Abstract Adenosine Triphosphate Research Articles

Protein active site prediction for early drug discovery and designing

Abstract Adenosine triphosphate (ATP) is an energy compound present in living organisms and is required by living cells for performing operations such as replication, molecules transportation, chemical synthesis, etc. ATP connects with living cells through specialized sites called ATP-sites. ATP-sites are present in various proteins of a living cell. The life span of a cell can be controlled by controlling ATP compounds and without the provision of energy to ATP compounds, cells cannot survive. Countless diseases treatment (such as cancer, diabetes) can be possible once protein active sites are predicted. Considering the need for an algorithm that predicts ATP-sites with higher accuracy and effectiveness, this research work predicts protein ATP sites in a very novel way. Till now Position-specific scoring matrix (PSSM) along with many physicochemical properties have been used as features with deep neural networks in order to create a model that predicts the ATP-sites. To overcome this problem of complex computation, this exertion proposes k-mer feature vectors with simple machine learning (ML) models to attain the same or even better performance with less computation required. Using 2-mer as feature vectors, this research work trained and tested five different models including KNN, Conv1D, XGBoost, SVM and Random Forest. SVM gave the best performance on k-mer features. The accuracy of the created model is 96%, MCC 90% and ROC-AUC is 99%, which are the same or even better in some aspects than the state-of-the-art results. The state-of-the-art results have an accuracy of 97%, MCC 78% and ROC-AUC is 92%. One of the benefits of the created model is that it is much simpler and more accurate.

Alterations in pore-forming subunits of adenosine triphosphate-sensitive potassium channels in pressure overload rat cardiomyocytes.

Adenosine triphosphate (ATP)-sensitive potassium channels (KATP) link cellular metabolic state and electrical activity of cardiomyocytes. Pharmacological studies indicated the involvement of KATP in pressure overload-induced left ventricular hypertrophy, but the alteration of pore-forming subunits, Kir6.1 and Kir6.2, at membranes and subcellular fractions is unclear. The objective of this study was to investigate the changes in the distribution and levels of Kir6.1 and Kir6.2 in rat cardiomyocytes responding to chronic pressure overload. Male Wistar rats were separated into a sham-operated group (sham) and a pressure overload or transverse aortic constriction (TAC) group. Echocardiography was performed to assess cardiac structure and functions at the 4th month after operation. Ventricular cardiomyocytes in both groups were isolated and then subjected to extract protein into cytoplasm, organelle membrane, and plasma membrane fractions. Immunoblotting and immunohistochemistry techniques were performed to detect and measure Kir6.1 and Kir6.2 protein levels. Echocardiographic parameters showed left ventricular hypertrophy with hypercontractility in TAC rats. The immunoblotting showed the presence of Kir6.1 and Kir6.2 at plasma membranes and only Kir6.1 at organelle membranes. Significantly, Kir6.1 and Kir6.2 levels were decreased in the plasma membrane fraction of the TAC group (n = 8 and 6 for Kir6.1 and Kir6.2, respectively), whereas Kir6.1 in the organelle membrane fraction tended to be higher in TAC but did not reach statistical significance (n = 7). These results seemed to relate to a left ventricular immunohistochemistry study, which showed the trend of decreased staining of Kir6.1 and Kir6.2 in pressure overload left ventricular tissue. In conclusion, both Kir6.1 and Kir6.2 plasma membrane subunits were decreased significantly in pressure overload-induced left ventricular hypertrophy.

Влияние пуринергической сигнальной системы на свойства клеток крови человека при старении организма

Adenosine triphosphate (ATP) molecules and nucleotide derivatives contribute to a wide range of cellular reactions through the activation of A2 and P2 purinergic receptors located on the cell surface. Almost any type of cells expresses such receptors, including red blood cells and white blood cells. The study of purinergic transmission of signals mediating calcium mobilization, actin polymerization, chemotaxis, release of mediators, tissue oxygenation, cell maturation, cytotoxicity, cell death and cell–cell interactions in dynamics at different stages of ontogenesis is an important issue in biology and medicine today. This paper aimed to explore the biophysical properties of blood cells during the activation of the purinergic signalling system in middle-aged people and older adults. To activate the elements of the purinergic signalling system, an in vitro model of mechanical stress was used. Blood samples were analysed using the methods of atomic force microscopy, namely, force spectroscopy and the Kelvin probe. When the purinergic signalling system is activated, an increase in ATP blood concentration is identified in the blood of both middle-aged people and older adults. In the blood of older adults, a smaller amount of ATP was recorded compared with the blood of middle-aged people, which can be due to changes in the expression of purinoreceptors on the surface of red blood cells at ageing. Erythrocytes and granulocytes in older adults are characterized by increased surface stiffness and potential upon activation of the purinergic signalling system, compared with cells of middle-aged people. In response to mechanical stress, lymphocyte membrane stiffness in older adults increases more significantly than in middle-aged people. Thus, the results indicate an important role of the ATP molecule and its derivatives in the regulation of functioning of blood cells through binding to receptors of the P2 family. Research on the changes in the biophysical properties of red blood cells and white blood cells at activation of the purinergic signalling system will allow us to identify new effects of purine compounds on intercellular interaction during ageing.

Strategies to gain novel Alzheimer's disease diagnostics and therapeutics using modulators of ABCA transporters.

Adenosine-triphosphate-(ATP)-binding cassette (ABC) transport proteins are ubiquitously present membrane-bound efflux pumps that distribute endo- and xenobiotics across intra- and intercellular barriers. Discovered over 40 years ago, ABC transporters have been identified as key players in various human diseases, such as multidrug-resistant cancer and atherosclerosis, but also neurodegenerative diseases, such as Alzheimer’s disease (AD). Most prominent and well-studied are ABCB1, ABCC1, and ABCG2, not only due to their contribution to the multidrug resistance (MDR) phenotype in cancer, but also due to their contribution to AD. However, our understanding of other ABC transporters is limited, and most of the 49 human ABC transporters have been largely neglected as potential targets for novel small-molecule drugs. This is especially true for the ABCA subfamily, which contains several members known to play a role in AD initiation and progression. This review provides up-to-date information on the proposed functional background and pathological role of ABCA transporters in AD. We also provide an overview of small-molecules shown to interact with ABCA transporters as well as potential in silico, in vitro, and in vivo methodologies to gain novel templates for the development of innovative ABC transporter-targeting diagnostics and therapeutics.

Nicorandil Affects Mitochondrial Respiratory Chain Function by Increasing Complex III Activity and ROS Production in Skeletal Muscle Mitochondria.

Adenosine triphosphate (ATP)-dependent potassium channels openers (KATP) protect skeletal muscle against function impairment through the activation of the mitochondrial KATP channels (mitoKATP). Previous reports suggest that modulators of the mitochondrial KATP channels have additional effects on isolated mitochondria. To determine whether the KATPchannel opener nicorandil has non-specific effects that explain its protective effect through the mitochondrial function, chicken muscle mitochondria were isolated, and respiration rate was determined pollarographically. The activity of the electron transport chain (ETC) complexes (I-IV) was measured using a spectrophotometric method. Reactive oxygen species(ROS)levels and lipid peroxidation were assessed using flow cytometry and thiobarbituric acid assay, respectively. Both KATP channel opener nicorandil and KATP channel blocker 5-hydroxydecanoate (5-HD) decreased mitochondrial respiration; nicorandil increased complex III activity and decreased complex IV activity. The effects of nicorandil on complex III were antagonized by 5-HD. Nicorandil increased ROS levels, effect reverted by either 5-HD or the antioxidant N-2-mercaptopropionyl glycine (MPG). None of these drugs affected lipid peroxidation levels. These findings suggest that KATPchannel opener nicorandil increases mitochondrial ROS production from complex III. This results by partially blocking electron flow in the complex IV, setting electron carriers in a more reduced state, which is favored by the increase in complex III activity by nicorandil. Overall, our study showed that nicorandil like other mitochondrial KATP channel openers might not act through mitoKATP channel activation.

New role in cells suggested for ATP

Adenosine triphosphate (ATP) performs many jobs in a cell. It carries energy, serves as a signaling molecule, and is the source of adenosine in DNA and RNA. But cells contain far more ATP—as much as 5 mM in the cytoplasm—than these known uses seem to require. That might be because ATP also can solubilize proteins, suggests a new study (Science 2017, DOI: 10.1126/science.aaf6846). ATP has the general characteristics of a hydrotrope, an amphiphilic molecule that has both a hydrophilic and a hydrophobic component but does not assemble into structures such as micelles. Hydrotropes are used industrially to solubilize hydrophobic species in aqueous solution. The hydrophobic portion of hydrotropes—such as ATP’s adenosine—likely interacts with the hydrophobic species, while the hydrophilic part—such as ATP’s triphosphate—allows the species to stay in solution. In the new work, a team led by Yamuna Krishnan of the University of Chicago and Anthony A. Hyman of the

Genomic Analysis of ATP Efflux in Saccharomyces cerevisiae.

Adenosine triphosphate (ATP) plays an important role as a primary molecule for the transfer of chemical energy to drive biological processes. ATP also functions as an extracellular signaling molecule in a diverse array of eukaryotic taxa in a conserved process known as purinergic signaling. Given the important roles of extracellular ATP in cell signaling, we sought to comprehensively elucidate the pathways and mechanisms governing ATP efflux from eukaryotic cells. Here, we present results of a genomic analysis of ATP efflux from Saccharomyces cerevisiae by measuring extracellular ATP levels in cultures of 4609 deletion mutants. This screen revealed key cellular processes that regulate extracellular ATP levels, including mitochondrial translation and vesicle sorting in the late endosome, indicating that ATP production and transport through vesicles are required for efflux. We also observed evidence for altered ATP efflux in strains deleted for genes involved in amino acid signaling, and mitochondrial retrograde signaling. Based on these results, we propose a model in which the retrograde signaling pathway potentiates amino acid signaling to promote mitochondrial respiration. This study advances our understanding of the mechanism of ATP secretion in eukaryotes and implicates TOR complex 1 (TORC1) and nutrient signaling pathways in the regulation of ATP efflux. These results will facilitate analysis of ATP efflux mechanisms in higher eukaryotes.

Assessment of the quality of cleaning of surfaces in care rooms of intensive care units: feasibility of the use of ATP-metry

Adenosine triphosphate (ATP) is the basic energy molecule for all living cells. Its presence on a surface is a sign of contamination by microorganisms. ATP-metry is an ATP detection technique based on chemiluminescence. It is used in the food industry to assess the quality of cleaning of surfaces.

Investigation of oxidative phosphorylation in continuous cultures. A non-equilibrium thermodynamic approach to energy transduction for Escherichia coli in aerobic condition

Abstract Adenosine triphosphate (ATP) production in living cells is very important. Different researches have shown that in terms of mathematical modeling, the domain of these investigations is essentially restricted. Recently the thermodynamic models have been suggested for calculation of the efficiency of oxidative phosphorylation process and rate of energy loss in animal cells using chemiosmotic theory and non-equilibrium thermodynamics equations. In our previous work, we developed a mathematical model for mitochondria of animal cells. In this research, according to similarities between oxidative phosphorylation process in microorganisms and animal cells, Golfar's model was developed to predict the non-equilibrium thermodynamic behavior of the oxidative phosphorylation process for bacteria in aerobic condition. With this model the rate of energy loss, P/O ratio, and efficiency of oxidative phosphorylation were calculated for Escherichia coli in aerobic condition. The results then were compared with experimental data given by other authors. The thermodynamic model had an acceptable agreement with the experimental data.

ATP induced microvesicles stimulate macrophage activation (89.18)

Abstract Adenosine triphosphate (ATP) has a unique role as a damage associated molecular pattern (DAMP) as it is normally kept within cells unless there is cellular/tissue injury. As a DAMP, ATP has several known mechanisms for immune cell activation. Our lab and others have reported that exogenous ATP administration to macrophages in cell culture induces rapid shedding of microvesicles (MV). In this study, we harvested MV from ATP treated macrophages and demonstrated that MV are capable of stimulating naive macrophages to release pro-inflammatory cytokines and induce co-stimulatory surface marker upregulation. MV- induced activation is dependent on a cAMP/p38 MAPK pathway and also results in NFkappaB translocation. Activation is partially dependent on TLR4, but is not due to the TLR4 agonist HMGB1, which is present in MV. TNFalpha is also present in MV but is not required to activate macrophages since MV from TNF-deficient cells are also stimulatory. Biochemical fractionation of MV was then used to show that the stimulatory component is phospholipid in nature, and we are currently working to identify the bioactive compound. These findings suggest a novel means by which macrophages at sites of cellular damage can become sterilely activated by ATP to further propagate cellular damage via shed microvesicles.

MRP transporters as membrane machinery in the bradykinin-inducible export of ATP

Adenosine triphosphate (ATP) plays the role of an autocrine/paracrine signal molecule in a variety of cells. So far, however, the membrane machinery in the export of intracellular ATP remains poorly understood. Activation of B2-receptor with bradykinin-induced massive release of ATP from cultured taenia coli smooth muscle cells. The evoked release of ATP was unaffected by gap junction hemichannel blockers, such as 18alpha-glycyrrhetinic acid and Gap 26. Furthermore, the cystic fibrosis transmembrane regulator (CFTR) coupled Cl(-) channel blockers, CFTR(inh)172, 5-nitro-2-(3-phenylpropylamino)-benzoic acid, Gd3(+) and glibenclamide, failed to suppress the export of ATP by bradykinin. On the other, the evoked release of ATP was greatly reduced by multidrug resistance protein (MRP) transporter inhibitors, MK-571, indomethacin, and benzbromarone. From western blotting analysis, blots of MRP 1 protein only, but not MRP 2 and MRP 3 protein, appeared at 190 kD. However, the MRP 1 protein expression was not enhanced after loading with 1 muM bradykinin for 5 min. Likewise, niflumic acid and fulfenamic acid, Ca2(+)-activated Cl(-) channel blockers, largely abated the evoked release of ATP. The possibility that the MRP transporter system couples with Ca2(+)-activated Cl(-) channel activities is discussed here. These findings suggest that MRP transporters, probably MRP 1, unlike CFTR-Cl(-) channels and gap junction hemichannels, may contribute as membrane machinery to the export of ATP induced by G-protein-coupled receptor stimulation.

Real-Time Testing of Bioburdens in Metalworking Fluids Using Adenosine Triphosphate as a Biomass Indicator

Adenosine triphosphate (ATP) assays have been used to quantify bioburdens (biomass) in low-organic-compound-content fluids (freshwater, seawater, cooling tower water, and similar fluids) since the early 1950s. The original methodology was labor intensive and required considerable laboratory skill. Over the past half-century, the protocol has been simplified substantially, but until recently, chemical interferences made it impractical to use the ATP test in metalworking fluids (MWF). This article presents precision and bias statistics for a new test protocol for ATP in emulsifiable oil, semisynthetic and synthetic MWF at end-use dilutions. Additionally it presents the results of field tests in which ATP data are compared with other MWF condition monitoring data. The field evaluation demonstrates the applicability of the new protocol to MWF bioburden condition monitoring.

Vasodilatory effect of adenosine triphosphate does not change cerebral blood flow: a PET study with 15O-water

Adenosine triphosphate (ATP) is the parent compound of adenosine and well known as a powerful vasodilator. To investigate the effect of ATP on cerebral blood flow (CBF) and cerebral vessels, (15)O-water positron emission tomography (PET) studies were performed to evaluate changes in CBF and blood volume before and after ATP administration. Ten healthy young volunteers underwent (15)O-water PET scans under the conditions of baseline, 3 and 1 min after ATP continuous infusion. CBF values in cortical regions of the bilateral middle cerebral arteries and basal ganglia were obtained for each subject. Statistical parametric mapping (SPM) was applied for analysis of regional changes. Physiologic parameters, such as blood gas and blood pressure were also measured. Cortical CBF showed no significant change after continuous infusion of ATP compared with the baseline. Dilatation of major vessels induced by ATP was visualized on SPM analysis. Heart rates increased and mean blood pressure decreased during ATP administration while blood gas data showed no changes between the different conditions. Intravenous ATP administration caused dilatation of major cerebral vessels but no significant change in CBF under normoventilation and decrease in systemic blood pressure, indicating that this no change in CBF under vasodilatory effect of ATP may be caused by cerebral microvascular autoregulation.

Computer Model of Nucleotide Transport in a Realistic Porcine Aortic Trifurcation

Adenosine triphosphate (ATP) is a ubiquitous blood borne agonist which is responsible for the regulation of vascular tone via purinogenic signalling pathways. The present study models the transport of ATP in a realistic porcine aortic trifurcation, which includes multiple branches and bifurcations. The focus of the present study is understanding how pulsatile flow effects mass transfer, observing both mean and transient variations. Unlike in the many idealized models which model transport of low diffusion coefficient species, the realistic geometry leads to very different mass transfer characteristics. These include spiral patterns in the distribution of low concentration fluid. Furthermore, the mean ATP distribution was found to be elevated compared with the steady state; this is attributed to the effects of convective mixing. The results strongly implicate that under certain conditions mass transport in pulsatile flow exhibits different hydrolysis characteristics at the endothelium compared with steady state. Transient variations throughout the the cardiac cycle were found to be small. This small transient response is primarily due to low ATP diffusivity.

Citric Acid Interferes with Adenosine Triphosphate Determination by Bioluminescence

Adenosine triphosphate (ATP) measurement by bioluminescence is used in microbial adsorption studies for which colony-forming units (CFU) may underestimate true microbial numbers. We observed that citric acid profoundly affected ATP measurement by luciferin-luciferase assay, and we assessed the effect of citric acid on ATP detection in water and buffer systems in the pH range 4.9 to 7.4. Bioluminescence depended on the buffer system, increasing as pH increased in each buffer system and decreasing as the buffer system changed from Na-phosphate to citrate-phosphate. Citrate-phosphate buffer decreased bioluminescence by 82% relative to water at the same pH. Consequences of poor buffer selection are potentially reduced luciferase activity in the bioassay system and underestimation of microbial numbers.

Monitoring ATP release from individual cells with a biosensor

Adenosine triphosphate (ATP) is a neurotransmitter/neuromodulator in both central and peripheral nervous systems. Particularly in the taste bud, a peripheral taste organ, ATP serves as an afferent neurotransmitter. To examine the mechanism that mediates ATP secretion in taste cells, we elaborated an approach for monitoring ATP in an extracellular medium by employing a biosensor, that is, cells responsive to ATP. Two lines of ATP-sensitive cells, HEK-293 and COS-1, which endogenously express P2Y receptors, were employed. In addition, HEK-293 cells transfected with P2X3 receptors were also used. By most relevant parameters (threshold response, inactivation kinetics of ATP responses, and refractory period), COS-1 cells were more suitable as an ATP sensor than HEK-293 cells, both native and transfected. For the HEK-293 cell-based biosensor, one of pitfalls was that they were highly responsive to mechanical disturbances, e.g., solution flux elicited by application of a chemical stimulus, owing to the expression of mechanosensitive Ca2+-permeable cation channels. In COS-1 cells, ATP-dependent Ca2+ transients were generated mostly due to Ca2+ release, the feature allowing one to control the activity of ATP-releasing cells electrophysiologically and to monitor the ATP secretion by Ca2+ responses of the ATP-biosensor. By using this technique, it was demonstrated that individual taste cells of a mouse released ATP in response to membrane depolarization.

ATP Inhibits Gravitropism

Adenosine triphosphate (ATP) is both an energy source when intracellular and a signaling molecule when extracellular. In plants, ATP is normally much more concentrated inside the cells than in the extracellular space. Tang et al. determined that ATP inhibited gravitropism of the roots of Arabidopsis seedlings and at higher concentrations promoted root curling. Exogenous addition of ATP also stimulated auxin signaling as measured by increased expression of an auxin-sensitive reporter gene in root tips. Consistent with ATP enhancing auxin signaling, lower concentrations of exogenous auxins were required to produce growth inhibition in the presence of ATP than without added ATP. ATP decreased basipetal auxin transport and increased auxin retention in roots. The authors discuss whether the effect of ATP is due to alteration of the steepness of the intracellular and extracellular ATP gradient or whether ATP may be acting as a signaling molecule stimulating gravitropism. W. Tang, S. R. Brady, Y. Sun, G. K. Muday, S. J. Roux, Extracellular ATP inhibits root gravitropism at concentrations that inhibit polar auxin transport. Plant Physiol. 131 , 147-154 (2003). [Abstract] [Full Text]

Cell cycle arrest and the induction of apoptosis in pancreatic cancer cells exposed to adenosine triphosphate in vitro

Adenosine triphosphate (ATP) has been shown to be an inhibitory or a stimulatory agent for cell growth in various types of cells. Here, we studied the effects of extracellular ATP on two pancreatic cancer cell lines, PK-1 and YAPC established by us. In both cell lines, ATP inhibited cell growth in a time- and dose-dependent manner, whereas the same doses of ATP stimulated DNA synthesis. Flow cytometric analysis of the cells incubated with or without ATP demonstrated the ATP-induced striking increase in cells at S-phase. The same analysis showed also the increase in sub-G0/G1 population in the same analysis and the electrophoretic pattern of DNA showed the occurrence of ATP-induced cell disintegration likely to be apoptosis. We suggest that extracellular ATP is cytotoxic for pancreatic cancer cells because of its induction of cell cycle arrest at S-phase and cell death, possibly apoptosis, overcoming the promotion of the entry into S-phase.

Differential modulation of inspiratory motoneurons and respiratory rhythm generating circuits by ATP

Adenosine triphosphate (ATP) gates a diverse family of P2 (P2X1-7 and P2Y1-11) receptors that are expressed throughout the CNS, including respiratory regions of the brainstem [1]. Activation of ATP receptors potentiates activity of inspiratory motoneurons (MN) [2], modulates respiratory behaviour in vivo when applied to medullary respiratory nuclei and, via activation of pH-sensitive P2X2 receptors, may contribute to central respiratory chemosensitivity [3]. Aims of this study were to: i) determine the effects of ATP receptor activation within the pre-Botzinger complex (preBotC, proposed site of rhythm generation) on respiratory behaviour; ii) test the hypothesis that respiratory networks are endogenously modulated by ATP; and iii) compare the sensitivity of inspiratory MN pools and rhythm generating networks to purinergic modulation. To determine the effects of ATP on respiratory rhythm and inspiratory motor output, ATP was pressure-injected into the pre-BotC and XII nuclei of rhythmically-active medullary slice preparations from neonatal rats, while monitoring XII nerve and MN output. Effects on phrenic MNs were determined by locally applying drugs over the phrenic MN pool of brainstem spinal cord preparations. ATP (10 s, 0.01–1 mM) caused up to a 4-fold, suramin-sensitive (0.05–1.0 mM), increase in frequency, that was followed by a brief (22 ± 5%) reduction. To test whether this post-ATP inhibition was due to hydrolysis of ATP to adenosine and activation of adenosine receptors, we applied ATPγs, a non-hydrolyzable ATP analogue. Peak potentiation of frequency by 0.1 mM ATPγs (3.20 ± 0.3 fold increase, n = 7) was similar to that evoked by ATP (3.30 ± 0.3, n = 7), but the effects of ATPγs were longer lasting (102.9 s ± 10.64 vs 29.3 s ± 2.06 for ATP). The secondary reduction in frequency was also absent following ATPγs. Since P2 receptor antagonists also antagonize glutamate receptors which are essential for rhythm generation, the role of endogenous ATP in modulating respiratory rhythm was investigated via bath application of ectoATPase inhibitors (DEPC, pCMPS) and an allosteric modulator of P2X2 recptors (Cu2+). DEPC (100 μM, n = 4) and pCMPS (30 μM, n = 3) increased respiratory frequency 1.30–1.40 fold, while Cu2+ (10–50 μM, n = 6) increased frequency ~1.5-fold. Local application of ATP (1–10 mM) over XII and phrenic nuclei produced a biphasic response comprising an initial potentiation of burst amplitude (1.40 ± 0.20 of control and 1.22 ± 0.7 respectively) followed by a decrease in burst amplitude (to 0.82 ± 0.05 and 0.90 ± 0.05 of control respectively) that was theophylline-sensitive and absent following application of ATP-γ-s. The doses of ATP required to potentiate burst amplitude by 1.20–1.40 of control [2] were ~100 times higher than required to increase rhythm 2.5- to 4-fold. Results show that exogenous ATP potently increases respiratory frequency and that rhythm generating networks are significantly more sensitive to ATP than respiratory motoneurons. In addition, effects of ectoATPase inhibitors and allosteric modulators of ATP receptors suggest that respiratory networks are endogenously modulated by ATP. The differential sensitivity of rhythm generating elements and motoneurons provides an opportunity to explore the physiological significance of P2 receptor diversity to respiratory control.

Development of ATP assay as a surrogate indicator of viability of Cryptosporidium parvum oocysts

Adenosine Triphosphate (ATP) determination was applied to evaluate viability of Cryptosporidium oocysts. Three pretreatment methods, such as incubation in acidified Hanks balanced salt solution (HBSS), excystation and sonication were investigated for ATP extraction from oocysts. Incubation in acidified HBSS was insufficient to extract ATP from oocysts, but a linear relationship between the number of oocysts and the concentration of ATP extracted was observed in the test of excystation and sonication treatments. Sonicationtreatment was able to extract ATP from oocysts more rapidly and precisely than excystation treatment. ATP amount per oocyst by sonication treatment (ATPs) was evaluated to be 2.9×10–8 μg on average, andits detection limit was 500 oocysts/100 μl. Ozone treatment experiments were conducted in batch condition to evaluate differences among ATP concentrations extracted, in vitro excystation and DAPI/PI permeability assays. ATPs assay was observed to have a linear relationship with DAPI/PI permeability assay (R2=0.98). As a result, ATP assay is applicable as a surrogate indicator of the viability of C. parvum, and is superior to in vitro excystation and DAPI/PI permeability assay, because of its rapid, accurate and simple procedure.