Arginase Enzyme Activity in Human Serum as a Marker of Cognitive Function: The Role of Inositol in Combination with Arginine Silicate

Authors: Katie Emerson, Sara Perez-Ojalvo, Jim Komorowski, Danielle Greenberg

Abstract:

The purpose of this study was to evaluate arginase activity levels in response to combinations of an inositol-stabilized arginine silicate (ASI; Nitrosigine®), L-arginine, and Inositol. Arginine acts as a vasodilator that promotes increased blood flow resulting in enhanced delivery of oxygen and nutrients to the brain and other tissues. Arginase, found in human serum, catalyzes the conversion of arginine to ornithine and urea, completing the last step in the urea cycle. Decreasing arginase levels maintains arginine and results in increased nitric oxide production. This study aimed to determine the most effective combination of ASI, L-arginine and inositol for minimizing arginase levels and therefore maximize ASI’s effect on cognition. Serum was taken from untreated healthy donors by separation from clotted factors. Arginase activity of serum in the presence or absence of test products was determined (QuantiChrom™, DARG-100, Bioassay Systems, Hayward CA). The remaining ultra-filtrated serum units were harvested and used as the source for the arginase enzyme. ASI alone or combined with varied levels of Inositol were tested as follows: ASI + inositol at 0.25 g, 0.5 g, 0.75 g, or 1.00 g. L-arginine was also tested as a positive control. All tests elicited changes in arginase activity demonstrating the efficacy of the method used. Adding L-arginine to serum from untreated subjects, with or without inositol only had a mild effect. Adding inositol at all levels reduced arginase activity. Adding 0.5 g to the standardized amount of ASI led to the lowest amount of arginase activity as compared to the 0.25 g, 0.75 g or 1.00g doses of inositol or to L-arginine alone. The outcome of this study demonstrates an interaction of the pairing of inositol with ASI on the activity of the enzyme arginase. We found that neither the maximum nor minimum amount of inositol tested in this study led to maximal arginase inhibition. Since the inhibition of arginase activity is desirable for product formulations looking to maintain arginine levels, the most effective amount of inositol was deemed preferred. Subsequent studies suggest this moderate level of inositol in combination with ASI leads to cognitive improvements including reaction time, executive function, and concentration.

Keywords: Arginine, blood flow, colorimetry, urea cycle.

References:

[1] L. Castillo, T. E. Chapman, M. Sanchez, Y. M. Yu, J. F. Burke, A. M. Ajami, J. Vogt, and V. R. Young, “Plasma arginine and citrulline kinetics in adults given adequate and arginine-free diets,” Proceedings of the National Academy of Sciences, vol. 90, no. 16, pp. 7749-7753, 1993.
[2] N. E. Flynn, C. J. Meininger, T. E. Haynes, and G. Wu, “The metabolic basis of arginine nutrition and pharmacotherapy,” Biomedicine & Pharmacotherapy, vol. 56, no. 9, pp. 427-438, 2002/11/01/, 2002.
[3] J. L. Boucher, C. Moali, and J. P. Tenu, “Nitric oxide biosynthesis, nitric oxide synthase inhibitors and arginase competition for L-arginine utilization,” Cellular and Molecular Life Sciences, vol. 55, no. 9, pp. 1015, 1999.
[4] LS-Bio. "Arginase Assay Kit (Colorimetric) LS-K334," https:// www.lsbio.com/assaykits/arginase-assay-kit-colorimetric-ls-k334/334.
[5] G. Wu, and S. M. Morris, Jr., “Arginine metabolism: nitric oxide and beyond,” Biochemical Journal, vol. 336, no. 1, pp. 1-17, 1998.
[6] J. Satriano, “Agmatine: At the Crossroads of the Arginine Pathways,” Annals of the New York Academy of Sciences, vol. 1009, no. 1, pp. 34-43, 2003, 2003
[7] H. Dubey, A. Dubey, K. Gulati, and A. Ray, “Protective effects of L-arginine on cognitive deficits and biochemical parameters in an experimental model of type-2 diabetes mellitus induced Alzheimer's disease in rats,” J Physiol Pharmacol, vol. 73, no. 1, Feb, 2022.
[8] P. Mone, A. Pansini, S. S. Jankauskas, F. Varzideh, U. Kansakar, A. Lombardi, V. Trimarco, S. Frullone, and G. Santulli, “L-Arginine Improves Cognitive Impairment in Hypertensive Frail Older Adults,” Front Cardiovasc Med, vol. 9, pp. 868521, 2022.
[9] M. G. Fleszar, J. Wiśniewski, M. Zboch, D. Diakowska, A. Gamian, and M. Krzystek-Korpacka, “Targeted metabolomic analysis of nitric oxide/L-arginine pathway metabolites in dementia: association with pathology, severity, and structural brain changes,” Scientific Reports, vol. 9, no. 1, 2019.
[10] Y.-q. Zhang, Y.-b. Tang, E. Dammer, J.-r. Liu, Y.-w. Zhao, L. Zhu, R.-j. Ren, H.-z. Chen, G. Wang, and Q. Cheng, “Dysregulated Urinary Arginine Metabolism in Older Adults With Amnestic Mild Cognitive Impairment,” Frontiers in Aging Neuroscience, vol. 11, 2019.
[11] G. Carlomagno, and V. Unfer, “Inositol safety: clinical evidences,” Eur Rev Med Pharmacol Sci, vol. 15, no. 8, pp. 931-6, Aug, 2011.
[12] B. J. Holub, "The Nutritional Significance, Metabolism, and Function of myo-Inositol and Phosphatidylinositol in Health and Disease," pp. 107-141: Springer US, 1982.
[13] L. K. Parthasarathy, L. Ratnam, S. Seelan, C. Tobias, M. F. Casanova, and R. N. Parthasarathy, "Mammalian Inositol 3-phosphate Synthase: Its Role in the Biosynthesis of Brain Inositol and its Clinical Use as a Psychoactive Agent," pp. 293-314: Springer US.
[14] A. Kiani, S. Paolacci, A. Calogero, R. Cannarella, G. Di Renzo, S. Gerli, C. Della Morte, G. Busetto, E. De Berardinis, and F. Del Giudice, “From Myo-inositol to D-chiro-inositol molecular pathways,” Eur. Rev. Med. Pharmacol. Sci, vol. 25, no. 5, pp. 2390, 2021.
[15] PubChem, “PubChem Compound Summary for CID 892, Inositol,” NIH: National Library of Medicine. National Center for Biotechnology Information, 2022.
[16] S. K. Fisher, J. E. Novak, and B. W. Agranoff, “Inositol and higher inositol phosphates in neural tissues: homeostasis, metabolism and functional significance,” Journal of Neurochemistry, vol. 82, no. 4, pp. 736-754, 2002.
[17] C. B. Brink, S. L. Viljoen, S. E. de Kock, D. J. Stein, and B. H. Harvey, “Effects of myo-inositol versus fluoxetine and imipramine pretreatments on serotonin 5HT2A and muscarinic acetylcholine receptors in human neuroblastoma cells,” Metab Brain Dis, vol. 19, no. 1-2, pp. 51-70, Jun, 2004.
[18] C. P. Downes, “Inositol phospholipids and neurotransmitter-receptor signalling mechanisms,” Trends in Neurosciences, vol. 6, pp. 313-316, 1983.
[19] S. J. Park, H. Park, M.-G. Kim, S. Zhang, S. E. Park, S. Kim, and C. Chung, “Inositol Pyrophosphate Metabolism Regulates Presynaptic Vesicle Cycling at Central Synapses,” Science, vol. 23, no. 4, pp. 101000, 2020.
[20] F. Donato, C. M. Alberini, D. Amso, G. Dragoi, A. Dranovsky, and N. S. Newcombe, “The Ontogeny of Hippocampus-Dependent Memories,” The Journal of Neuroscience, vol. 41, no. 5, pp. 920-926, 2021.
[21] N. M. Van Strien, N. L. M. Cappaert, and M. P. Witter, “The anatomy of memory: an interactive overview of the parahippocampal–hippocampal network,” Nature Reviews Neuroscience, vol. 10, no. 4, pp. 272-282, 2009.
[22] D. Kalman, P. D. Harvey, S. Perez Ojalvo, and J. Komorowski, “Randomized Prospective Double-Blind Studies to Evaluate the Cognitive Effects of Inositol-Stabilized Arginine Silicate in Healthy Physically Active Adults,” Nutrients, vol. 8, no. 11, Nov 18, 2016.
[23] J. Heller, and A. Heller, “Loss of Activity or Gain in Stability of Oxidases upon Their Immobilization in Hydrated Silica:  Significance of the Electrostatic Interactions of Surface Arginine Residues at the Entrances of the Reaction Channels,” Journal of the American Chemical Society, vol. 120, no. 19, pp. 4586-4590, 1998.
[24] K. R. Martin, "Silicon: The Health Benefits of a Metalloid," Interrelations between Essential Metal Ions and Human Diseases, Metal Ions in Life Sciences A. Sigel, H. Sigel and R. K. O. Sigel, eds., pp. 451-473, Dordrecht: Springer Netherlands, 2013.
[25] H. Jakubowski, A. Zioła-Frankowska, M. Frankowski, J. Perła-Kaján, H. Refsum, C. A. de Jager, and A. D. Smith, “B Vitamins Prevent Iron-Associated Brain Atrophy and Domain-Specific Effects of Iron, Copper, Aluminum, and Silicon on Cognition in Mild Cognitive Impairment,” Journal of Alzheimer's Disease, vol. 84, pp. 1039-1055, 2021.
[26] S. Gillette-Guyonnet, S. Andrieu, and B. Vellas, “The potential influence of silica present in drinking water on Alzheimer's disease and associated disorders,” The Journal of Nutrition, Health & Aging, vol. 11, no. 2, pp. 119-24, 2007.
[27] J. L. Tartar, D. Kalman, and S. Hewlings, “A Prospective Study Evaluating the Effects of a Nutritional Supplement Intervention on Cognition, Mood States, and Mental Performance in Video Gamers,” Nutrients, vol. 11, no. 10, Oct 1, 2019.
[28] D. S. Kalman, S. Feldman, A. Samson, and D. R. Krieger, “A clinical evaluation to determine the safety, pharmacokinetics, and pharmacodynamics of an inositol-stabilized arginine silicate dietary supplement in healthy adult males,” Clin Pharmacol, vol. 7, pp. 103-9, 2015.
[29] D. Kalman, and D. R. Krieger, A randomized, crossover, double-blind clinical trial to evaluate the pharmacokinetics of Nitrosigine as compared to arginine-HCl in healthy adult males, Nutrition21 Proprietary Data, 2015.
[30] S. M. Morris Jr, “Arginine: beyond protein,” The American journal of clinical nutrition, vol. 83, no. 2, pp. 508S-512S, 2006.
[31] S. M. Morris Jr, “Recent advances in arginine metabolism: roles and regulation of the arginases,” British Journal of Pharmacology, vol. 157, no. 6, pp. 922-930, 2009.
[32] J. Pernow, and C. Jung, “Arginase as a potential target in the treatment of cardiovascular disease: reversal of arginine steal?,” Cardiovascular Research, vol. 98, no. 3, pp. 334-343, 2013.
[33] J. L. Gills, A. Campitelli, M. Jones, S. Paulson, J. R. Myers, E. N. Madero, J. M. Glenn, J. Komorowski, and M. Gray, “Acute Inositol-Stabilized Arginine Silicate Improves Cognitive Outcomes in Healthy Adults,” Nutrients, vol. 13, no. 12, 2021.
[34] R. Sowinski, D. Gonzalez, D. Xing, C. Yoo, V. Jenkins, K. Nottingham, B. Dickerson, M. Humphries, M. Leonard, and J. Ko, “Effects of Inositol-Enhanced Bonded Arginine Silicate Ingestion on Cognitive and Executive Function in Gamers,” Nutrients, vol. 13, no. 11, pp. 3758, 2021.
[35] K. Emerson, S. Perez Ojalvo, and D. Greenberg, “Effects of a Bonded Arginine Silicate Inositol combination (nooLVL®) on Cognitive function in esports Gamers,” The FASEB Journal, vol. 36, no. S1, 2022.
[36] L. Bell, D. Lamport, D. Field, L. Butler, and C. Williams, “Practice effects in nutrition intervention studies with repeated cognitive testing,” Nutrition and Healthy Aging, vol. 4, pp. 309-322, 2018.
[37] E. H. Martínez-Lapiscina, P. Clavero, E. Toledo, R. Estruch, J. Salas-Salvadó, B. San Julián, A. Sanchez-Tainta, E. Ros, C. Valls-Pedret, and M. Á. Martinez-Gonzalez, “Mediterranean diet improves cognition: the PREDIMED-NAVARRA randomised trial,” Journal of Neurology, Neurosurgery & Psychiatry, vol. 84, no. 12, pp. 1318-1325, 2013.