Summary of Study ST003342

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR002078. The data can be accessed directly via it's Project DOI: 10.21228/M8124D This work is supported by NIH grant, U2C- DK119886.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

Study ID	ST003342
Study Title	Glucosinolate profile and specifier protein activity determine the glucosinolate hydrolysis product formation in kohlrabi (Brassica oleracea var. gongylodes) in a tissue-specific way
Study Summary	Glucosinolates, commonly found in Brassica vegetables, are hydrolyzed by myrosinase to form bioactive isothiocyanates unless specifier proteins redirect the degradation to less bioactive nitriles and epithionitriles. Here, the tissue-specific impact of specifier proteins on the outcome of glucosinolate hydrolysis in nine kohlrabi tissues was investigated. Glucosinolate and their hydrolysis product profiles, epithiospecifier protein and myrosinase activity and protein abundance patterns of key glucosinolate biosynthesis, transport and hydrolysis proteins were determined and correlated to the metabolites in the kohlrabi tissues. Leaf tissues were rich in glucoraphanin, while bulb tissues contained more glucoerucin. Moreover, a higher proportion of isothiocyanates was formed in leaf stalk, bulb peel, stem and root, tissues with relatively higher ESM1 abundance and a higher ratio of myrosinase activity to ESP activity. This study shows that tissue-specific glucosinolate hydrolysis is mediated by glucosinolate biosynthesis and profiles, myrosinase activity and specifier protein as well as modifier protein abundance.
Institute	Leibniz Institute of Vegetable and Ornamental Crops (IGZ) e.V.
Last Name	Hanschen
First Name	Franziska
Address	Theodor-Echtermeyer-Weg 1, 14979 Großbeeren, Brandenburg, Germany
Email	hanschen@igzev.de
Phone	00493370178250
Submit Date	2024-07-12
Raw Data Available	Yes
Raw Data File Type(s)	mzML
Analysis Type Detail	GC-MS
Release Date	2024-11-15
Release Version	1

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Project:

Project ID:	PR002078
Project DOI:	doi: 10.21228/M8124D
Project Title:	Glucosinolate profile and specifier protein activity determine the glucosinolate hydrolysis product formation in kohlrabi (Brassica oleracea var. gongylodes) in a tissue-specific way
Project Summary:	Glucosinolates, commonly found in Brassica vegetables, are hydrolyzed by myrosinase to form bioactive isothiocyanates unless specifier proteins redirect the degradation to less bioactive nitriles and epithionitriles. Here, the tissue-specific impact of specifier proteins on the outcome of glucosinolate hydrolysis in nine kohlrabi tissues was investigated. Glucosinolates and their hydrolysis product profiles, epithiospecifier protein and myrosinase activity and protein abundance patterns of key glucosinolate biosynthesis, transport and hydrolysis proteins were determined and correlated to the metabolites in the kohlrabi tissues. Leaf tissues were rich in glucoraphanin, while bulb tissues contained more glucoerucin. Moreover, a higher proportion of isothiocyanates was formed in leaf stalk, bulb peel, stem and root, tissues with relatively higher ESM1 abundance and a higher ratio of myrosinase activity to ESP activity. This study shows that tissue-specific glucosinolate hydrolysis is mediated by glucosinolate biosynthesis and profiles, myrosinase activity and specifier protein as well as modifier protein abundance.
Institute:	Leibniz Institute of Vegetable and Ornamental Crops (IGZ) e.V.
Department:	Plant Quality and Food Security
Last Name:	Hanschen
First Name:	Franziska
Address:	Theodor-Echtermeyer-Weg 1
Email:	hanschen@igzev.de
Phone:	00493370178250
Funding Source:	Leibniz Association
Project Comments:	J16/2017

Subject:

Subject ID:	SU003463
Subject Type:	Plant
Subject Species:	Brassica oleracea
Genotype Strain:	Brassica oleracea var. gongylodes cv. 'Kolibri'
Age Or Age Range:	adult

Factors:

Subject type: Plant; Subject species: Brassica oleracea (Factor headings shown in green)

mb_sample_id	local_sample_id	Sample source	Tissue
SA364633	K25	Brassica plant tissues	Bulb core
SA364634	K22	Brassica plant tissues	Bulb core
SA364635	K23	Brassica plant tissues	Bulb core
SA364636	K24	Brassica plant tissues	Bulb core
SA364637	K21	Brassica plant tissues	Bulb core
SA364638	K304	Brassica plant tissues	Bulb core
SA364639	K303b	Brassica plant tissues	Bulb core
SA364640	K302	Brassica plant tissues	Bulb core
SA364641	K301	Brassica plant tissues	Bulb core
SA364642	K300	Brassica plant tissues	Bulb core
SA364643	K305	Brassica plant tissues	Bulb middle part
SA364644	K306	Brassica plant tissues	Bulb middle part
SA364645	K307	Brassica plant tissues	Bulb middle part
SA364646	K29	Brassica plant tissues	Bulb middle part
SA364647	K28	Brassica plant tissues	Bulb middle part
SA364648	K27	Brassica plant tissues	Bulb middle part
SA364649	K30	Brassica plant tissues	Bulb middle part
SA364650	K26	Brassica plant tissues	Bulb middle part
SA364651	K308	Brassica plant tissues	Bulb middle part
SA364652	K309	Brassica plant tissues	Bulb middle part
SA364653	K310	Brassica plant tissues	Bulb peel
SA364654	K311	Brassica plant tissues	Bulb peel
SA364655	K314	Brassica plant tissues	Bulb peel
SA364656	K312	Brassica plant tissues	Bulb peel
SA364657	K31	Brassica plant tissues	Bulb peel
SA364658	K32	Brassica plant tissues	Bulb peel
SA364659	K33	Brassica plant tissues	Bulb peel
SA364660	K34	Brassica plant tissues	Bulb peel
SA364661	K35	Brassica plant tissues	Bulb peel
SA364662	K313	Brassica plant tissues	Bulb peel
SA364663	K287	Brassica plant tissues	Leaf lamina
SA364664	K288	Brassica plant tissues	Leaf lamina
SA364665	K289	Brassica plant tissues	Leaf lamina
SA364666	K7	Brassica plant tissues	Leaf lamina
SA364667	K6	Brassica plant tissues	Leaf lamina
SA364668	K9	Brassica plant tissues	Leaf lamina
SA364669	K8	Brassica plant tissues	Leaf lamina
SA364670	K10	Brassica plant tissues	Leaf lamina
SA364671	K286	Brassica plant tissues	Leaf lamina
SA364672	K285	Brassica plant tissues	Leaf lamina
SA364673	K11	Brassica plant tissues	Leaf margin
SA364674	K12	Brassica plant tissues	Leaf margin
SA364675	K13	Brassica plant tissues	Leaf margin
SA364676	K294	Brassica plant tissues	Leaf margin
SA364677	K293	Brassica plant tissues	Leaf margin
SA364678	K292	Brassica plant tissues	Leaf margin
SA364679	K291	Brassica plant tissues	Leaf margin
SA364680	K290	Brassica plant tissues	Leaf margin
SA364681	K14	Brassica plant tissues	Leaf margin
SA364682	K15	Brassica plant tissues	Leaf margin
SA364683	K1	Brassica plant tissues	Leaf midvein
SA364684	K284	Brassica plant tissues	Leaf midvein
SA364685	K283	Brassica plant tissues	Leaf midvein
SA364686	K3	Brassica plant tissues	Leaf midvein
SA364687	K4	Brassica plant tissues	Leaf midvein
SA364688	K5	Brassica plant tissues	Leaf midvein
SA364689	K280	Brassica plant tissues	Leaf midvein
SA364690	K2	Brassica plant tissues	Leaf midvein
SA364691	K281	Brassica plant tissues	Leaf midvein
SA364692	K282	Brassica plant tissues	Leaf midvein
SA364693	K18	Brassica plant tissues	Leaf stalk
SA364694	K16	Brassica plant tissues	Leaf stalk
SA364695	K19	Brassica plant tissues	Leaf stalk
SA364696	K20	Brassica plant tissues	Leaf stalk
SA364697	K17	Brassica plant tissues	Leaf stalk
SA364698	K299	Brassica plant tissues	Leaf stalk
SA364699	K298b	Brassica plant tissues	Leaf stalk
SA364700	K297	Brassica plant tissues	Leaf stalk
SA364701	K296	Brassica plant tissues	Leaf stalk
SA364702	K295	Brassica plant tissues	Leaf stalk
SA364703	K323	Brassica plant tissues	Root
SA364704	K324	Brassica plant tissues	Root
SA364705	K321	Brassica plant tissues	Root
SA364706	K320	Brassica plant tissues	Root
SA364707	K322	Brassica plant tissues	Root
SA364708	K44	Brassica plant tissues	Root
SA364709	K43	Brassica plant tissues	Root
SA364710	K45	Brassica plant tissues	Root
SA364711	K41	Brassica plant tissues	Root
SA364712	K42	Brassica plant tissues	Root
SA364713	K37	Brassica plant tissues	Stem
SA364714	K315	Brassica plant tissues	Stem
SA364715	K316	Brassica plant tissues	Stem
SA364716	K317	Brassica plant tissues	Stem
SA364717	K318	Brassica plant tissues	Stem
SA364718	K319	Brassica plant tissues	Stem
SA364719	K38	Brassica plant tissues	Stem
SA364720	K39	Brassica plant tissues	Stem
SA364721	K40	Brassica plant tissues	Stem
SA364722	K36	Brassica plant tissues	Stem

Showing results 1 to 90 of 90

Showing results 1 to 90 of 90

Collection:

Collection ID:	CO003456
Collection Summary:	The kohlrabi plants were harvested at maturity (68 days after sowing). Nine organs were harvested from each kohlrabi plant (Figure 1B). Regarding the leaf tissues, the third youngest leaf was harvested and separated into four parts (leaf midvein, leaf lamina, leaf margin and leaf stalk). Further, three parts of the bulb (bulb core, bulb middle part and bulb peel) the stem and the root were harvested as well (Figure 1B). Five biological replicates for each plant organ were prepared comprising of three pooled plants. The harvested plant organ materials were weighed, put into 20 mL vials (polyvial V; Zinsser Analytics GmbH, Eschborn, Germany), immediately flash frozen in liquid nitrogen, stored at −20 °C and then subsequently freeze-dried. For the homogenization metal balls (8 mm ⌀) were added to each vial and the samples were ground to a fine powder within 2 min using a mixer mill at 30 Hz (MM400, Retsch GmbH, Haan, Germany). The kohlrabi stem sample which was tough, was allowed to cool (at room temperature) and the grinding was repeated to get the fine powder. The samples were then stored at −20 °C until the subsequent experiments. The whole experiment was performed two times independently.
Collection Protocol Filename:	ProtocolKohlrabi.pdf
Sample Type:	Plant

Treatment:

Treatment ID:	TR003472
Treatment Summary:	The seeds of kohlrabi (B. oleracea var. gongylodes cv. Kolibri) obtained from Bejo Samen GmbH, (Sonsbeck, Germany) were grown in seed trays and then transferred to single 15 cm diameter polypropylene pots containing soil (Einheitserde T, Einheitserde Werkverband e.V., Germany) after germination. The plants were grown under controlled growth conditions in a phytochamber (ENGIE Deutschland GmbH, Köln, Germany) with mercury to metal halide conversion lamp (Clean Ace (Daylight) Mogul Base, 6500K CCT/90CRI; EYE Lighting International, Mentor, OH) at the Leibniz Institute of Vegetable and Ornamental Crops in Großbeeren, Germany. The following growth conditions were used: temperature 22 °C to 18 °C (day to night), 12 h photoperiod, light intensity 500 μmol m−2 s−1 and 70 % air humidity. The plants were watered as needed and no fertilizer was added during the growth period.

Treatment ID:

TR003472

Treatment Summary:

The seeds of kohlrabi (B. oleracea var. gongylodes cv. Kolibri) obtained from Bejo Samen GmbH, (Sonsbeck, Germany) were grown in seed trays and then transferred to single 15 cm diameter polypropylene pots containing soil (Einheitserde T, Einheitserde Werkverband e.V., Germany) after germination. The plants were grown under controlled growth conditions in a phytochamber (ENGIE Deutschland GmbH, Köln, Germany) with mercury to metal halide conversion lamp (Clean Ace (Daylight) Mogul Base, 6500K CCT/90CRI; EYE Lighting International, Mentor, OH) at the Leibniz Institute of Vegetable and Ornamental Crops in Großbeeren, Germany. The following growth conditions were used: temperature 22 °C to 18 °C (day to night), 12 h photoperiod, light intensity 500 μmol m−2 s−1 and 70 % air humidity. The plants were watered as needed and no fertilizer was added during the growth period.

Sample Preparation:

Sampleprep ID:	SP003470
Sampleprep Summary:	The volatile glucosinolate (GLS) hydrolysis products were extracted from the homogenized plant material and quantified according to Friedrich, Wermter, Andernach, Witzel, and Hanschen (2022) with small modifications. Briefly, 250 µL of water was added to 25 mg of freeze-dried material and incubated for one hour at room temperature. After incubation, the GLS hydrolysis products were then extracted with methylene chloride three times (with 2 mL in the first and 1.5 mL in the second and third extraction step) in the presence of 0.2 µmol benzonitrile internal standard. After each extraction steps, samples were centrifuged and the methylene chloride phase was then filtered over anhydrous sodium sulfate, collected and finally concentrated under a nitrogen stream to 300 µL. Reference: Friedrich, K., Wermter, N. S., Andernach, L., Witzel, K., & Hanschen, F. S. (2022). Formation of volatile sulfur compounds and S-methyl-L-cysteine sulfoxide in Brassica oleracea vegetables. Food Chemistry, 383, 132544.
Sampleprep Protocol Filename:	ProtocolKohlrabi.pdf

Combined analysis:

Analysis ID	AN005477
Analysis type	MS
Chromatography type	GC
Chromatography system	Agilent 7890A
Column	SGE BPX5 (30m × 0.25mm; 0.25um)
MS Type	EI
MS instrument type	Single quadrupole
MS instrument name	Agilent 5975C
Ion Mode	POSITIVE
Units	µmol/g dry weight of tissue

Chromatography:

Chromatography ID:	CH004160
Chromatography Summary:	The volatile compounds were analyzed by gas chromatography mass spectrometry (GC-MS) system (7890 A GC with 5975C Inert XL MSD, Agilent Technologies Deutschland GmbH, Waldbronn, Germany) by using a SGE BPX5 column (30 m × 0.25 mm × 0.25 μm) (Trajan Scientific Europe Ltd., Victoria, Australia). After splitless injection of 1 µL of sample solution at 190°C, analytes were separated with He as carrier gas (1.8 mL/min) and a temperature gradient starting at 37°C (for 3 min) rising with 9°C/min to 90°C (2 min hold) then to 110°C with 3°C/min, further increased to 210°C with 9°C/min, then to 223°C with 3°C/min, then to 230°C with 9°C/min and finally the GC was heated to 310°C with 35°C/min (6 min hold).
Instrument Name:	Agilent 7890A
Column Name:	SGE BPX5 (30m × 0.25mm; 0.25um)
Column Temperature:	starting at 37°C (for 3 min) rising with 9°C/min to 90°C (2 min hold) then to 110°C with 3°C/min, further increased to 210°C with 9°C/min, then to 223°C with 3°C/min, then to 230°C with 9°C/min and finally the GC was heated to 310°C with 35°C/min (6 min hold)
Flow Gradient:	-
Flow Rate:	-
Solvent A:	-
Solvent B:	-
Chromatography Type:	GC

MS:

MS ID:	MS005203
Analysis ID:	AN005477
Instrument Name:	Agilent 5975C
Instrument Type:	Single quadrupole
MS Type:	EI
MS Comments:	The identification of glucosinolate hydrolysis products was based on comparison of their mass spectra to those of authentic standards and retention time as well as by comparison of mass spectra with literature data (Spencer & Daxenbichler, 1980). The response factors of the compounds relative to the internal standard benzonitrile obtained from calibration curves of authentic standards was used to quantify the hydrolysis products. For any given compound without a standard, the response factor of the chemically most similar compound was used. MassHunter version 10.2 (AgilentTechnologies Deutschland GmbH, Waldbronn, Germany) was used for data analysis and quantification of compounds. Agilent MSD ChemStation data were transformed to MassHunter format using the MassHunter GC/MS Translator B.0705 2479. Reference: Spencer, G. F., & Daxenbichler, M. E. (1980). Gas chromatography‐mass spectrometry of nitriles, isothiocyanates and oxazolidinethiones derived from cruciferous glucosinolates. Journal of the Science of Food and Agriculture, 31, 359-367.
Ion Mode:	POSITIVE
Analysis Protocol File:	ProtocolKohlrabi.pdf