High-performance liquid chromatographic methods for base and nucleoside analysis in extracellular fluids and in cells

Simmonds, R. J.; Harkness, R. A.

doi:10.1016/s0378-4347(00)86071-5

Cited by 150 publications

(48 citation statements)

References 26 publications

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“…A drawback is that half the xanthine present is measured as hypoxanthine resulting in a somewhat high hypoxanthine value. The xanthine/hypoxanthine ratio in plasma and other body fluids is normally 20-45% (27,28) and decreases to between 5 and 12% in hypoxia (27,29). Thus the error becomes almost negligible, at least in the hypoxic state.…”

Section: Methodsmentioning

confidence: 99%

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Hypoxanthine as an Indicator of Hypoxia: Its Role in Health and Disease through Free Radical Production

Saugstad¹

1988

Pediatr Res

374

206

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Hypoxia is a common insult during the perinatal and neonatal period. New and better ways to evaluate hypoxia are needed. In 1975 we demonstrated high concentrations of the purine metabolite hypoxanthine in umbilical cord plasma after intrauterine hypoxia and proposed that hypoxanthine could be used as an indicator of hypoxia (1). Since then a large number of studies have been published dealing with different aspects of hypoxanthine in hypoxia. However, investigators in this field have encountered several methodologic problems: 1) Hypoxanthine leaks rapidly from erythrocytes (2, 3); if plasma is not separated promptly from red cells, falsely elevated plasma hypoxanthine concentrations will be found. 2) There are large variations in purine metabolism among species. 3) No clear definition of clinical hypoxia is available. Many authors do not distinguish between the terms hypoxemia and hypoxia, which adds to the confusion. We define tissue hypoxia as: "oxygen deficiency resulting in altered or interrupted energy metabolism" (4). It is important to be aware that hypoxia has two stages. In the first stage it is compensated because the cells are able to meet energy demands through anaerobic metabolism and other mechanisms. (We are not dealing with physiologic adaptation to hypoxia.) In the second stage of hypoxia or uncompensated hypoxia, energy demands are not met and cell injury ensues. At present there are no techniques for distinguishing between these two stages in clinical medicine, and such a distinction would be useful. Theoretically hypoxanthine should only be elevated in uncompensated hypoxia, while pH and lactate changes occur in compensated hypoxia. It seems, however, that hypoxanthine is also elevated to some extent in uncompensated hypoxia.Renewed interest in hypoxanthine developed when it was realized that hypoxanthine is a potential free radical generator (5, 6). Hypoxanthine seems to play a role in posthypoxic reoxygenation cell injury through oxygen radical production (7) and is therefore involved in the pathogenesis of a number of diseases. Hypoxanthine also modulates a number of other processes because it reacts with benzodiazepine receptors (8) and inhibits phosphodiesterase in the brain (9). Hypoxanthine inhibits the effect of several cytotoxic drugs and may therefore influence treatment with such drugs (10).Herein we summarize the extensive literature on hypoxanthine published during the last decade and try to answer the question: How useful in clinical medicine are hypoxanthine measurements in plasma and other body fluids? Is hypoxanthine a better marker of hypoxia than lactate or pH? Further, we discuss the significance of hypoxanthine as a potential oxygen radical generator and put forward a hypothesis for the pathogenesis of an "oxygen radical disease in neonatology". In hypoxia there is an accelerated breakdown of AMP to hypoxanthine. Figure 1 illustrates several important aspects of hypoxanthine metabolism in hypoxia. Normally about 90% of the hypoxanthine formed is reutilized through the sa...

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Section: Methodsmentioning

confidence: 99%

“…Fluorimetry (30, 3 1) and high-performance liquid chromatography (HPLC) (28,(32)(33)(34)(35) provide methods with greater sensitivity. Although several other methods have been described, pOz, fluorimetric, or HPLC methods are the most generally used.…”

Section: Methodsmentioning

confidence: 99%

Hypoxanthine as an Indicator of Hypoxia: Its Role in Health and Disease through Free Radical Production

Saugstad¹

1988

Pediatr Res

374

206

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“…H x analysis. Hx levels in plasma were analyzed according to a HPLC method described in former studies (23,24). Plasma samples were prepared by centrifugation for 30 min through a filter with 30.000 molecular wt cut-off (YMT-filter, MPS-1 System; Amicon Corp., Danvers, MA).…”

Section: Methodsmentioning

confidence: 99%

Reactive Oxygen Metabolites Produce Pulmonary Vasoconstriction in Young Pigs

1991

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ABSTRACT. Reactive oxygen metabolites appear to modulate pulmonary vascular changes. To study the effects of free radical formation in vivo, we investigated five groups of young pigs by recording hemodynamic changes after xanthine oxidase infusion alone and after pretreatment with hypoxanthine or possible blocking agents. The pulmonary vascular pressure increased rapidly in the groups without inhibition reaching maximum levels 25 min after the start of the experiment. The pulmonary artery blood flow declined toward minimum values at the same time. Compared to baseline levels, the calculated vascular lung resistance increased by 300% when the pigs were pretreated with hypoxanthine, and by 150% when xanthine oxidase was given alone. These findings suggest enhanced pulmonary vasoconstriction as a result of high initial hypoxanthine levels probably capable of forming larger quantities of oxygen radicals. The vascular reaction was attenuated when the pigs were pretreated with indomethacin (cyclooxygenase inhibitor) or allopurinol (xanthine oxidase inhibitor). Furthermore, the presence of catalase (hydrogen peroxide scavenger) reduced the pulmonary vasoconstriction significantly. We observed less decline in arterial oxygen tension and oxygen saturation when the animals had been pretreated with inhibitory agents, compared to the blood gas changes found in the xanthine oxidase group. The systemic pressure recordings in the carotid artery remained at baseline levels in all groups. We conclude that oxygen radicals formed by the hypoxanthine-xanthine oxidase system produce severe pulmonary vascular constriction in young pigs. (Pediatr Res 29: 543-547, 1991) Abbreviations Hx, hypoxanthine PVR, pulmonary vascular resistance XO, xanthine oxidase Pao2, arterial oxygen tensionThe complex mechanisms governing the pulmonary circulation in the perinatal period have recently been given extensive consideration in several review articles (1-3). Oxygen-derived free radicals play an important role in experimentally induced hypoxic-reoxygenation injury (4, 5), and may be responsible for both hypoxic and hyperoxic aspects of pulmonary damage (6-

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“…Analysis of hypoxanthine. Hypoxanthine was analyzed using an HPLC method (13,14) with modifications as previously described (1 5).…”

Section: Csf Cerebrospinal Fluidmentioning

confidence: 99%

Hypoxemia and Reoxygenation with 21% or 100% Oxygen in Newborn Pigs: Changes in Blood Pressure, Base Deficit, and Hypoxanthine and Brain Morphology

et al. 1992

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ABSTRACT. To study whether room air is as effective as 100% 0 2 in resuscitation after hypoxia, hypoxemia (Pao2 2.3-4.3 kPa) was induced in newborn pigs (2-5 d old) by ventilation with 8% O2 in nitrogen. When systolic blood pressure had fallen to 20 mm Hg, animals were randomly reoxygenated with either 21% Oz (group 1, n = 9) or 100% 0 2 (group 2, n = 11) for 20 min followed by 21% O2 in both groups. Controls (group 3, n = 5) were ventilated with 21% 0 2 throughout the experiment. Base deficit peaked at 31 f 5 mmol/L (mean f SD) for both hypoxic groups at 5 min of reoxygenation and then normalized over the following 3 h. There were no statistically significant differences between the two groups during reoxygenation concerning blood pressure, heart rate, base deficit, or plasma hypoxanthine. Hypoxanthine peaked at 165 +. 40 and 143 +. 42 ~m o l / L in group 1 and 2 (NS), respectively, and was eliminated monoexponentially in both groups with an initial half-life for excess hypoxanthine of 48 f 21 and 51 +. 27 min (NS), respectively. Blinded pathologic examination of cerebral cortex, cerebellum, and hippocampus after 4 d showed no statistically significant differences with regard to brain damage. We conclude that 21% 0 2 is as effective as 100% O2 for normalizing blood pressure, heart rate, base deficit, and plasma hypoxanthine after severe neonatal hypoxemia in piglets and that the extent of the hypoxic brain damage is similar in the two groups. (Pediatr Res 32: 107-113,1992)

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High-performance liquid chromatographic methods for base and nucleoside analysis in extracellular fluids and in cells

Cited by 150 publications

References 26 publications

Hypoxanthine as an Indicator of Hypoxia: Its Role in Health and Disease through Free Radical Production

Hypoxanthine as an Indicator of Hypoxia: Its Role in Health and Disease through Free Radical Production

Reactive Oxygen Metabolites Produce Pulmonary Vasoconstriction in Young Pigs

Hypoxemia and Reoxygenation with 21% or 100% Oxygen in Newborn Pigs: Changes in Blood Pressure, Base Deficit, and Hypoxanthine and Brain Morphology

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