Effect of different additives on amylase activity

Abstract

Enzymes are catalysts of biological origin, and according to their chemical composition, they are simple or complex proteins. There are several theories about the enzyme's mechanism of action. Today, the Michaelis-Menten theory is generally accepted. According to this theory, during enzymatic reactions, an intermediate compound is created between the enzyme and the substrate. After the formation of this complex, the enzyme catalyzes a chemical reaction that changes the substrate into another molecule, which we call the product. The product is then separated and released from the active site of the enzyme, which is then ready to bind the next substrate molecule. Enzyme activity can be affected by different molecules. The purpose of this study is to use the spectrophotometric approach to determine whether sodium benzoate and ascorbic acid (vitamin C) serve as activators or inhibitors of enzymatic reactions. The obtained results show that both additives bind to the enzyme-substrate complex, causing non-competitive inhibition.

Keywords

• Enzymes
• additives
• ascorbic acid
• sodium benzoate
• inhibition

References

1. [1] Bisswanger H. Enzyme Kinetics. Principles and Methods. 2nd Ed., WILEY-VCH Verlag GmbH & Co. KGaA; 2008.
2. [2] Boehlke C, Zierau O, Hannig C. Salivary amylase—the enzyme of unspecialized euryphagous animals. Arch Oral Biol. 2015; 60(8): 1162–1176. https://doi.org/10.1016/j.archoralbio.2015.05.008.
3. [3] Brayer GD, Luo Y, Withers SG. The structure of human pancreatic 𝛼-amylase at 1.8 ˚A resolution and comparisons with related enzymes. Protein Sci. 1995; 4(9): 1730–1742. https://doi.org/10.1002/pro.5560040908.
4. [4] Fujisawa T, Ikegami H, Inoue K, Kawabata Y, Ogihara T. Effect of two 𝛼-glucosidase inhibitors, voglibose and acarbose, on postprandial hyperglycemia correlates with subjective abdominal symptoms. Metabolism. 2005; 54(3): 387–390. https://doi.org/10.1016/j.metabol.2004.10.004.
5. [5] Jayaraj S, Suresh S, Kadeppagari RK. Amylase inhibitors and their biomedical applications. Starch. 2013; 65(7-8): 535–542. https://doi.org/10.1002/star.201200194.
6. [6] Zajácz A, Gyémánt G, Vittori N, Kandra L. Aleppo tannin: structural analysis and salivary amylase inhibition. Carbohydr Res. 2007;342(5):717-723. https://doi.org/10.1016/j.carres.2006.12.016.
7. [7] Luo H, Sun X, Xu L, He W, Liang X. A review on stress determination and control in metal-based additive manufacturing. Theor Appl Mech Lett. 2023; 13(1): 100396. https://doi.org/10.1016/j.taml.2022.100396.
8. [8] Lennerz B, Vafai SB, Delaney NF, Clish CB, Deik AA, Pierce KA, Ludwig DS, Mootha VK. Effects of sodium benzoate, a widely used food preservative, on glucose homeostasis and metabolic profiles in humans. Mol Genet Metab. 2015; 114(1): 73–79. https://doi.org/10.1016/j.ymgme.2014.11.010.

9. [9] Guo J, Gutierrez A, Tan L, Kong L. Inhibitory Effect of ascorbic acid on in vitro enzymatic digestion of raw and cooked starches. Front Nutr. 2021; 8: 758367. https://doi.org/10.3389/fnut.2021.758367.
10. [10] Clissold SP, Edwards C. Acarbose. A preliminary review of its pharmacodynamic and pharmacokinetic properties, and therapeutic potential. Drugs. 1988; 35(3): 214–243. https://doi.org/10.2165/00003495-198835030-00003.
11. [11] Feldmann RJ, Maibach HI. Absorption of some organic compounds through the skin in man. J Invest Dermatol. 1970; 54(5): 399-404. https://doi.org/10.1111/1523-1747.ep12259184.
12. [12] Feillet F, Leonard JV. Alternative pathway therapy for urea cycle disorders. J Inherit Meta Dis. 1998; 21(1): 101-111. https://doi.org/10.1023/a:1005365825875.
13. [13] Kubota K, Horai Y, Kushida K, Ishizaki T. Determination of benzoic acid and hippuric acid in human plasma and urine by high performance liquid chromatography. J Chromatog. 1988; 425(1): 67-75. https://doi.org/10.1016/0378-4347(88)80007-0.
14. [14] Fujii T, Omori T, Tagucji T, Ogata M. Urinary excretion of hippuric acid after administration of sodium benzoate (biological monitoring 1). J Food Hyg. 1991; 32(3): 177-182. https://doi.org/10.1007/BF01805533.
15. [15] Kubota K. Ishizaki T. Dosedependent pharmacokinetics of benzoic acid following oral administration of sodium benzoate to humans. Eur J Clin Pharmacol. 1991; 41(4): 363-368. https://doi.org/10.1007/BF00314969.
16. [16] Matos R, Angelli M, Lago C, Angnes L. Flow injection analysis-amperometric determination of ascorbic and uric acids in urine using arrays of gold microelectrodes modified by electrodeposition of palladium. Anal Chim Acta. 2000; 404: 151-157. https://doi.org/10.1016/S0003-2670(99)00674-1.
17. [17] Akylmaz E, Dinskaya E. A new enzyme electrode based on ascorbate oxidase immobilized in gelatin for specific determination of L-ascorbic acid. Talanta. 1990; 50:87-93. https://doi.org/10.1016/s0039-9140(99)00107-1.
18. [18] Zhu M, Huang X, Li J, Shen H. Peroxidase-based spectrophotometric methods for the determination of ascorbic acid, norepinephrine, epinephrine, dopamine and levodopa. Anal Chim Acta. 1997; 357:261-267. https://doi.org/10.1016/S0003-2670(97)00561-8.
19. [19] Volotovsky V, Kim N. Ascorbic acid determination with an ion-sensitive field effect transistor-based peroxidase biosensor. Anal Chim Acta. 1998; 359(1-2):143-148. https://doi.org/10.1016/S0003-2670(97)00653-3.
20. [20] Shekhovtsova TN, Muginova SV, Luchinina JA, Galimova AZ. Enzymatic methods in food analysis: determination of ascorbic acid. Anal Chim Acta. 2006;573-574:125-132. https://doi.org/10.1016/j.aca.2006.05.015.
21. [21] Tang B, Wang Y, Du M, Ge JC, Chen ZZ. Catalytic spectrophotometric determination of ascorbic acid in tea drink with 1,5-bis(p-hydroxybenzaldene)thiocarbohydrazone as the substrate for horseradish peroxidase. J Agric Food Chem. 2003; 51(15): 4198−4201. https://doi.org/10.1021/jf030012e.
22. [22] Rowe AW, Weill CE. The inhibition of β-amylase by ascorbic acid. Biochim Biophys Acta.1962; 65(2): 245-251. https://doi.org/10.1016/0006-3002(62)91043-0.