Advertisement

Food Analytical Methods

, Volume 12, Issue 11, pp 2518–2526 | Cite as

Determination of native lactoferrin in milk by HPLC on HiTrapTM Heparin HP column

  • Meixia Chen
  • Fang Wen
  • Yangdong Zhang
  • Peng Li
  • Nan Zheng
  • Jiaqi WangEmail author
Article
  • 44 Downloads

Abstract

Foods supplemented with lactoferrin are an increasing trend owing to its multiple functions. Therefore, a reliable analytical method for the nutritional assessment and quality control of lactoferrin containing foods must be established. A method for the determination of native lactoferrin in milk using a HiTrapTM Heparin HP column coupled with HPLC was developed and validated. The conditions for lactoferrin enrichment in milk using the HiTrapTM Heparin HP column were optimized. A linear range of 2 to 100 mg L-1 was obtained for lactoferrin standards, with a R2 value of 0.9989. The detection limit (LOD) and quantification limit (LOQ) of the lactoferrin standards were 2.00 and 6.67 mg L-1, respectively. Considering the enrichment factor involved in the pre-treatment procedures, LOD and LOQ for detection of milk sample were 0.57 and 1.90 mg L-1, respectively. In addition, milk samples spiked with lactoferrin at three concentration levels (2, 5, and 10 mg/L) showed overall mean recovery of 88.3%, 90.2% and 95.1%, respectively. The relative standard deviations of intra-day and inter-day precision were 1.3% ~ 4.8%, and 2.1% ~ 5.7%, respectively, indicating that the present method showed good performances. Furthermore, the successful quantification of lactoferrin in raw milk and processed milk demonstrated the usefulness and effectiveness of the proposed method.

Keywords

lactoferrin heparin HPLC milk enrichment 

Notes

Acknowledgement

This work was supported by a grant from the Agricultural Science and Technology Innovation Program (ASTIP-IAS12), Special Fund for Agro-scientific Research in the Public Interest (201403071) and Modern Agro-Industry Technology Research System of the PR China (CARS-36). We thank Simon Partridge, PhD, from Liwen Bianji, Edanz Editing China (www.liwenbianji.cn/ac), for editing the English text of a draft of this manuscript.

Compliance with ethical standards

Conflicts of Interest

The authors declare no conflicts of interest.

Ethical approval

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

Informed consent

Not applicable.

Supplementary material

12161_2019_1572_MOESM1_ESM.docx (863 kb)
ESM 1 (DOCX 863 kb)

References

  1. Anderson BF, Baker HM, Dodson EJ, Norris GE, Rumball SV, Waters JM, Baker EN (1987) Structure of human lactoferrin at 3.2-A resolution. Proc Natl Acad Sci 84(7):1769–1773PubMedCrossRefGoogle Scholar
  2. Baker EN (2005) LactoferrinLactoferrin: A multi-tasking protein par excellence. Cell Mol Life Sci:Cmls 62(22):2529–2530PubMedCrossRefGoogle Scholar
  3. Bordin G, Raposo FC, Calle BDL, Rodriguez AR (2001) Identification and quantification of major bovine milk proteins by liquid chromatography. J Chromatogr A 928(1):63–76PubMedCrossRefGoogle Scholar
  4. Brock JH (2002) The physiology of lactoferrin. Biochem Cell Biol 80(80):1–6PubMedCrossRefGoogle Scholar
  5. Capila I, Linhardt RJ (2002) Heparin–Protein Interactions. Angew Chem Int Ed 41(3):390–412CrossRefGoogle Scholar
  6. Conneely OM (2001) Antiinflammatory activities of lactoferrin. J Am Coll Nutr 20(5 Suppl):389S–395SPubMedCrossRefGoogle Scholar
  7. Ding X, Yang Y, Zhao S, Li Y, Wang Z (2011) Analysis of α-lactalbumin, β-lactoglobulin A and B in whey protein powder, colostrum, raw milk, and infant formula by CE and LC. Dairy Sci Technol 91(2):213–225CrossRefGoogle Scholar
  8. Elagamy EI (2000) Effect of heat treatment on camel milk proteins with respect to antimicrobial factors: a comparison with cows' and buffalo milk proteins. Food Chem 68(2):227–232CrossRefGoogle Scholar
  9. Finete VDLM, Gouvêa MM, Netto ADP (2015) Validation of a method of high performance liquid chromatography with fluorescence detection for melamine determination in UHT whole bovine milk. Food Control 51:402–407CrossRefGoogle Scholar
  10. Fox P, Uniacke-Lowe T, McSweeney P, and O’Mahony J 2015 Biologically Active Compounds in Milk. Pages 415-497 in Dairy Chemistry and Biochemistry. Springer.Google Scholar
  11. Ghafari S, Aziz HA, Isa MH, Zinatizadeh AA (2009) Application of response surface methodology (RSM) to optimize coagulation–flocculation treatment of leachate using poly-aluminum chloride (PAC) and alum. J Hazard Mater 163(2–3):650–656PubMedCrossRefGoogle Scholar
  12. Hayashida K, Kaneko T, Takeuchi T, Shimizu H, Ando K, Harada E (2004) Oral administration of lactoferrin inhibits inflammation and nociception in rat adjuvant-induced arthritis. J Vet Med Sci 66(2):149–154PubMedCrossRefGoogle Scholar
  13. Hetherington SV, Spitznagel JK, Quie PG (1984) An enzyme-linked immunoassay (ELISA) for measurement of lactoferrin. J Immunol Methods 65(1-2):183–190CrossRefGoogle Scholar
  14. Legrand D, Mazurier J (2010) A critical review of the roles of host lactoferrin in immunity. Biometals: An International Journal on the Role of Metal Ions in Biology Biochemistry & Medicine 23(3):365–376CrossRefGoogle Scholar
  15. Levay PF, Viljoen M (1995) Lactoferrin: a general review. Haematologica 80(3):252–267PubMedGoogle Scholar
  16. Li H, Zhao G, Niu S, Luan Y (2007) Technologic parameter optimization of gas quenching process using response surface method. Comput Mater Sci 38(4):561–570CrossRefGoogle Scholar
  17. Li J, Ding X, Chen Y, Song B, Zhao S, Wang Z (2012) Determination of bovine lactoferrin in infant formula by capillary electrophoresis with ultraviolet detection. J Chromatogr A 1244(12):178–183PubMedCrossRefGoogle Scholar
  18. Maran JP, Manikandan S (2012) Response surface modeling and optimization of process parameters for aqueous extraction of pigments from prickly pear ( Opuntia ficus-indica ) fruit. Dyes Pigments 95(3):465–472CrossRefGoogle Scholar
  19. Mata L, Sanchez L, Headon DR, Calvo M (1998) Thermal denaturation of human lactoferrin and its effect on the ability to bind iron. J Agric Food Chem 46(10):3964–3970CrossRefGoogle Scholar
  20. Murphy ME, Kariwa H, Mizutani T, Tanabe H, Yoshimatsu K, Arikawa J, Takashima I (2001) Characterization of in vitro and in vivo antiviral activity of lactoferrin and ribavirin upon hantavirus. J Vet Med Sci 63(6):637–645PubMedCrossRefGoogle Scholar
  21. Nithipatikom K, Mcgown LB (1987) Homogeneous immunochemical technique for determination of human lactoferrin using excitation transfer and phase-resolved fluorometry. Anal Chem 59(3):423–427PubMedCrossRefGoogle Scholar
  22. Ounis WB, Gauthier SF, Turgeon SL, Roufik S, Pouliot Y (2008) Separation of minor protein components from whey protein isolates by heparin affinity chromatography. Int Dairy J 18(10–11):1043–1050CrossRefGoogle Scholar
  23. Palmano KP, Elgar DF (2002) Detection and quantitation of lactoferrin in bovine whey samples by reversed-phase high-performance liquid chromatography on polystyrene-divinylbenzene. J Chromatogr A 947(2):307–311CrossRefGoogle Scholar
  24. Pellegrino LM 2005 ISO International Standard 13875:2005 - Liquid milk - Determination of acid soluble beta-lactoglobulin content - Reverse phase HPLC method. International Organization for Standardization.Google Scholar
  25. Riechel P, Weiss T, Weiss M, Ulber R, Buchholz H, Scheper T (1998) Determination of the minor whey protein bovine lactoferrin in cheese whey concentrates with capillary electrophoresis. J Chromatogr A 817(1–2):187–193PubMedCrossRefGoogle Scholar
  26. Sandomirsky BP, Galchenko SE, Galchenko KS (2003) Antioxidative properties of lactoferrin from bovine colostrum before and after its lyophilization. Cryo-Lett 24(5):275–280Google Scholar
  27. Sun GQ, Kang XH, Liu WX, and Xin-Yu HU. 2009. Detection of lactoferrin in milk product by capillary electrophoresis. Food Research & Development.Google Scholar
  28. Takakura N, Wakabayashi H, Ishibashi H, Teraguchi S, Tamura Y, Yamaguchi H, Abe S (2003) Oral Lactoferrin Treatment of Experimental Oral Candidiasis in Mice. Antimicrob Agents Chemother 47(8):2619–2623PubMedPubMedCentralCrossRefGoogle Scholar
  29. Tomassetti M, Martini E, Campanella L, Favero G, Sanzò G, Mazzei F (2013) Lactoferrin determination using flow or batch immunosensor surface plasmon resonance: Comparison with amperometric and screen-printed immunosensor methods. Sensors Actuators B Chem 179(2):215–225CrossRefGoogle Scholar
  30. Tsakali E, Petrotos K, Chatzilazarou A, Stamatopoulos K, D’Alessandro AG, Goulas P, Massouras T, Impe JFMV (2014) Short communication: Determination of lactoferrin in Feta cheese whey with reversed-phase high-performance liquid chromatography. J Dairy Sci 97(8):4832–4837PubMedCrossRefGoogle Scholar
  31. Tsuda H, Sekine K, Fujita K, Ligo M (2002) Cancer prevention by bovine lactoferrin and underlying mechanisms--a review of experimental and clinical studies. Biochem Cell Biol 80(1):131–136PubMedCrossRefGoogle Scholar
  32. Van Berkel PH, Geerts ME, van Veen HA, Mericskay M, de Boer HA, Nuijens JH (1997) N-terminal stretch Arg2, Arg3, Arg4 and Arg5 of human lactoferrin is essential for binding to heparin, bacterial lipopolysaccharide, human lysozyme and DNA. Biochem J 328(Pt 11):145–151PubMedPubMedCentralCrossRefGoogle Scholar
  33. Wen H, An X, Hai N, Zhen J, Wang J (2009) Optimised ultrasonic-assisted extraction of flavonoids from Folium eucommiae and evaluation of antioxidant activity in multi-test systems in vitro. Food Chem 114(3):1147–1154CrossRefGoogle Scholar
  34. Yuana GU, Cheng LH, Jiang JD, Yang JB, Qian WB, Jing-Lanab WU, and Ying HJ. 2011. Determination of Lactoferrin in infant formula milk powder by Capillary Electrophoresis. China Dairy Industry.Google Scholar
  35. Zhang J, Lai S, Cai Z, Chen Q, Huang B, Ren Y (2014) Determination of bovine lactoferrin in dairy products by ultra-high performance liquid chromatography-tandem mass spectrometry based on tryptic signature peptides employing an isotope-labeled winged peptide as internal standard. Anal Chim Acta 829:33–39PubMedCrossRefGoogle Scholar
  36. Zhang Y 2005a. The application of lactoferrin in infant formula milk powder. Dairy Guide.Google Scholar
  37. Zhang YJ 2005b. Function of lactoferrin and the application of lactoferrin in infant formula milk powder. China Dairy Industry.Google Scholar
  38. Zou S, Hurley W (1992) Effect of ph and some cations on binding of heparin to lactoferrin. J Nanjing Agr UnivGoogle Scholar
  39. Zou S, Magura CE, Hurley WL (1992) Heparin-binding properties of lactoferrin and lysozyme. Comp Biochem Physiol B Comp Biochem 103(4):889–895CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Meixia Chen
    • 1
    • 2
    • 3
  • Fang Wen
    • 1
    • 2
    • 3
  • Yangdong Zhang
    • 1
    • 2
    • 3
  • Peng Li
    • 1
    • 2
    • 3
  • Nan Zheng
    • 1
    • 2
    • 3
  • Jiaqi Wang
    • 1
    • 2
    • 3
    Email author
  1. 1.Key Laboratory of Quality & Safety Control for Milk and Dairy Products of Ministry of Agriculture and Rural Affairs, Institute of Animal SciencesChinese Academy of Agricultural SciencesBeijingPeople’s Republic of China
  2. 2.Laboratory of Quality and Safety Risk Assessment for Dairy Products of Ministry of Agriculture and Rural Affairs, Institute of Animal SciencesChinese Academy of Agricultural SciencesBeijingPeople’s Republic of China
  3. 3.State Key Laboratory of Animal Nutrition, Institute of Animal ScienceChinese Academy of Agricultural SciencesBeijingPeople’s Republic of China

Personalised recommendations