Mass spectrometry detection of L-valine

I. Principles and Technical Characteristics of Mass Spectrometry Detection

As a polar amino acid, L-valine contains amino (-NH₂) and carboxyl (-COOH) groups in its molecular structure. In mass spectrometry detection, it is usually ionized to form charged ions, which are then separated and quantified based on the mass-to-charge ratio (m/z). The detection technology has the following characteristics:

High Sensitivity: Capable of detecting samples as low as the nanogram level, suitable for trace amino acid analysis.

High Specificity: Distinguishes structural analogs through m/z and fragment ion information, avoiding interference from other amino acids.

Quantitative Accuracy: Achieves high-precision quantification combined with the internal standard method, applicable to detection in complex matrices (such as biological samples and fermentation broths).

II. Common Mass Spectrometry Ionization Modes and Detection Strategies

Electrospray Ionization (ESI)

Principle: Under the action of an electric field, the sample solution is atomized to form charged droplets, and gas-phase ions (such as [M+H]⁺ or [M-H]⁻) are generated after solvent evaporation.

Application: L-valine easily forms protonated ions [M+H]⁺ (m/z = 118.1, molecular weight 117.15 Da) in positive ion mode, which can be directly detected by primary mass spectrometry (MS). For structural confirmation, secondary mass spectrometry (MS/MS) can be used to generate characteristic fragments (such as decarboxylated and deaminated fragments) through collision-induced dissociation (CID).

Advantages: Suitable for polar compounds, can be coupled with liquid chromatography (LC) (LC-MS) to achieve integrated separation and detection, reducing matrix interference.

Matrix-Assisted Laser Desorption/Ionization (MALDI)

Principle: After the sample is mixed and crystallized with a matrix (such as α-cyano-4-hydroxycinnamic acid), it is excited by laser irradiation to generate gas-phase ions.

Application: Mainly used for molecular weight determination. The [M+H]⁺ peak can be seen in the MALDI-MS spectrum of L-valine, and the molecular weight can be accurately determined by combining with a calibrator.

Advantages: Simple operation, suitable for rapid detection, especially for molecular weight confirmation of high-purity samples.

III. Molecular Weight Determination Technology and Data Verification

Steps for Accurate Molecular Weight Determination

Sample Preparation: Dissolve L-valine in polar solvents such as water or methanol at a concentration of usually 1-10 μg/mL, and filter through a 0.22 μm membrane to remove impurities.

Mass Spectrometry Parameter Optimization:

ESI positive ion mode: Spray voltage 2.5-4 kV, capillary temperature 250-350℃, sheath gas flow 10-30 arb.

MALDI mode: Select an appropriate matrix, and adjust the laser energy to the threshold that just produces a signal.

Data Collection and Analysis: Obtain molecular ion peaks through primary mass spectrometry, verify the charge state by combining isotope peaks (such as ¹³C labeling), and consider isotope abundance correction for molecular weight calculation.

Mass Spectrometry Technology in Quantitative Analysis

LC-MS/MS Quantification: Adopt the multiple reaction monitoring (MRM) mode, select the parent ion [M+H]⁺ (m/z 118.1) and characteristic daughter ions (such as m/z 86.1, corresponding to decarboxylated fragments), and quantify by the standard curve method (such as using deuterated L-valine as an internal standard). The linear range is usually 0.1-100 μg/mL, and the detection limit can reach 0.01 μg/mL.

IV. Precautions and Interference Factors

Matrix Effect: Proteins, salts, etc., in biological samples (such as serum and cell lysates) will inhibit ionization, requiring pretreatment by solid-phase extraction (SPE) or protein precipitation.

Isomer Differentiation: L-valine and D-valine are isomers of each other and need to be detected by mass spectrometry after separation by chiral chromatography.

Instrument Calibration: Regularly calibrate the mass axis of the mass spectrometer with standards (such as sodium formate) to ensure that the molecular weight determination error is <5 ppm.

V. Technical Application Scenarios

Biomedicine: Quality control of L-valine content in amino acid injections and nutritional supplements.

Fermentation Engineering: Monitoring the synthesis efficiency of L-valine in microbial fermentation broths.

Food Detection: Quantitative analysis of free amino acids in dairy products and grains.

Through the high-resolution and high-sensitivity characteristics of mass spectrometry technology, accurate molecular weight determination and trace quantification of L-valine can be achieved, providing a reliable analytical means for its application in scientific research and industry.