought and D + S, with higher accumulation in Zhongmian 23. Phenols content in Zhongmian 23 remained unchanged under salinity as relative to control, but were significantly reduced in Zhongmian 41. In addition, callose content, chitinase activities and abscisic acid (ABA) and Indole‐3‐acetic acid (IAA) were more induced in Zhongmian 23 under drought, salinity and D + S, than in Zhongmian 41. Our results suggest that high tolerance to D + S stress in Zhongmian 23 is closely related to elevated callose, chitinase, flavonoids and phenols contents and higher secondary metabolism‐related enzyme activities and their transcript levels.

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“…Endogenous ABA and IAA concentration was assayed in leaves depending on the methods of Islam et al (2016) and Nakurte et al (2012).…”

Section: Measurement Of Aba and Iaa Contentmentioning

confidence: 99%

Genotypic differences in leaf secondary metabolism, plant hormones and yield under alone and combined stress of drought and salinity in cotton genotypes

Ibrahim

Zhu²,

Chen

et al. 2018

Physiologia Plantarum

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“…We successfully quantified IAA and eight additional IAA‐related compounds from 10 mg of Arabidopsis shoots, which included cotyledons, hypocotyls, and leaves, in a single experiment. In contrast, previous research typically used 10 mg to over 100 mg of plant tissues for the quantification of IAA . Therefore, our quantification method is potent and sensitive enough for the quantification of IAAs.…”

Section: Discussionmentioning

confidence: 94%

“…In contrast, previous research typically used 10 mg to over 100 mg of plant tissues for the quantification of IAA. [34][35][36] Therefore, our quantification method is potent and sensitive enough for the quantification of IAAs.…”

Section: Discussionmentioning

confidence: 98%

Practical optimization of liquid chromatography/mass spectrometry conditions and pretreatment methods toward the sensitive quantification of auxin in plants

Sugahara

Kitao

Yamagaki

et al. 2020

Rapid Comm Mass Spectrometry

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Rationale The plant hormone auxin, indole‐3‐acetic acid, regulates many aspects of plant growth and development. Auxin quantification should offer broad insights into its mechanistic action in plants. However, limited auxin content in plant tissues hampers the establishment of quantification methods without the highest graded instruments or deeply specialized experimental techniques. Methods In this study, we detailed optimized conditions for high‐performance liquid chromatography coupled with triple‐quadrupole mass spectrometry (LC/MS). We compared LC/MS conditions, such as columns, mobile phases, parameters of acquisition methods (selective or multiple ion monitoring), dwell times (DTs), and channel numbers, using differentially mixed authentic auxin and its related compounds. We further investigated pretreatment methods through the optimization of auxin recovery and irrelative compound removal from plant tissues prior to the LC/MS analysis. Results Our LC/MS analysis demonstrated the particular importance of the column, DTs, and channel numbers on detection sensitivity. Our comparative analysis developed optimal pretreatment methods, including the pulverization of plants, concentration of extract through centrifugal evaporation, and removal of irrelative metabolites using liquid–liquid extraction and a spin filter. We injected plant samples into our LC/MS system, quantified auxin and eight related compounds in a single measurement, and determined the auxin increase in an auxin over‐producing mutant. Conclusions Our practical optimization of LC/MS conditions and pretreatment methods provides detailed experimental processes toward the sensitive quantification of auxin from 10 mg of plant tissue. These processes have not always been clearly documented; therefore, our protocol could broadly contribute to technical advances in plant growth and development research.

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“…ABA was estimated from leaf and de-rinded internode tissues using the method of Nakurte et al (2012). Samples were ground in liquid nitrogen and extracted with 80% methanol.…”

Section: Abscisic Acid Extractionmentioning

confidence: 99%

Role of ABA Signaling in Regulation of Stem Sugar Metabolism and Transport under Post- Flowering Drought Stress in Sweet Sorghum

et al. 2019

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Development and Validation of a Reversed-Phase Liquid Chromatography Method for the Simultaneous Determination of Indole-3-Acetic Acid, Indole-3-Pyruvic Acid, and Abscisic Acid in Barley (Hordeum vulgareL.)

Cited by 44 publications

References 21 publications

Genotypic differences in leaf secondary metabolism, plant hormones and yield under alone and combined stress of drought and salinity in cotton genotypes

Genotypic differences in leaf secondary metabolism, plant hormones and yield under alone and combined stress of drought and salinity in cotton genotypes

Practical optimization of liquid chromatography/mass spectrometry conditions and pretreatment methods toward the sensitive quantification of auxin in plants

Role of ABA Signaling in Regulation of Stem Sugar Metabolism and Transport under Post- Flowering Drought Stress in Sweet Sorghum

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