SPL-M100 Application NoteNo. AS200603EN

Compositional Comparison of Crops byOn-line Solid-phase Derivatization SPE-GC/MSIntroduction

In conventional GC/MS analysis of metabolomics, extraction, lyophilization, and derivatization wereSPL-M100

complicated and time-consuming, and the data obtained tended to be highly inconsistent. We havefor SPE-GC system

dramatically shortened the time required, simplified the process, and achieved high accuracy throughour proprietary “solid-phase derivatization” technology. Amino acids and organic acids are held in theSample

solid phase by ion-exchange interactions, and then passed through acetonitrile to dehydrate andwash the solid phase. In this study, we attempted to compare the composition of vegetables andfruits using the SGI-M100, a fully automated system that performs these processes.

SamplePretreatment flow

On-line solid-phase derivatization

A. Spinach

Dry ice freezing and pulverizing

SPE-GC：SPL-M100

Spinach contains high levels of

100~200 g of edible parts ofcrops were frozen and crushedwith dry ice.

Vial

oxalic acid, which binds withcalcium in the body and cancause stones in the kidneys andurinary tract.

B. TomatoSample 10 g

Automation
Pretreatment：15 min.

Flash-SPE ACXs

Add Water 10 mL

Conditioning
ACN-Water(1/1)
ACN

Load[Retain] Supernatant
50 µL Isolation

Shaking

It is used in ketchup, tomato

Add ACN 20 mL
Shaking extraction 10 min

sauce, pizza sauce, etc.

Centrifuge 3500 rpm, ５ min

Dewater ACN

Supernatant 500 µL

Infiltration Methoxyamine/Pyridine

because of its umami taste due
to its very high concentration ofglutamic acid and its acidity andmoisture content.

C. Plum
It is characterized by its strong
acidity and contains high levels

Washing ACN-水(1/1)

SPD Methoximization, 3 min

Add ACN 500 µL
Shaking 37 ℃, 5 min

Infiltration MSTFA-Toluene(3/1)

Cetrifuge 14000 rpm, 3 min

SPD TMS, 1 min

Add 0.1N NaOH

vegetable・fruit

Information【Sample】
・ Spinach・ Tomato
・ Plum【Target】
・Amino acids
・Amines・Nucleobases,
・organic acids

Elution Hexane

Extract

GC-MS Sprit 1:50

of citric acid and other organicacids.

Analytical condition
PTV Injector
LVI-S250 (AiSTI Science)
Insert Type
Spiral Insert
Injector Temp. 220℃(0.5min)-50℃/min-290℃(16min)
GC-MS
Inlet Mode
Split 1:50Flow Mode
Constant Flow, 1 ml/min
Pre-Column
0.25mm i.d. x 1m
ColumnVf-5ms, 0.25mm i.d. x 30m, df;0.25µm
Oven Temp.
100℃(2min)-10℃/min-220℃-30℃/min-320℃
Trans. Temp. 290℃
MS Method
SCAN, m/z;70-470

SPE-GC-MS system for metabolome analysis
SPL-M100 / GCMS-TQ8040NX

Solid phase extraction to GC-MS injection process（fully automatic process）
Automatic pretreatment time （Purification,Dewater, Derivatization）: 10 min

Time fromderivatization toinjection is always
constantElution・Injection
FlashSPE

Product

SPE-GC system

SPL-M100

Solid phase cartridge

Flash-SPEGC Lage Volume Injection

LVI-S250

GC-MS
SPAD

Conditioning Loading

AiSTI SCIENCE

Washing・Dewater

AiSTI SCIENCE CO.,Ltd.

LVI-S250
Large Volume Injection

Tel：+81-73-475-0033
as@aisti.co.jp

E-mail：as@aisti.co.jp
www.aisti.co.jp

HP：www.aisti.co.jp

AiSTI Application Note

Citric acid-4TMS

Glutamine-3TMS

Citric acid-4TMS
Glutamine-3TMS

Glutamic acid-3TMS

Glutamic acid-3TMS

Asparagine-3TMS

oxalic acid

Citric acid-4TMS

Malic acid-3TMS
GABA-3TMS

Asparagine-3TMS

Malic acid-3TMS

Aspartic acid-3TMS
GABA-3TMS

Malic acid-3TMSAspartic acid-3TMS
GABA-3TMS

Serine-3TMS

Phosphoric acid-3TMS
Phosphoric acid-3TMS

Phosphoric acid-3TMS

Valine-2TMS

Oxalic acid-2TMS■SCAN total ion chromatograms for each crop using this method ■ Comparison of components of each crop【Results and Discussion】

Glutamic acidcitric acidPeak shapes and separations are also excellent for all threesamples in the TIC. It can be inferred that washing,dehydration, and derivatization in the solid phase areefficiently performed. The solid-phase derivatization of thisanalytical method improves the accuracy and shortens thetime required, and the automation of the method is also
expected to increase efficiency. The method is expected tomaximize the advantages of GC/MS, such as high separation,high qualitative capability, and a full database, by overcoming
the complexity and long time required for pretreatment,
which had been a weak point of the method.
In addition, a comparison of the component area of eachsample show that the differences can be clearly determined.
The data obtained from the highly accurate analytical method

is expected to contribute to highly reliable analysis results,
and can be utilized for future improvement of crop varietiesand development of cultivation methods.

HOU:Spinach, TOM:tomate, UME:plum