Skip to main content
SpringerLink
Log in

Carbohydrate Analysis in Fermentation Samples by Reversed-Phase Liquid Chromatography with Mass Spectrometry Detection Using Precolumn Derivatization with Dibenzylamine

  • Original
  • Published:
Chromatographia Aims and scope Submit manuscript

Abstract

Liquid chromatography–mass spectrometry (LC–MS) technique has been widely used for the determination of trace carbohydrate compounds in biological samples. However, the ionization efficiency of carbohydrates under the conditions of common electrospray ion sources is poor, and thus chemical derivatization treatment is usually needed. Here, a pre-column derivatization based on dibenzylamine method coupling with LC–MS/MS was developed for the analysis of glucose, xylose, ribose, galactose, maltose, and maltotriose. Aldosaccharide compounds were labeled by dibenzylamine within 3 h at 75 °C. The derivatized aldosaccharides were separated on a polar-copolymerized C18 column and identified with electrospray ionization mass spectrometer in positive mode. The dibenzylamine-derived product of 3-O-methyl-D-glucopyranose was used as the internal standard for the quantitation of derivatized aldosaccharide products. Under the optimized condition, good correlation coefficients (R2 > 0.9903) for all carbohydrates were obtained. Outstanding sensitivity with limits of quantitation (S/N 10) of 0.35–20 ng mL−1 and limits of detection (S/N 3) of 0.1–7 ng mL−1 were achieved. The developed method was successfully utilized for the determination of aldosaccharides in sample of corn stalk fermentation. The recoveries of the method were in the range of 92.59–103.22% with relative standard deviations of 1.08–6.08%.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Scheme 1
Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  1. 1.Wang J, Chae M, Beyene D, Sauvageau D, Bressler DC (2021) Co-production of ethanol and cellulose nanocrystals through self-cycling fermentation of wood pulp hydrolysate. Bioresour Technol 330:124969. https://www.ncbi.nlm.nih.gov/pubmed/33740586

  2. Granados-Arvizu JA, Madrigal-Perez LA, Canizal-Garcia M, Gonzalez-Hernandez JC, Garcia-Almendarez BE, Regalado-Gonzalez C (2019) Effect of cytochrome bc(1) complex inhibition during fermentation and growth of Scheffersomyces stipitis using glucose, xylose or arabinose as carbon sources. Fems Yeast Research 19. <Go to ISI>://WOS:000463868300008

  3. Qin W, Zhang Z, Qin R, Han J, Zhao R, Gu Y, Pan Y, Gu J, Ren S (2019) Providing bionic glycome as internal standards by glycan reducing and isotope labeling for reliable and simple quantitation of N-glycome based on MALDI-MS. Anal Chim Acta 1081:112–119. https://www.ncbi.nlm.nih.gov/pubmed/31446948

  4. Costa S, Rugiero I, Pedrini P, Bernardi T, Tamburini E (2017) Simultaneous quantification of carbohydrates and metabolites in multicomponent fermentation broths by means of high-performance thin-layer chromatography. JPC J Planar Chromatogr Modern TLC 30:170–174

    Article  CAS  Google Scholar 

  5. Santos TM, Alonso MV, Oliet M, Dominguez JC, Rigual V, Rodriguez F (2018) Effect of autohydrolysis on Pinus radiata wood for hemicellulose extraction. Carbohydr Polym 194:285–293. https://www.ncbi.nlm.nih.gov/pubmed/29801841

  6. Li G, Zhang H, Lu Y, Xue H (2019) Solid state fermentation process coupled biological pretreatment with cellulase production by piptoporus betulinus for enhanced cellulose hydrolysis. Cellulose 26:3815–3824

    Article  CAS  Google Scholar 

  7. Chen H, Chen H, Lu H, Tang X, Zhang H, Chen YQ, Chen W (2021) Carbohydrate analysis of mortierella alpina by colorimetry and HPLC-ELSD to reveal accumulation differences of sugar and lipid. Biotechnol Lett 43:1289–1301. https://www.ncbi.nlm.nih.gov/pubmed/33864523

  8. McCormick W, Muldoon C, McCrudden D (2021) Electrochemical flow injection analysis for the rapid determination of reducing sugars in potatoes. Food Chem 340:127919. https://www.ncbi.nlm.nih.gov/pubmed/32889210

  9. Rodriguez-Morato J, Pozo OJ, Marcos J (2018) Targeting human urinary metabolome by LC–MS/MS: a review. Bioanalysis 10:489–516. <Go to ISI>://WOS:000433569600008

  10. Wang W, Chen F, Zheng F, Russell BT (2020) Optimization of synthesis of carbohydrates and 1-phenyl-3-methyl-5-pyrazolone (PMP) by response surface methodology (RSM) for improved carbohydrate detection. Food Chem 309:125686. https://www.ncbi.nlm.nih.gov/pubmed/31670130

  11. Chuanxiang W, Lian X, Lijie L, Fengli Q, Zhiwei S, Xianen Z, Jinmao Y (2016) A sensitive and efficient method for determination of N-acetylhexosamines and N-acetylneuraminic acid in breast milk and milk-based products by high-performance liquid chromatography via UV detection and mass spectrometry identification. J Chromatogr B Analyt Technol Biomed Life Sci 1011:14–23. https://www.ncbi.nlm.nih.gov/pubmed/26751589

  12. Zhang P, Wang Z, Xie M, Nie W, Huang L (2010) Detection of carbohydrates using a pre-column derivatization reagent 1-(4-isopropyl) phenyl-3-methyl-5-pyrazolone by high-performance liquid chromatography coupled with electrospray ionization mass spectrometry. J Chromatogr B Analyt Technol Biomed Life Sci 878:1135–1144. https://www.ncbi.nlm.nih.gov/pubmed/20382576

  13. Sun Z, Song C, Xia L, Wang X, Suo Y, You J (2010) Comprehensive comparisons between 1-phenyl-3-methyl-5-pyrazolones, 1-(4-methoxyphenyl)-3-methyl-5-pyrazolones and 1-(2-naphthyl)-3-methyl-5-pyrazolones as labeling reagents used in LC-DAD-ESI-MS-MS analysis of neutral aldoses and uronic acids. Chromatographia 71:789–797

    Article  CAS  Google Scholar 

  14. Chen S, Sathuvan M, Zhang X, Zhang W, Tang S, Liu Y, Cheong KL (2021) Characterization of polysaccharides from different species of brown seaweed using saccharide mapping and chromatographic analysis. BMC Chem 15:1. https://www.ncbi.nlm.nih.gov/pubmed/33430936

  15. Galermo AG, Nandita E, Barboza M, Amicucci MJ, Vo TT, Lebrilla CB (2018) Liquid chromatography-tandem mass spectrometry approach for determining glycosidic linkages. Anal Chem 90:13073–13080. https://www.ncbi.nlm.nih.gov/pubmed/30299929

  16. Ruhaak LR, Steenvoorden E, Koeleman CA, Deelder AM, Wuhrer M (2010) 2-picoline-borane: a non-toxic reducing agent for oligosaccharide labeling by reductive amination. Proteomics 10:2330–2336. https://www.ncbi.nlm.nih.gov/pubmed/20391534

  17. Alzweiri M, Al-Marabeh S, Bardaweel SK, Alfar R, Al-Hiari YM (2017) Stability determination for cyclized 2,4-dinitrophenyl hydrazone derivative of glucose. J Anal Sci Technol. https://doi.org/10.1186/s40543-017-0117-x

    Article  Google Scholar 

  18. Uchiyama S, Inaba Y, Kunugita N (2011) Derivatization of carbonyl compounds with 2,4-dinitrophenylhydrazine and their subsequent determination by high-performance liquid chromatography. J Chromatogr B Analyt Technol Biomed Life Sci 879:1282–1289. https://www.ncbi.nlm.nih.gov/pubmed/20970389

  19. Li L, Ma L, Guo Y, Liu W, Wang Y, Liu S (2020) Analysis of oligosaccharides from Panax ginseng by using solid-phase permethylation method combined with ultra-high-performance liquid chromatography-Q-Orbitrap/mass spectrometry. J Ginseng Res 44:775–783. https://www.ncbi.nlm.nih.gov/pubmed/33192120

  20. Silva NGS, Maia TF (2021) Mulinari DR (2021) Effect of acetylation with perchloric acid as catalyst in sugarcane bagasse waste. J Nat Fibers 10(1080/15440478):1875352

    Google Scholar 

  21. Directive EC (2002) 657/EC of 12 August, 1990 on implementing Council Directive 96/23/EC concerning the performance of analytical methods and the interpretation of results. J Eur Communit L221:8–36

    Google Scholar 

  22. Guo Z, Wang C, Liang T, Liang X (2010) Polar-copolymerized approach based on horizontal polymerization on silica surface for preparation of polar-modified stationary phases. J Chromatogr A 1217:4555–4560. https://www.ncbi.nlm.nih.gov/pubmed/20564780

  23. Wei J, Shen A, Yan J, Jin G, Yang B, Guo Z, Zhang F, Liang X (2016) Separation analysis of macrolide antibiotics with good performance on a positively charged C18HCE column. J Sep Sci 39:1073–1081. <Go to ISI>://WOS:000373617100007

  24. Wang C, Guo Z, Zhang J, Zeng J, Zhang X, Liang X (2011) High-performance purification of quaternary alkaloids from Corydalis yanhusuo W. T. Wang using a new polar-copolymerized stationary phase. J Sep Sci 34:53–58. https://www.ncbi.nlm.nih.gov/pubmed/21171176

  25. Ling Z, Xu P, Zhong Z, Wang F, Shu N, Zhang J, Tang X, Liu L, Liu X (2016) Sensitive determination of glucose in Dulbecco’s modified Eagle medium by high-performance liquid chromatography with 1-phenyl-3-methyl-5-pyrazolone derivatization: application to gluconeogenesis studies. Biomed Chromatogr 30:601–605

    Article  CAS  Google Scholar 

  26. Trufelli H, Palma P, Famiglini G, Cappiello A (2011) An overview of matrix effects in liquid chromatography-mass spectrometry. Mass Spectrom Rev 30:491–509. <Go to ISI>://WOS:000289532100007

  27. Gu H, Liu G, Wang J, Aubry A-F, Arnold ME (2014) Selecting the correct weighting factors for linear and quadratic calibration curves with least-squares regression algorithm in bioanalytical LC–MS/MS assays and impacts of using incorrect weighting factors on curve stability, data quality, and assay performance. Anal Chem 86:8959–8966

    Article  CAS  Google Scholar 

Download references

Acknowledgements

This work was funded by the National Natural Science Foundation of China (No. 21505014).

Author information

Authors and Affiliations

  1. School of Chemical Engineering, Dalian University of Technology, Dalian, China

    Zekun Tang, Fanfan Ye, Yuming Song, Qinqin Du & Hua Zhang

  2. Instrumental Analysis Center, Dalian University of Technology, Dalian, China

    Huihui Wan, Rui Cai & Qiang Xu

  3. School of Biological Engineering, Dalian University of Technology, Dalian, China

    Weiwei Fan & Zhanyou Chi

Authors
  1. Zekun Tang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Fanfan Ye
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Huihui Wan
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Yuming Song
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Weiwei Fan
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Zhanyou Chi
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Qinqin Du
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Rui Cai
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Qiang Xu
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Hua Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Huihui Wan or Hua Zhang.

Ethics declarations

Conflict of interest

The authors declare that there is no conflict of interest.

Ethical Approval

This article does not contain any studies with human participants or animals performed by any of the authors.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary Information

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Tang, Z., Ye, F., Wan, H. et al. Carbohydrate Analysis in Fermentation Samples by Reversed-Phase Liquid Chromatography with Mass Spectrometry Detection Using Precolumn Derivatization with Dibenzylamine. Chromatographia 85, 395–403 (2022). https://doi.org/10.1007/s10337-022-04147-x

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10337-022-04147-x

Keywords

Search

Navigation