Elsevier

Talanta

Volume 176, 1 January 2018, Pages 344-349
Talanta

Rapid and sensitive serum glucose determination using chemical labeling coupled with black phosphorus-assisted laser desorption/ionization time-of-flight mass spectrometry

https://doi.org/10.1016/j.talanta.2017.08.055Get rights and content

Highlights

  • BBII was synthesized as a chemical labeling reagent for labeling glucose used in BP/ALDI-TOF MS.

  • The entire analysis of serum glucose was completed within 5 min.

  • Endogenous glucose was detected in small volume (0.5 μL) of blood serum.

  • The results were consistent with those obtained using the hexokinase method in a clinical laboratory.

Abstract

Monitoring the concentration of blood glucose in patients is a key component of good medical diagnoses. Therefore, developing an accurate, rapid and sensitive strategy for monitoring blood glucose is of vital importance. We proposed a strategy for serum glucose determination combining 2-(4-boronobenzyl) isoquinolin-2-ium bromide chemical labeling with black phosphorus assisted laser desorption ionization-time of flight mass spectrometry (CL-BP/ALDI-TOF MS). The entire analytical process consisted of 1 min of protein precipitation and 3 min of chemical labeling in a microwave oven prior to the BP/ALDI-TOF MS analysis. The analysis can be completed in 5 min with high throughput and extremely low sample consumption. Good linearity for glucose was obtained with a correlation coefficient (R) of 0.9986. The limit of detection (LOD) and limit of quantification (LOQ) were 11.5 fmol and 37.5 fmol, respectively. Satisfied reproducibility and reliability were gained by evaluation of the intra- and inter-day precisions with relative standard deviations (RSDs) less than 7.2% and relative recoveries ranging from 87.1% to 108.1%, respectively. The proposed strategy was also applied for the analysis of endogenous glucose in various serum samples and the results were consistent with those obtained using the hexokinase method in a clinical laboratory. Considering the results, the proposed CL-BP/ALDI-TOF MS strategy has proven to be reliable, fast, and sensitive for quantitative analysis of serum glucose.

Introduction

Glucose is a hexose that serves as the primary energy source for living cells in the human body, and its concentration levels in serum are widely recognized as a critical clinical indicator of diabetes mellitus [1]. Over time, the presence of too much glucose in the blood can lead to severe secondary complications, such as retinopathy, nephropathy, neuropathy, and cardiovascular disease [2]. In addition, emerging research has shown that the serum glucose level is closely related to pancreatic, esophagus, liver, and colon/rectum cancers [3], indicating that serum glucose may be a potential biomarker for monitoring the development of cancer. Therefore, numerous methods have been investigated for blood glucose detection. In recent years, electrochemically or optically developed glucose sensors [4], [5] are commonly used due to their low-cost and portability advantages. However, low detection sensitivity and poor anti-interference capacity of these glucose-sensors seriously restrict their practical applications. In this regard, gas chromatography-mass spectrometry (GC-MS) [6] and liquid chromatography-mass spectrometry (LC-MS) [7], [8], [9], [10] approaches are introduced for blood glucose analysis because of their high accuracy and high sensitivity. However, it may be time-consuming because a gas- or liquid-phase separation process is required. The limitations of the current technology highlight the need for an accurate, sensitive and rapid method for the determination of blood glucose to produce convincing clinical measurements.

Matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) has become a crucial tool in the analysis of various macromolecules [11] due to its advantages of high throughput, high sensitivity, and low sample consumption [12]. However, very few MALDI MS methods [13], [14] have been reported for the analysis of glucose in blood because of background signal interferences in the small molecular region (< 700 Da) from organic matrices [15] Recently, our group proved black phosphorus as a new matrix for MALDI MS qualification and quantification of small molecule [16]. The research showed that black phosphorus assisted laser desorption/ionization-time of flight mass spectrometry (BP/ALDI-TOF MS) could produce a clean background during the analysis of small molecules. Moreover, the technique facilitated high detection sensitivity towards quaternary ammonium compounds. On the other hand, boronic acids can covalently bind with 1,2 or 1,3 cis-diols to form five- or six-membered cyclic esters [17], indicating that boronic acids can selectively bind to the cis-diols pairs of glucose. It inspired us to explore a reaction pathway for converting glucose to a quaternary ammonium compound via a chemical labeling reagent containing a boronic acid group and a quaternary ammonium moiety used in BP/ALDI-TOF MS detection. This may be a promising strategy for accurate, high–throughput, and sensitive measurements of glucose in clinical diagnosis applications.

Herein, a compound of 2-(4-boronobenzyl) isoquinolin-2-ium bromide (BBII) containing both boronic acid group and quanternary ammonium group was synthesized as the chemical labeling (CL) reagent. The CL-BP/ALDI-TOF MS strategy enabled a simple and high–throughput sample preparation procedure consisting a 1 min protein precipitation and a 3 min chemical labeling step before the MALDI MS analysis. In addition, due to the superior sensitivity of the detection method, glucose was successfully detected with a miniscule sample of 0.5 μL serum. With the proposed method, endogenous glucose in various serum samples were determined and quantified, and the results were in agreement with those obtained using the conventional clinical hexokinase method.

Section snippets

Chemicals and reagents

Glucose standard, ammonia hydrate (NH3·H2O, 25%, aqueous solution), formic acid (FA, 88%) were all obtained from Sinopharm Chemical Reagent (Shanghai, China). D-glucose-4,5,6,6′-d4 used as internal standard (IS), 4-(bromomethyl)phenylboronic acid and isoquinoline were all purchased from J&K Chemicals (Beijing). Acetonitrile (ACN, HPLC grade) was obtained from Tedia Co. (Fairfield, OH, U.S.A.). Purified water was obtained on the Milli-Q system (Milford, MA, USA). BP was purchased from XFNANO

General principle for the strategy

It was anticipated that high throughput, high sensitivity and low sample consumption exhibited by the MALDI MS would lend itself to the development of an accurate, rapid and sensitive method for detecting serum glucose. BP/ALDI-TOF MS has superior sensitivity for quaternary ammonium compounds, and the cis-diols pairs of glucose can bind well with a boronate reagent. Thus, we explored a reaction path way for converting glucose to a quanternary ammonium compound via a chemical labeling reagent

Conclusions

In summary, a reliable and simple CL-BP/ALDI-TOF MS strategy was developed for the rapid and sensitive analysis of glucose in blood serum samples. Endogenous glucose was selectively reacted with BBII and a CL-glucose derivative exhibited a strong response in BP/ALDI-TOF MS analysis. A simple protein precipitation procedure was needed before chemical labeling and MS detection, and the entire analysis of serum glucose was completed within 5 min. Moreover, due to the highly detection sensitivity of

Acknowledgments

The authors thank the National Natural Science Foundation of China (21475098, 21635006, 31670373), the Natural Science Foundation of Hubei Province, China (2014CFA002) and the Fundamental Research Funds for the Central Universities (2042016kf0035).

References (21)

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