Available online on 15.08.2025 at http://jddtonline.info

Journal of Drug Delivery and Therapeutics

Open Access to Pharmaceutical and Medical Research

Copyright  © 2025 The  Author(s): This is an open-access article distributed under the terms of the CC BY-NC 4.0 which permits unrestricted use, distribution, and reproduction in any medium for non-commercial use provided the original author and source are credited

Open Access Full Text Article                                                                             Research Article

Micronutrient Supplementation Augments Adenosine Triphosphate (ATP) production in Caco-2 cells

Gaurav K. Jain ^a*, Pavitra Solanki ^a, Parimal Mahajan ^b, Gajanan Panchal ^b, Maneesha Khalse ^b, Kamlesh Patel ^b

^a Centre for Advanced Formulation Technology, Department of Pharmaceutics, Delhi Pharmaceutical Sciences and Research University, Pushp Vihar, New Delhi 110017, India

^b Medical services, Lupin Ltd. Kalpataru Inspire Off., Western Expressway, Highway, Santacruz East, Mumbai, Maharashtra 400055, India. 

 

 

 

INTRODUCTION

Fatigue, tiredness, exhaustion, and lack of energy is a very common problem for which thorough examination and laboratory tests do not provide a satisfactory explanation¹. Fatigue and energy are interrelated and a common feature of fatigue is a ‘sense of energy depletion’, which can be related to an insufficient amount of cellular energy². Adenosine triphosphate (ATP) produced through oxidative phosphorylation in mitochondria is the source of energy for cellular processes such as metabolic reactions, muscle contraction, active transport, and nerve function³. Recent research has dramatically increased our understanding of the biochemical processes of cellular energy generation and demonstrated that low cellular ATP have effect on cognitive and psychological processes and causes restlessness, tiredness, physical and mental fatigue⁴. The association of low cellular ATP with anxiety, depression, sleep apnea, anemia, insomnia, thyroid and in few cases with genetic disorders has also been established^5-8. Recent studies suggest that limitation in energy production processes limit the rates of energy expenditure and hence performance. Thus, it is expected that a strategy for enhancing cellular ATP could have therapeutic prospects in overcoming physical and mental fatigue^9,10. Micronutrient supplements play key role in a variety of metabolic pathways that support vital cellular and molecular functions¹¹. It has been recognized that a lack of micronutrients may impair cellular energy production, resulting in symptoms of fatigue, tiredness, and lack of energy¹². Theoretically, micronutrient supplement that could increase ATP could also enhance performance and overcome fatigue. Claims regarding the effects of micronutrients particularly B vitamins and minerals on fatigue, cognition and psychological functions are authorized in several reports^13-25 and shown in Figure 1. B vitamins are involved in at least one of the several steps of the energy-production system within the cell and their adequate supply is essential for functioning of the energy-production system. Vitamin B1 (thiamine) is involved in dehydrogenase reactions and helps in formation of acetyl-CoA¹⁴. Vitamin B2 (riboflavin) is an integral part of the coenzymes flavin mononucleotide and flavin adenine dinucleotide and is critical for production of acetyl-CoA via metabolism of proteins, fats and carbohydrates¹⁵. Vitamin B3 (niacin) act as precursor for NAD and NADH, which play important role in citric acid cycle and as electron donors during phosphorylation¹⁶. Vitamin B5 (pantothenic acid) is an essential precursor in the biosynthesis of Coenzyme A¹⁷. The metabolically active form of vitamin B6 (pyridoxine) act as cofactors of enzymes facilitating metabolism of amino acids and lipids, gluconeogenesis, synthesis of niacin and heme and hormone action¹⁸. Vitamin B7 (biotin) serves as a cofactor for several carboxylases enzymes that play key role in energy production and storage through synthesis of fatty acids, catabolism of amino acids and gluconeogenesis¹⁹. Vitamin B9 (folic acid) apart from playing a vital role in synthesis of DNA and RBC is crucial for the synthesis of thymidylate and purines, obligatory for production of cytosolic and mitochondrial ATP, total nucleotide triphosphate (NTP) and deoxy-NTP²⁰. Vitamin B12 (cobalamin) is a cofactor of the methylmalonyl-CoA mutase enzyme which produces succinyl-CoA from methylmalonyl-CoA²¹. 

Minerals such as magnesium has a predominant role in the ATP production and utilization. Magnesium binds to ATP to form biologically functional forms (Mg-ATP complex). In mitochondria Mg-ATP complex. Cellular ATP is present as and this complex act as cofactors for several kinases active during glycolysis help export mitochondrial ATP into cytosol and thus deliver energy within the cell²². Similarly, role of other minerals such as zinc, copper, selenium, chromium picolinate in ATP production through various mechanisms is well documented. Zinc being important component of RNA polymerase plays vital role in protein synthesis and ATP production. Recent in vitro studies showed that zinc is conducive to mitochondrial pyruvate transport, oxidative phosphorylation, lipid, carbohydrate and energy metabolism²³. Copper is a component of cytochrome c oxidase, a vital enzyme in the electron transport chain that produces ATP¹³. Selenium, a component of glutathione peroxidase and selenoproteins, is shown to increase skeletal muscle mitochondrial volume density, mitochondrial biogenesis and metabolism leading to enhanced intracellular ATP²⁴. Chromium picolinate plays a role in regulating carbohydrate, fat and protein metabolism and studies have demonstrated its usefulness in reducing muscle fatigue²⁵. 

Though immense literature exists documenting the role of micronutrients in energy production but most of the reports are theoretical and lacks research data. Additionally, there is mounting evidence regarding toxic effects of long-term micronutrients use²⁶. To date, a vast number of over-the-counter micronutrient supplements containing variable concentrations of vitamins and minerals are being utilized by patients to overcome symptoms of fatigue and tiredness. Therefore, it is crucial to assess the efficacy of ATP production by micronutrient supplements. In this study, we compared the ATP production efficacy of three micronutrient supplements using Caco2 cells, a model of human intestinal epithelial cells. In the past, Caco-2 cells have also been used to study the release and hydrolysis of ATP. ^{27, 28}

 

 

Figure 1: Schematic representation of role of micronutrients in ATP production

 

 

Materials and Methods

Materials

Caco2 cells were purchased from National Centre for Cell Science (Pune, India). Basal cell culture medium, fetal bovine serum (FBS) and phosphate buffer saline (PBS) were purchased from Himedia (Thane, India). Centrifuge tubes (50 ml and 1.5 ml) and 6-well plates were purchased from Tarson (Noida, India). ATP colorimetric assay kit (ab83355, Abcam) was provided by Life Technologies (New Delhi, India). 

Micronutrient Supplements 

Three micronutrient supplements were selected to evaluate their effect on cellular ATP production. First supplement (TV), has therapeutic B complex with magnesium and vit C. Second supplement (TM) has therapeutic B complex with minerals. Third supplement (NM) is a nutraceutical multivitamin and multimineral. The detailed composition of TV, TM and NM is shown in Table 1. All the supplements were provided by Lupin Limited (Mumbai, India).

 

Table 1: Composition of commercial micronutrient supplements used in the study

Supplements	Micronutrient Composition
	TV	TM	NM
Vit B1 (mg)	10	0	1.1
Vit B2 (mg)	10	0	1.3
Vit B3 (mg)	100	100	14
Vit B5 (mg)	5	0	4
Vit B6 (mg)	3	3	1.5
Vit B7 (mcg)	260	0	30
Vit B12 (mcg)	15	15	1
Vit C (mg)	100	0	40
Vit E (mg)	0	0	10
Folic acid (mg)	1.5	1.5	0.117
Mg (mg)	32.4	0	2
Zn sulphate (mg)	0	22.5	10
Selenious acid (mcg)	0	100	30
Copper (mg)	0	0	1.7
Iodine (mcg)	0	0	140
Chromium (mcg)	0	250	30

 

Reagents and Standards

ATP assay buffer and ATP probe in DMSO was used as provided. ATP standard was reconstituted in 100 µL of double distilled water to generate a 10 mM ATP standard stock solution. ATP converter enzyme and developer mix was dissolved in 220 µL assay buffer before use. All the reagents were freshly prepared and kept on ice during the assay. ATP standard stock solution (10 µl) was diluted with dH2O (90 µl) to generate 1 mM ATP standard. The 0, 2, 4, 6, 8, 10 µl of this solution was added into series of wells and volume adjusted to 50 µl/well with assay buffer to generate 0, 2, 4, 6, 8, 10 nmol/well of standard.

Cell culture and treatment

Caco2 cells were cultured in media with 10% FBS and penicillin-streptomycin at 37°C in 5% CO₂ atmosphere (Memmert, Germany). Once the cells were 80% confluent, they were trypsinized and used for the experiments. Cells (0.25 x 10⁶ cells/ mL) were seeded in a 6-well plate and were allowed to grow overnight. Micronutrient supplements at different concentrations (12.5, 25, 50, 100, 200 µg/mL) were added to respective wells. The plates were incubated for different time (treatment time, 30 min, 1 h, 2h, 4 h) at 37°C in 5% CO₂ atmosphere. Well plate without supplement treatment were used as control (CO), whereas cells treated with sodium lauryl sulphate (5 µg/mL) for 30 min were used as negative control (NO). 

Following treatment and incubation, the spent media was removed and the cells were washed with PBS. The cells were resuspended in 100 µL of assay buffer and quickly homogenized by pipetting up and down a few times. The cells were then centrifuged for 5 min at 4°C at 13,000 g in a cold microcentrifuge (Remi, India) to remove insoluble material. The enzymes, if any, were removed from the sample by deproteinizing. The supernatant was collected and transferred to a new tube and kept on ice.

ATP Assay 

ATP reaction mix was prepared by mixing 88% assay buffer, 4% ATP probe, 4% converter enzyme and 4% developer mix. The control was also prepared in same manner; however, 4% converter enzyme was replaced with assay buffer. In the absence of converter enzyme, the assay detects only endogenous glycerol phosphate but not ATP. Briefly, reaction mix (50 µL) was added into each standard, sample, and control wells and the well plates were incubated at room temperature for 30 min protected from light. The readings were measured on a microplate reader (ELX800, Biotek) at 570 nm optical density. All assays were performed in duplicate and controls were run with all the samples. The correction in the absorbance were made by subtracting mean value of the blank from all sample readings. The corrected values for each standard were plotted as a function of the final concentration of ATP. The formula for straight line was used to calculate ATP concentration in the wells. 

Statistical Analysis

All the results were presented as the mean ± standard deviation (SD). Data between the groups was compared using the student t-test. The p < 0.05 was considered statistically significant. All statistical calculations were done using SPSS V26.0 (SPSS Inc., Chicago, USA).

RESULTS

Effect of treatment time 

The amount of ATP produced following treatment of cells with supplements for 30 min, 1 h, 2 h and 4 h is shown in Figure 2. With increase in treatment time from 30 min to 1 h, a significant (p< 0.05) increase in ATP production was observed. Thereafter, increasing the treatment time to 2 h and 4 h, resulted in small but insignificant (p>0.05) increase in ATP. Similar results were obtained for all the three supplements tested and therefore treatment time of 1 h was selected for further experiments. 

Effect of supplement concentration 

The effect of supplement concentration on ATP production was determined. Caco2 cells were treated with supplements at concentration of 12.5, 25, 50, 100, 200 µg/mL. The results are presented in Figure 3. A dose-dependent increase in ATP production was observed for all the supplements. The saturated concentration for TV and TM supplements was 100 µg/mL. Though at 200 µg/mL concentration the ATP production was higher, but the difference was not significant for TV and TM supplements. For NM supplement the ATP production was maximum at 200 µg/mL and the values were significantly higher than those observed at 100 µg/mL. For comparison of supplements the concentration of 50 and 100 µg/mL was selected.

 
 

Figure 2: Effect of treatment time of cells with supplements on ATP production. * p< 0.05, 1h vs 0.5 h; # p>0.05, 2h and 4h vs 1h

 

Figure 3: Effect of supplement concentration on ATP production. * p< 0.05 vs previous concentration and # p>0.05 vs previous concentration

 

Comparison of micronutrient supplements 

Micronutrient supplements (TV, TM and NM) at concentration of 50 and 100 µg/mL were compared for their efficacy to produce cellular ATP Caco2 cells. Control group without treatment and negative control group with SLS treatment was also used for comparison. The amount of ATP produced following treatment as determined by colorimetry is shown in Figure 4. Cells treated with supplements showed markedly higher ATP production compared to control and negative control cells. ATP level in control cells was significantly higher (p<0.05) than negative control cells. Cells treated with TV showed markedly higher ATP level compared to cells treated with TM and NM at both the concentration tested. At 50 µg/mL, ATP level was 1.7-fold and 1.2-fold higher in TV treated cells compared to NM and TM treatment, respectively. Similar results were observed at 100 µg/mL and ATP level was higher in TV treated cells. The decreasing order of ATP production in cells by supplement treatment is as follows: TV > TM > NM. Further, compared to control cells, the cells treated with TV produced nearly 3-fold higher ATP.

 

 

Figure 4: Comparative ATP produced following treatment of cells with control, negative control (NC), and micronutrient supplements (TV, TM and NM) at concentration of 50 and 100 µg/mL and at treatment time of 1 h. * p<0.05, # p<0.001.

 

 

DISCUSSION

Micronutrient supplements are widely available commercially as over the counter products. Most of these micronutrient supplements contains varied concentrations, ratios and proportions of vitamins and minerals, with B-vitamins and minerals being most prominent. The role of micronutrient supplements in cellular ATP production has been well reviewed, however, proof-of-concept studies to validate the same are limited¹³. Though, few studies describe the role of single micronutrient in ATP production^14-18 but the data is poorly elucidated. Thus, we conducted an in vitro studies using Caco2 cells to demonstrate the efficacy of micronutrient supplements in ATP production. Caco2 cells have been used in our study since they are easy to handle and they have been used in past for studying ATP release and hydrolysis^{27, 28.} Further, caco-2 cells are human intestinal epithelial cells widely used for oral absorption and permeability studies²⁹ and are best suited for efficacy study of oral micronutrient supplements. Cells were grown in basal cell medium with limited micronutrients. Use of basal cell medium allows production of ATP by micronutrient treatment which could be distinguishable and comparable. Since no similar study has been reported previously therefore to optimize maximum ATP production multiple experiments were conducted with varied treatment time (30 min to 4 h) and micronutrient concentration (6.5 µg/mL to 200 µg/mL). Our results showed that maximum ATP level was achieved when cells were treated with micronutrients for 1 h. This also reveal that following oral administration the effect of micronutrient supplement will be apparent within 1 h. Interestingly, micronutrient supplements dose-dependently increased ATP level, with maximal intracellular ATP level observed at 100 µg/mL. Our results were in agreement with previous reports where dose-dependent increase in intracellular ATP was observed following treatment with pyruvate³⁰ and kaempferol³¹. Notably, the ATP levels in cells treated with 100 µg/mL and 200 µg/mL of TV and TM were not significantly different. This showed that the process of ATP production is saturable. This could be plausible explanation for harmful effects observed at very high doses of vitamins and minerals²⁶. Several reports demonstrated neuropathy and ataxia with high doses (>200 mg/ day) of vitamin B6 and hepatotoxic and hypoglycemic effects at high doses (>3000 mg/ day) of vitamin B3. Minerals such as magnesium can cause diarrhea at doses greater than 400 mg/day and high doses of zinc are immunosuppressive and can also cause nausea and vomiting. Selenium at dose in excess of 0.91 mg/ day can cause brittle nails and hair, and peripheral neuropathies²⁶. 

We compared three micronutrient supplements with varied composition and concentration. TV supplement has therapeutic B complex with magnesium and vit C, TM has therapeutic B complex with mineral and NM contains nutraceutical multivitamins and multiminerals. These different compositions allowed comparison of efficacy of therapeutic and nutraceutical concentrations of vitamins and minerals. No treatment group was considered as control whereas treatment with SLS was considered as negative control. The ATP level in control group was significantly higher (p<0.05) than negative control group. This could be due to presence of basal medium that supports cell growth and ATP production. Further, in negative control group, SLS treatment may have caused cell death and thus significantly low production of cellular ATP³². Higher ATP level in supplement treated cells substantiated the therapeutic use of supplements in overcoming fatigue and tiredness. Cells treated with TV showed 1.2-fold higher ATP compared to TM treatment signifying that vitamins are comparatively more effective than minerals. The findings were in agreement with previous reports^{13, 33}.

Nevertheless, higher ATP levels in TM treated cells compared to NM and control, substantiates the role of minerals in ATP production. Interestingly, TM supplement showed ATP rise even in the absence of magnesium, which is considered as very important factor for ATP generation since ATP in cells in bound to magnesium ions²². It is hypothesized that chromium picolinate or selenious acid or both in TM supplement have performed activity similar to magnesium and resulted in ATP production in cells. Markedly higher ATP level in cells treated with TV and TM compared to NM provide scientific evidence that support the role of micronutrient concentration in ATP production. Further, low ATP level in cells treated with NM might be due to low vitamins and minerals in nutraceutical composition. This also supports significantly higher ATP level in cell treated with 200 µg/mL NM compared to 100 µg/mL. In contrast, cells treated with TV and TM showed saturation at 100 µg/mL concentration. Interestingly, despite 10-fold lower micronutrients in NM supplement, the ATP levels were only about 1.7-fold lower. This underscores the possibility of synergistic activity of vitamin and minerals. The decreasing order of ATP levels in cells is as followed: TV > TM > NM>CO>NC.

The present in vitro study has several limitations. Firstly, the study evaluated the efficacy of supplement composition rather than individual components. It was speculated that determining efficacy of individual component would be difficult. This is because cell growth requires micronutrients and in presence of other micronutrient in media it would be difficult to analyze the effect of individual component. Secondly, study was conducted on Caco2 cells, whereas previously NSCLC A549 cells have also been used. Nevertheless, use of ATP release and hydrolysis has been studied with Caco2 cells previously^{27, 28} and NSCLC A549 cells have been deployed previously for studying eATP with regard to drug resistance mechanisms and cancer metabolism and hence not used in this study. In conclusion, our studies provide the scientific evidence that support the therapeutic role of micronutrient supplements in fatigue, tiredness and lethargy or any other disease caused due to cellular energy depletion. Further, the composition and concentration of vitamins and minerals present in the supplements are crucial for optimal efficacy. Therapeutic B complex with magnesium and vitamin C or with minerals performed better than nutraceutical multivitamin and multimineral.

Acknowledgment. The authors extend their gratitude to DSIR-DPSRU-CRTDH for providing the infrastructure necessary for conducting the study.

Conflict of Interest: The co-authors Parimal Mahajan, Gajanan Panchal, Maneesha Khalse, Kamlesh Patel are the employees of Lupin Limited, Mumbai, India.

Funding Source: The study was supported by Lupin Limited, Mumbai, India.

Author Contributions: All authors have equal contributions in the preparation of the manuscript and compilation.

Informed Consent Statement: Not applicable. 

Data Availability Statement: The data presented in this study are available on request from the corresponding author.  

Ethical approval: Not applicable.

REFERENCES

1. Maisel P, Baum E, Donner-Banzhoff N. Fatigue as the Chief Complaint-Epidemiology, Causes, Diagnosis, and Treatment. Dtsch Arztebl Int. 2021;118(33-34):566-576. https://doi.org/10.3238/arztebl.m2021.0192 PMid:34196270 PMCid:PMC8579431

2. Constantin-Teodosiu D, Constantin D. Molecular Mechanisms of Muscle Fatigue. Int J Mol Sci. 2021;22(21):11587. Published 2021 Oct 27. https://doi.org/10.3390/ijms222111587 PMid:34769017 PMCid:PMC8584022

3. igoulet M, Bouchez CL, Paumard P, et al. Cell energy metabolism: An update. Biochim Biophys Acta Bioenerg. 2020;1861(11):148276. https://doi.org/10.1016/j.bbabio.2020.148276 PMid:32717222

4. Hultman E, Greenhaff PL. Skeletal muscle energy metabolism and fatigue during intense exercise in man. Sci Prog. 1991;75(298 Pt 3-4):361-370.

5. van Calker D, Biber K, Domschke K, Serchov T. The role of adenosine receptors in mood and anxiety disorders. J Neurochem. 2019;151(1):11-27. https://doi.org/10.1111/jnc.14841 PMid:31361031

6. Wang K, Huang S, Fu D, et al. The neurobiological mechanisms and therapeutic prospect of extracellular ATP in depression. CNS Neurosci Ther. 2024;30(2):e14536. https://doi.org/10.1111/cns.14536 PMid:38375982 PMCid:PMC10877668

7. Chikahisa S, Séi H. The role of ATP in sleep regulation. Front Neurol. 2011 Dec 27;2:87. https://doi.org/10.3389/fneur.2011.00087 PMid:22207863 PMCid:PMC3246291

8. Dautant A, Meier T, Hahn A, Tribouillard-Tanvier D, di Rago JP, Kucharczyk R. ATP Synthase Diseases of Mitochondrial Genetic Origin. Front Physiol. 2018 Apr 4;9:329. https://doi.org/10.3389/fphys.2018.00329 PMid:29670542 PMCid:PMC5893901

9. Rathmacher, J.A., Fuller, J.C., Baier, S.M. et al. Adenosine-5'-triphosphate (ATP) supplementation improves low peak muscle torque and torque fatigue during repeated high intensity exercise sets. J Int Soc Sports Nutr 9, 48 (2012). https://doi.org/10.1186/1550-2783-9-48 PMid:23046855 PMCid:PMC3483284

10. Castro-Marrero J, Sáez-Francàs N, Segundo MJ, et al. Effect of coenzyme Q10 plus nicotinamide adenine dinucleotide supplementation on maximum heart rate after exercise testing in chronic fatigue syndrome - A randomized, controlled, double-blind trial. Clin Nutr. 2016;35(4):826-834. https://doi.org/10.1016/j.clnu.2015.07.010 PMid:26212172

11. Maggini S, Óvári V, Ferreres Giménez I, Pueyo Alamán MG. Benefits of micronutrient supplementation on nutritional status, energy metabolism, and subjective wellbeing. Nutr Hosp. 2021;38(Spec No2):3-8. https://doi.org/10.20960/nh.03788

12. Barnish M, Sheikh M, Scholey A. Nutrient Therapy for the Improvement of Fatigue Symptoms. Nutrients. 2023;15(9):2154. https://doi.org/10.3390/nu15092154 PMid:37432282 PMCid:PMC10181316

13. Tardy AL, Pouteau E, Marquez D, Yilmaz C, Scholey A. Vitamins and Minerals for Energy, Fatigue and Cognition: A Narrative Review of the Biochemical and Clinical Evidence. Nutrients. 2020 Jan 16;12(1):228. https://doi.org/10.3390/nu12010228 PMid:31963141 PMCid:PMC7019700

14. Bager P, Hvas CL, Hansen MM, Ueland P, Dahlerup JF. B-vitamins, related vitamers, and metabolites in patients with quiescent inflammatory bowel disease and chronic fatigue treated with high dose oral thiamine. Mol Med. 2023 Oct 25;29(1):143. https://doi.org/10.1186/s10020-023-00741-3 PMid:37880581 PMCid:PMC10601301

15. Aragão MÂ, Pires L, Santos-Buelga C, Barros L, Calhelha RC. Revitalising Riboflavin: Unveiling Its Timeless Significance in Human Physiology and Health. Foods. 2024;13(14):2255. https://doi.org/10.3390/foods13142255 PMid:39063339 PMCid:PMC11276209

16. Bogan L.K., Brenner C. Nicotinic acid, nicotinamide, and nicotinamide riboside: A molecular evaluation of NAD+ precursor vitamins in human nutrition. Annu. Rev. Nutr. 2008;28:115-130. https://doi.org/10.1146/annurev.nutr.28.061807.155443 PMid:18429699

17. Leonardi R, Jackowski S. Biosynthesis of Pantothenic Acid and Coenzyme A. EcoSal Plus. 2007 Apr;2(2):10.1128/ecosalplus.3.6.3.4. https://doi.org/10.1128/ecosalplus.3.6.3.4 PMid:26443589 PMCid:PMC4950986

18. Parra M, Stahl S, Hellmann H. Vitamin B₆ and Its Role in Cell Metabolism and Physiology. Cells. 2018 Jul 22;7(7):84. https://doi.org/10.3390/cells7070084 PMid:30037155 PMCid:PMC6071262

19. Cui QL, Lin YH, Xu YKT, Fernandes MGF, Rao VTS, Kennedy TE, Antel J. Effects of Biotin on survival, ensheathment, and ATP production by oligodendrocyte lineage cells in vitro. PLoS One. 2020 May 29;15(5):e0233859. https://doi.org/10.1371/journal.pone.0233859 PMid:32470040 PMCid:PMC7259710

20. Zheng Y, Cantley LC. Toward a better understanding of folate metabolism in health and disease. J Exp Med. 2019 Feb 4;216(2):253-266. https://doi.org/10.1084/jem.20181965 PMid:30587505 PMCid:PMC6363433

21. Schubert HL, Hill CP. Structure of ATP-bound human ATP:cobalamin adenosyltransferase. Biochemistry. 2006 Dec 26;45(51):15188-96. https://doi.org/10.1021/bi061396f PMid:17176040 PMCid:PMC2532598

22. Kleczkowski LA, Igamberdiev AU. Magnesium and cell energetics: At the junction of metabolism of adenylate and non-adenylate nucleotides. J Plant Physiol. 2023;280:153901. https://doi.org/10.1016/j.jplph.2022.153901 PMid:36549033

23. Yang X., Wang H., Huang C., He X., Xu W., Luo Y., Huang K. Zinc enhances the cellular energy supply to improve cell motility and restore impaired energetic metabolism in a toxic environment induced by OTA. Sci. Rep. 2017;7:14669. https://doi.org/10.1038/s41598-017-14868-x PMid:29116164 PMCid:PMC5676743

24. Wesolowski LT, Semanchik PL, White-Springer SH. Beyond antioxidants: Selenium and skeletal muscle mitochondria. Front Vet Sci. 2022 Dec 1;9:1011159. https://doi.org/10.3389/fvets.2022.1011159 PMid:36532343 PMCid:PMC9751202

25. Pala R, Sari MA, Erten F, et al. The effects of chromium picolinate on glucose and lipid metabolism in running rats. J Trace Elem Med Biol. 2020;58:126434. https://doi.org/10.1016/j.jtemb.2019.126434 PMid:31778961

26. Renwick AG. Toxicology of micronutrients: adverse effects and uncertainty. J Nutr. 2006;136(2):493S-501S. https://doi.org/10.1093/jn/136.2.493S PMid:16424134

27. Schachter J, Alvarez CL, Bazzi Z, et al. Extracellular ATP hydrolysis in Caco-2 human intestinal cell line. Biochim Biophys Acta Biomembr. 2021;1863(10):183679. https://doi.org/10.1016/j.bbamem.2021.183679 PMid:34216588

28. Saffioti NA, Alvarez CL, Bazzi Z, et al. Dynamic recycling of extracellular ATP in human epithelial intestinal cells. PLoS Comput Biol. 2023;19(6):e1011196. https://doi.org/10.1371/journal.pcbi.1011196 PMid:37384797 PMCid:PMC10337955

29. van Breemen RB, Li Y. Caco-2 cell permeability assays to measure drug absorption. Expert Opin Drug Metab Toxicol. 2005;1(2):175-185. https://doi.org/10.1517/17425255.1.2.175 PMid:16922635

30. Ozawa, S., Ueda, S., Imamura, H. et al. Glycolysis, but not Mitochondria, responsible for intracellular ATP distribution in cortical area of podocytes. Sci Rep 5, 18575 (2016). https://doi.org/10.1038/srep18575 PMid:26677804 PMCid:PMC4683464

31. Akiyama M, Mizokami T, Miyamoto S, Ikeda Y. Kaempferol increases intracellular ATP content in C2C12 myotubes under hypoxic conditions by suppressing the HIF-1α stabilization and/or by enhancing the mitochondrial complex IV activity. J Nutr Biochem. 2022;103:108949. https://doi.org/10.1016/j.jnutbio.2022.108949 PMid:35122998

32. Maher S, Geoghegan C, Brayden DJ. Safety of surfactant excipients in oral drug formulations. Adv Drug Deliv Rev. 2023;202:115086. https://doi.org/10.1016/j.addr.2023.115086 PMid:37739041

33. Costa-Pinto R, Gantner D. Macronutrients, minerals, vitamins and energy. Anaesthesia & Intensive Care Medicine. 2020 Mar 1;21(3):157-61. https://doi.org/10.1016/j.mpaic.2019.12.006