European Journal of Nutrition

, Volume 56, Issue 3, pp 973–979 | Cite as

Fenugreek increases insulin-stimulated creatine content in L6C11 muscle myotubes

  • Kristyen A. Tomcik
  • William J. Smiles
  • Donny M. Camera
  • Helmut M. Hügel
  • John A. HawleyEmail author
  • Rani Watts
Original Contribution



Creatine uptake by muscle cells is increased in the presence of insulin. Accordingly, compounds with insulin-like actions may also augment creatine uptake. The aim of this study was to investigate whether Trigonella foenum-graecum (fenugreek), an insulin mimetic, increases total intracellular creatine levels in vitro.


Total cellular creatine content was measured fluorometrically in L6C11 muscle myotubes treated for 1, 4, and 24 h with 0.5 mM creatine (CR), CR and 20 μg/mL fenugreek seed extract (CR + FEN), CR and 100 nM insulin (CR + INS), and CR + INS + FEN (n = 6 per treatment group). Alterations in the expression of the sodium- and chloride-dependent creatine transporter, SLC6A8, and key signaling proteins in the PI3-K/Akt pathway were determined.


Compared to control (CON), CR + INS + FEN increased total creatine content after 4 h (P < 0.05), whereas all conditions increased SLC6A8 protein expression above CON at this time (P < 0.05). Changes in insulin signaling were demonstrated via increases in AktThr308 phosphorylation, with CR + INS > CON and CR at 1 h (P < 0.05) and with CR + INS + FEN > CON, CR, and CR + INS at 4 h (P < 0.05). In contrast, no changes in PKCζ/λ or GLUT4 phosphorylation were detected.


Fenugreek, when combined with insulin, modulates creatine content via a mechanism which is independent of the activity of SLC6A8, suggesting that an alternative mechanism is responsible for the regulation and facilitation of insulin-mediated creatine uptake in skeletal muscle cells.


Creatine content Creatine transport Fenugreek supplement Insulin signaling 



The authors of this paper would like to thank A/Prof. Theo Macrides and the RMIT Natural Products Research Group (Dr. Lynn Hodges, Mr. Donald Harney, and Ms. Nicolette Kalafatis) as well as Mr. Robert Brkljaca for their invaluable help in the extraction process. We appreciate and thank Mr. Frank Antolasic for the analysis of the fenugreek extract samples. Additionally, we would like to thank Mr. Griffin D’costa for his input and valuable discussions about fenugreek and cell culture technique.

Compliance with ethical standards

Conflict of interest

On behalf of all authors, the corresponding author states that there is no conflict of interest.

Supplementary material

394_2015_1145_MOESM1_ESM.jpg (62 kb)
Figure 1 A liquid chromatography-mass spectroscopy (LC-MS) method was developed and validated for the analysis of (2S,3R,4S)- 4-hydroxyisoleucine (4-HIL) in fenugreek seed extract. A- A representative analytical scale of separation and abundance for fenugreek seed extract run on an Electrospray Ionization ESI mass spectrometer. Extract analysis was performed by monitoring the area under the 4-HIL peak at mass/charge of 148 which corresponds to the [4HIL+1] parent ion in the mass spectrum in each extract sample. B- A comparison of the percentage of 4-HIL per extract by weight. A calibration curve was created from 4-HIL standards and fenugreek extract 4-HIL samples. The weight of the 4-HIL content from each fenugreek extract sample was determined using area under the curve and calculated as a percentage. (JPEG 62 kb)


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Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Kristyen A. Tomcik
    • 1
  • William J. Smiles
    • 1
  • Donny M. Camera
    • 1
  • Helmut M. Hügel
    • 2
  • John A. Hawley
    • 1
    • 3
    Email author
  • Rani Watts
    • 1
  1. 1.Centre for Exercise and Nutrition, Mary MacKillop Institute for Health ResearchAustralian Catholic UniversityMelbourneAustralia
  2. 2.School of Applied Sciences and Health Innovations Research InstituteRMIT UniversityMelbourneAustralia
  3. 3.Research Institute for Sports and Exercise SciencesLiverpool John Moores UniversityLiverpoolUK

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