Diurnal variations in the activities of the glycogen metabolizing enzymes in mouse liver

The glycogen level in mouse liver was maximal during the night and decreased to the lowest level during the light period. The peak activity of phosphorylase alpha was observed during the light hours and thus paralleled the decline of hepatic glycogen concentrations. The period of rapid glycogen synthesis (1800-2200 hr) was immediately preceded by maximum glycogen synthase alpha activity. Significant diurnal rhythms for phosphorylase kinase and phosphorylase phosphatase activities were also observed and appear to play a role in regulating the diurnal rhythm of phosphorylase alpha activity.     

Diurnal Rhythm of the Muscular Performance of Elbow Flexors During Isometric Contractions

The influence of time of day on elbow flexion torque was studied. Thirteen physical education students, 7 males and 6 females, made maximal and submaximal isometric contractions at 90° of elbow flexors using a dynamometer. The torque developed was measured on each contraction. The myoelectric activity of the biceps muscle was also measured at the same time by surface electromyography (EMG) and quantified from the root mean square (RMS) activity. Torque and surface EMGs were measured at 6:00, 9:00, 12:00, 15:00, 18:00, 21:00, and 24:00 h over the same day. Oral temperature before each test session was measured on each occasion after a 30-min rest period. We observed a diurnal rhythm in elbow flexor torque with an acro-phase at 18:00 h and a bathyphase at 6:00 h, in phase with the diurnal rhythm in oral temperature. However, the diurnal rhythm of temperature did not appear to have any influence on the torque. Links between neuromuscular efficiency and RMS/torque ratio were evaluated by measuring muscle activity along with torque. We also assessed variations in the level of maximal activity of the muscle under maximal voluntary contraction. Neuromuscular efficiency fluctuated during the day, with maximal and minimal efficiency at 18:00 h and 9:00 h, respectively, whereas activation level was maximal at 18:00 h and minimal at 9:00 h. The diurnal rhythm of torque was accounted for by variations in both central nervous system command and the contractile state of the muscle.     

Glucocorticoid effects on the diurnal rhythm of circulating leptin levels

It is known that circulating leptin shows diurnal variation with a nocturnal rise; however, the mechanisms generating this rhythm have not been fully elucidated. Glucocorticoids are a potent stimulator of leptin secretion, and there is a reciprocal relationship between circulating leptin and glucocorticoid levels. We hypothesized that glucocorticoids could modulate the diurnal rhythm of circulating leptin. We therefore explored the diurnal variation of leptin under situations in which subjects showed no or some shift of glucocorticoid diurnal rhythm, such as prednisolone-administered humans, and adrenalectomized and corticosterone-replaced (ADX+B) rats. The peak level of plasma cortisol immunoreactivity was shifted from early morning to noon by prednisolone administration. The nocturnal increment of plasma leptin in prednisolone-administered patients (71.2 +/- 14.2% from 08:00 h value) was significantly greater than that in normal volunteers (12.2 +/- 7.5% from 08:00 h value), but the timing of nadir and the peak of plasma leptin was not shifted. In normal rats, the plasma concentration of leptin showed the diurnal rhythm with the bottom at 16:00 h and the top between midnight and early morning. The amplitude of leptin diurnal rhythm was significantly reduced in ADX+B rats (08:00 h: 3.0 +/- 0.2, 16:00 h: 2.7 +/- 0.2, 00:00 h; 3.7 +/- 0.2 ng/ml) compared with sham operated rats (08:00 h: 3.0 +/- 0.2, 16:00 h 2.2 +/- 0.2, 00:00 h: 4.7 +/- 0.4 ng/ml); but ADX+B rats still retained similar timing of nadir and the peak of plasma leptin as observed in sham rats. These results indicate that glucocorticoids enhance the amplitude of leptin diurnal rhythm, and are consistent with previous findings showing that glucocorticoids increase leptin secretion. Glucocorticoids appear to play modulatory, but not essential roles in generating leptin diurnal rhythm.     

In vitro TRH release from hypothalamus slices varies during the diurnal cycle

We have previously described a daily rhythm in thyrotropin releasing hormone (TRH) and TRH mRNA in the rat hypothalamus. To determine whether TRH release fluctuates in a diurnal manner, we have measured basal and potassium stimulated release from hypothalamic slices, and compared it to release from olfactory bulb slices, during the diurnal cycle. Basal TRH release was higher at 7:00 h than at any other time (1:00, 13:00 or 19:00 h) in either hypothalamus or olfactory bulb. The ratio of stimulated over basal release was higher in the hypothalamus at 19:00 h, when TRH content was highest. Potassium stimulated TRH release from olfactory bulb was not different from basal release at any time. TRH release fluctuations were not due to a rhythm of extracellular inactivation: the activity of pyroglutamyl aminopeptidase II, an ectoenzyme responsible for TRH inactivation, was constant throughout the cycle. Our data demonstrate that diurnal variations of TRH release occur in vitro and that the enhanced responsiveness to potassium stimulation in hypothalamus is correlated with increased levels of peptide.     

Diurnal Variations of Striatal D2 Dopaminergic Receptors and its Relation with Haloperidol-induced Catalepsy

In spite of the clear evidences for the blockade of dopaminergic D2 receptors as the mechanism of action for haloperidol-induced catalepsy, the contribution of pharmacokinetic and pharmacodynamic aspects on the diurnal modulation of haloperidol-induced catalepsy is controversial. We studied the diurnal variations of striatal dopamine receptors and its relation with catalepsy diurnal variations. The [ 3 H]-spiperone binding to dopamine receptors had a clear rhythm with a peak at 00:00 to 03:00 h, and a trough at 12:00 to 18:00 h. Haloperidol-produced catalepsy measured with the four-cork test, also showed a clear rhythm, with a peak at 00:00 h and trough at 9:00 h. The dose-response curves at peak and trough of catalepsy had the same ED 50 (0.12 mg), with time-related changes in the maximal effect. Similar diurnal variations between catalepsy and dopamine receptor binding, indicate a relevant role of temporal pharmacodynamics of haloperidol on the modulation of its behavioral effects.     

Diurnal Variations of Striatal D2 Dopaminergic Receptors and its Relation with Haloperidol-induced Catalepsy

In spite of the clear evidences for the blockade of dopaminergic D2 receptors as the mechanism of action for haloperidol-induced catalepsy, the contribution of pharmacokinetic and pharmacodynamic aspects on the diurnal modulation of haloperidol-induced catalepsy is controversial. We studied the diurnal variations of striatal dopamine receptors and its relation with catalepsy diurnal variations. The [3 H]-spiperone binding to dopamine receptors had a clear rhythm with a peak at 00:00 to 03:00 h, and a trough at 12:00 to 18:00 h. Haloperidol-produced catalepsy measured with the four-cork test, also showed a clear rhythm, with a peak at 00:00 h and trough at 9:00 h. The dose-response curves at peak and trough of catalepsy had the same ED 50 (0.12 mg), with time-related changes in the maximal effect. Similar diurnal variations between catalepsy and dopamine receptor binding, indicate a relevant role of temporal pharmacodynamics of haloperidol on the modulation of its behavioral effects.     

Malate Dehydrogenase Activity of Rat Testis: Circadian Rhythm and its Ontogeny

Malate dehydrogenase activity and soluble protein content in testes from rats exposed to a 14:00 h light:10:00 h dark photoperiod, have been determined every two or four hours over a 24 hour period in 5, 15, 25 and 120 day-old rats. By using the Cosinor method, the ontogeny of an unimodal rhythm was studied for MDH activity and soluble protein content in testis. In 5 and 15 day-old rats, the MDH acrophases were recorded around 19:00 h and 17:00 h, respectively. Rats aged 25 and 110 days showed the MDH acrophases during the dark period. An inversion of the MDH circadian rhythms was detected in 25 day-old compared to those of 5 and 15 day-old rats. An inversion of the protein circadian rhythm was also detected at 15 days compared to that at 5 days. These inversions persist in the adult rats. The amplitude of the MDH and protein rhythms reached the lowest value in adulthood. The mean daily value of testicular MDH increased between day 5 and 15, decreasing at day 35 and remaining unchanged until adulthood. The variation of malate dehydrogenase activity, soluble protein content levels, and the circadian rhythm parameters during the maturation process may be related to gonad development.     

Malate Dehydrogenase Activity of Rat Testis: Circadian Rhythm and its Ontogeny

Malate dehydrogenase activity and soluble protein content in testes from rats exposed to a 14:00 h light:10:00 h dark photoperiod, have been determined every two or four hours over a 24 hour period in 5, 15, 25 and 120 day-old rats. By using the Cosinor method, the ontogeny of an unimodal rhythm was studied for MDH activity and soluble protein content in testis. In 5 and 15 day-old rats, the MDH acrophases were recorded around 19:00 h and 17:00 h, respectively. Rats aged 25 and 110 days showed the MDH acrophases during the dark period. An inversion of the MDH circadian rhythms was detected in 25 day-old compared to those of 5 and 15 day-old rats. An inversion of the protein circadian rhythm was also detected at 15 days compared to that at 5 days. These inversions persist in the adult rats. The amplitude of the MDH and protein rhythms reached the lowest value in adulthood. The mean daily value of testicular MDH increased between day 5 and 15, decreasing at day 35 and remaining unchanged until adulthood. The variation of malate dehydrogenase activity, soluble protein content levels, and the circadian rhythm parameters during the maturation process may be related to gonad development.     

Rat liver cholesterol 7 alpha-hydroxylase. Pretranslational regulation for circadian rhythm.

A complete cDNA encoding cholesterol 7 alpha-monooxygenase (EC 1.14.13.17) which had been isolated from rat liver cDNA libraries by using specific antibodies to the enzyme (Noshiro, M., Nishimoto, M., and Okuda, K. (1989) FEBS Lett. 257, 97-100) was totally sequenced. The cDNA contained a 1,509-base pair open reading frame encoding 503 amino acid residues (Mr = 56,880) and an unusually long 3'-untranslated region rich in AT sequence in the total length of 3,545 base pairs. The predicted amino acid sequence displays less than 30% similarity to other sequenced cytochrome P-450s indicating that the 7 alpha-hydroxylase constitutes a novel family of cytochrome P-450. The AT-rich region often contained ATTTA motifs, 5'-AAT-3' or 5'-TAA-3' trinucleotides which were reported to be involved in rapidly degrading mRNA. Employing the specific antibodies and the cDNA as probes, a diurnal variation of the levels of the three factors, i.e. enzyme protein, mRNA, and enzyme activity, was studied on rat livers prepared at various times of the day. In normal animals, all three factors exhibited maximum level at 10:00 p.m. and minimum at 10:00 a.m. No significant sexual difference was observed. Cholestyramine feeding increased all three factors at 10:00 a.m. close to the maximum levels of the normal rats, but did not show a significant increase at 10:00 p.m. On the contrary, starvation markedly decreased all three factors either at 10:00 a.m. or at 10:00 p.m., while maintaining still the diurnal variation. A good correlation of the levels of mRNA to the enzyme activities and the protein levels demonstrates that pretranslational regulation is most likely a mechanism for the circadian rhythm of 7 alpha-hydroxylase. The marked diurnal fluctuation of the amount of protein and the level of mRNA also indicates their rapid turnover. The short half-life of mRNA could be correlated with the structure of the 3'-untranslated region of the mRNA characteristic of rapidly degrading mRNA, i.e. abundance of motif, AUUUA, and existence of 5'-AAU-3' or 5'-UAA-3' trinucleotides in single-stranded regions of the secondary structure.     

Regulation of neutral cholesterol esterase and acyl-CoA : cholesterol acyltransferase in the rat adrenal gland.

The activities of neutral cholesterol esterase and acyl-CoA : cholesterol acyltransferase in rat adrenal gland were measured at various time intervals over 24 h. The activity of cholesterol esterase displayed diurnal rhythm, with a major peak at the onset of darkness coinciding with the peak in the diurnal rhythm of plasma corticosterone concentration. The activity of acyl-CoA : cholesterol acyltransferase also exhibited a characteristic diurnal rhythm, with the minimum activity occurring 3 h after the onset of darkness. The profile of the rhythm exhibited by the activity of the esterifying enzyme was similar to the mirror image of the pattern of diurnal rhythm in the activity of 3-hydroxy-3-methylglutaryl-CoA reductase. Microsomal non-esterified cholesterol showed a gradual decline with a significant decrease in concentration at the onset of darkness, thus suggesting that diurnal removal of cholesterol in the environment of the esterifying enzyme and hydroxymethylglutaryl-CoA reductase leads to such diurnal decrease or increase in the activities of these two enzymes. Acute administration of corticotropin led to a 3-fold increase in the activity of cholesterol esterase, a 50% decrease in the activity of acyl-CoA : cholesterol acyltransferase and a 2-fold increase in the activity of hydroxymethylglutaryl-CoA reductase. Corticotropin administration also resulted in a significant decrease in microsomal non-esterified cholesterol and increase in plasma corticosterone concentration. These observations suggest that corticotropin plays an important part in generating the diurnal rhythm in the activities of the three enzymes.     

Diurnal Variation of Plasmatic Melatonin,Corticosterone and Variation of General Activity in Pigeons under Light-Dark Cycle and Constant Light

This work analyzed the diurnal variation of general activity and plasmatic levels of melatonin and corticosterone in pigeons submitted to a 12:00:12:00 h light-dark cycle (lights on at 6:00 a.m.) or to constant light. In both conditions pigeons were observed in 5-min sessions at times 03:00, 06:00, 09:00, 12:00, 15:00, 18:00, 21:00 and 24:00 h during two successive days. Behavior was video taped in the home cages for posterior categorization and quantification. Radioimmunoassays were used to evaluate plasmatic levels of melatonin and corticosterone. Blood samples were obtained at the times of behavioral observation. In the light-dark condition the results showed day-night variation of general activity (p < 0.001) and a robust diurnal rhythm of plasmatic melatonin (p < 0.001). Both of these variations as well as the oscillatory secretion of corticosterone disappeared under constant light condition. The parallel changes in general activity and blunting of melatonin rhythm secretion in constant light condition agree with previous evidences that melatonin may regulate behavioral oscillations in the pigeon. The present data are related to the proposition that the timing system in pigeons may involve neuroendocrine relations characterized by interactions between blood born signalization by melatonin and corticosterone.     

Extremely low frequency magnetic field exposure modulates the diurnal rhythm of the pain threshold in mice

The aim of this study was to determine whether exposure to extremely low frequency magnetic field (ELF-MF) affects the normal diurnal rhythm of the pain threshold in mice. Pain thresholds were evaluated in mice using the hot plate test. A significant increase of pain threshold during night was observed compared to that during day. This rhythm was attenuated by both constant exposure to light (LL) and constant exposure to darkness (DD) for 5 days. Under DD exposure, the diurnal rhythm in pain threshold was restored when mice were exposed to ELF-MF (60 Hz, 1.5 mT for 12 h daily, from 08:00 to 20:00 h) for 5 days. The diurnal rhythm was not reversed under dark with reversed ELF-MF cycle (exposure to 1.5 mT from 20:00 to 08:00 h, next day) for 5 days, although pain threshold in the ELF-MF exposed period of night was slightly decreased. The diurnal rhythm of melatonin analgesic effect related to pain threshold was also observed under DD by the exposure of ELF-MF for 5 days, but not for 5 nights. The present results suggest that ELF-MF may participate in the diurnal rhythm of pain threshold by acting on the system that is associated with environmental light-dark cycle.     

Mass-spectral identification and purification of phycoerythrobilin and phycocyanobilin.

The hepatic output of triacylglycerol and cholesterol from very-low-density lipoprotein (VLD lipoprotein), and the activity of 3-hydroxy-3-methylglutaryl-coenzyme A reductase were compared in the isolated perfused rat-liver preparation and in the intact rat. The output of triacylglycerol and cholesterol from VLD lipoprotein by the perfused liver was stimulated by oleate concomitant with stimulation of hepatic microsomal hydroxymethylglutaryl-coenzyme A reductase activity. In the intact animal treated with Triton WR-1339, the magnitude of secretion of triacylglycerol and cholesterol from VLD lipoprotein coincided with the diurnal rhythm of hepatic hydroxymethylglutaryl-coenzyme A reductase activity, which was maximal at 24:00 h and minimal at 12:00 h. These observations suggest that the stimulation of the reductase and of the secretion of cholesterol from VLD lipoprotein by non-esterified fatty acids, as observed with the isolated perfused rat liver preparation in vitro, may also be an important physiological mechanism in vivo. Hepatic cholesterogenesis may be stimulated under conditions conductive to the secretion of the VLD lipoprotein, the primary transport form for triacylglycerol in the postabsorptive state.     

Diurnal variation and age-related changes of bone turnover markers in female Göttingen minipigs

We investigated diurnal variation and age-related changes in bone turnover markers in female Gottingen minipigs. Ten females, 6-9 months of age, were used for confirmation of diurnal variation. Blood was collected at 3 h intervals for 24 h, and bone-specific alkaline phosphatase and intact osteocalcin (OC) levels were determined by enzyme immunoassay and radioimmunoassay, respectively. Urine was collected at 3 h intervals for 24 h using a tray attached to the bottom of the cage. The levels of N-terminal telopeptide of type I collagen (NTX) were determined by enzyme immunoassay. Pyridinoline and deoxypyridinoline were measured by high performance liquid chromatography. OC and NTX exhibited diurnal variation (Kruskal-Wallis test, P < 0.05), with the highest and lowest levels at 18:00 h (76.7 +/- 26.2 ng/ml) and 06:00 h (44.3 +/- 10.3 ng/ml), and at 03:00-05:59 h (550.4 +/- 82.4 nmol/micromol Cr) and 12:00-14:59 h (297.8 +/- 152.5 nmol/micromol Cr), respectively. In the study of age-related changes, blood and urine samples from 66 females (age range, 3-76 months) were examined to determine the bone turnover markers. All markers showed high correlations with age (0.569 < R(2) < 0.818). High levels of bone turnover markers were observed in young animals, decreasing with age (Kruskal-Wallis test, P < 0.01). The diurnal variation and age-related changes revealed in the present study will be useful in studies of bone diseases using female Gottingen minipigs.     

DIFFERENCES IN THE REGULATION OF TYROSINE AMINOTRANSFERASE IN BRAIN AND LIVER

Abstract— l -Tyrosine:2-oxoglutarate aminotransferase (EC 2.6.1.5) activity in rat brain is not regulated in the same way as in rat liver. No diurnal rhythm in the activity of the cerebral enzyme was found in rats fed ad lib. although there was a marked diurnal variation in the activity of the hepatic enzyme. In adrenalectomized rats, hydrocortisone and glucagon induced the enzyme in liver but had no effect on the enzyme in brain. In normal rats, treatment with reserpine or exposure to cold elevated the activity of the hepatic enzyme without affecting the enzyme in brain. Thus, the tyrosine aminotransferase of brain differed from the enzyme in liver since it did not exhibit diurnal variations of activity and was not affected by hormones, drugs, or stress.     

Light Regulation of Diurnal Variation of Sucrose-Phosphate Synthase in Potato

Light regulation of diurnal variation of sucrose-phosphate synthase (SPS) activity was studied in potato leaves during pre-tuber initiation stage. SPS activity was greatest at 14:00 and coincident with highest irradiance [2 000 μmol(PAR) m-2 s-1], and lowest at 18:00 during lowest irradiance (200 μmol m-2 s-1). In contrast, fructose-1,6-bisphosphatase (FBPase) activity was greatest at 12:00 (irradiance 1 700 μmol m-2 s-1). Inhibition of SPS activity by phosphorylation was least at 14:00 and highest at 18:00. Considerable inhibition was also observed at 10:00. SPS showed hyperbolic saturation kinetics in response to varying UDP-glucose concentrations. Diurnal variation of SPS activity was due to a change of affinity for its substrate (Km), but not due to change of Vmax. Phosphate (Pi) decreased the activation state of SPS. Inhibition by Pi was most severe under limiting UDP-glucose concentration. Sensitivity towards Pi inhibition at limiting substrate concentration was highest at 18:00 and lowest at 14:00. Change of tissue phosphate concentration followed the opposite pattern to that of diurnal variation of SPS activity and irradiance, being highest at 18:00 and lowest at 14:00. A dual mode of regulation is considered to exist for diurnal variation of potato leaf SPS: covalent modification and effector (Pi) regulation. Thus the change of tissue phosphate concentration may play a pivotal role in regulating potato leaf SPS. This revised version was published online in July 2006 with corrections to the Cover Date.     

The role of glucocorticoids in the regulation of the diurnal rhythm of hepatic beta-hydroxy-beta-methylglutaryl-coenzyme A reductase and cholesterol 7 alpha-hydroxylase.

The microsomal activities of the hepatic enzymes hydroxymethylglutaryl-CoA reductase and cholesterol 7 alpha-hydroxylase exhibit a diurnal rhythm with maximum activities observed during the dark period and minimum activities around noon (12:00h). This diurnal rhythm was maintained for both enzymes after adrenalectomy, but the amplitude of variation for the activity of both enzymes was greatly decreased. A single injection of cortisol administered to adrenalectomized rats 3h before the expected maximum in enzyme activity resulted in a twofold increase in the activity of both enzymes 3h later, at values similar to those observed for control rats killed at the same time. This response appeared to require protein synthesis, since it was blocked by actinomycin D. However, the administration of cortisol to adrenalectomized rats 3 h before the expected minimum did not result in significant change in the activity of hydroxymethylglutaryl-CoA reductase and cholesterol 7 alpha-hydroxylase 3 h later. Kinetic studies of cholic acid metabolism in vivo demonstrated that adrenalectomy results in a significant decrease in the rate of synthesis of cholic acid and a considerable decrease in the pool size of cholic acid and its metabolic products. Treatment of adrenalectomized rats with cortisol increased the rate oonsistent with the effects of adrenalectomy and cortisol treatment on the activity of cholesterol 7alpha-hydroxylase.     

Temporal Variation in Hepatic Superoxide Dismutase Activity in Mice

Circadian variations in superoxide dismutase (SOD) activity were determined in liver homogenates of Balb-C mice that were synchronized under controlled environmental conditions with 12 h light: 12 h dark. The activity of hepatic SOD exhibited a significant circadian rhythm, with a minimum at 01:00 h and maximum at 10:00-13:00 h. It is concluded that fluctuations in hepatic SOD activity render mice more susceptible to the toxic effects of reactive oxygen radicals at particular times of the day.     

Diurnal rhythm in mRNA expression of genes encoding amino acid transporter and circadian gene cry in intestinal mucosa of piglets

Long chain PUFA contents in plasma and liver both exhibited diurnal rhythms in pigs. However, whether mRNA expression of amino acid transporter and circadian gene Cry in intestinal mucosa is also rhythmic is yet to be known. The purpose of this study aims to investigate the diurnal rhythm in mRNA expression of genes encoding amino acid transporter and whether their rhythm was related to the expression of circadian gene Cry in intestinal mucosa of piglets. Thirty-six piglets (Duroc?×?Landrace?×?Large Yorkshire) at the age of 35 days were selected and fed for three weeks, and then samples were collected at 3:00 am (Clo3), 7:00 am (Clo7), 11:00 am (Clo11), 3:00 pm (Clo15), 7:00 pm (Clo19), and 11:00 pm (Clo23) at the age of 56 days. At each time point, small intestinal mucosa samples were collected from duodenum, jejunum, and ileum for detection of mRNA expression of the amino acid transporters and circadian gene Cry. The results showed that mRNA expression of most amino acid transporters in intestinal mucosa was higher at night and lower during the daytime. Expression of SLC1A2, SLC6A20, SLC7A1, and SLC6A14 in duodenal mucosa reached the peak at Clo3 and Clo7; the diurnal rhythm of expression of SLC1A2, SLC6A20, and SLC7A1 was similar to Cry1, while the diurnal rhythm of expression of SLC6A14 had a similar trend to Cry2. Expression of SLC16A10, SLC1A2, and SLC7A1 in jejunal mucosa reached the peak at Clo7, while SLC6A14 reached the peak at Clo3; the diurnal rhythm of expression of SLC1A2 showed a similarity with Cry1, while the diurnal rhythm of expression of SLC16A10, SLC7A1, and SLC6A14 was similar to Cry2. Expression of SLC6A14, SLC6A20, and SLC7A1 in ileal mucosa reached the peak at Clo3; the diurnal rhythm of expression of SLC6A20 has a similarity with Cry1, while the diurnal rhythm of expression of SLC7A1 and SLC6A14 was similar to Cry2. The results suggested that the mRNA expression of most genes encoding amino acid transporters exhibited diurnal rhythms in the intestinal mucosa of piglets, and SLC7A1, SLC6A14, and SLC1A2 have a similar rhythm with circadian clock genes Cry1 and 2, and they reached the peak at Clo3 and Clo7.     

Clinical pharmacokinetics of 5-fluorouracil with consideration of chronopharmacokinetics

Even though 5-fluorouracil (FU) is one of the oldest anticancer drugs, its use in cancer chemotherapy continues to increase. Fluorouracil is a pro-drug that requires intracellular activation to exert its effects. This makes it difficult to associate blood drug concentration with cell toxicity directly, although data from the literature show the existence of such a relationship. The relationship between FU pharmacokinetics and patient response has been explored extensively and reports attest a link between systemic drug exposure and response and survival. This has led to the concept of maximal tolerated exposure, and strategies to achieve this rely on pharmacokinetic follow-up and individual dose adjustment. More than 80% of the administered FU dose is eliminated by catabolism through dihydropyrimidine dehydrogenase (DPD), the rate-limiting enzyme. Dihydropyrimidine dehydrogenase activity is found in most tissues but is highest in the liver. Peripheral blood mononuclear cells (PBMC) are used to monitor clinically DPD activity. A significant, but weak correlation between PBMC and liver DPD activity has been observed. The relationship between PBMC-DPD activity and FU systemic clearance is weak (r²=0.10); thus, simply determining PBMC-DPD is not sufficient to predict accurately FU clearance. Population pharmacokinetic analysis identified patient co-variables that influence FU clearance; drug kinetics is significantly reduced by increased age, high serum alkaline phosphatase, length of drug infusion, and low PBMC-DPD. Autoregulation of FU metabolism also is suggested; inhibition of DPD activity was observed after FU administration in both colorectal cancer patients and an animal model. Circadian rhythmicity in DPD activity is suggested from both human and animal investigations. In patients receiving protracted low dose 5-FU infusion, the circadian rhythm in FU plasma concentration peaks at 11:00h and is lowest at 23:00h, on average. The inverse relationship observed between the circadian profile of FU plasma concentration and PBMC-DP activity in these same patients suggests a link between DPD activity and FU pharmacokinetics. The impact of the biological time of drug administration was also studied with short venous infusions; clearance was 70% greater at 13:00h than at 01:00h. Similarly, peak drug concentration occurred in the first half of the night in patients receiving constant rate 5-FU infusion for 2-5 d. Several studies describe wide interindividual variation in the timing of the peak and trough of the 24h rhythm in DPD activity. The rational for FU chronomodulated therapy has been the circadian rhythm in host drug tolerance, which is greatest during the night time when the proliferation of normal target tissue is least. A randomized study of chronomodulated FU therapy with maximal delivery rate at 04:00h was shown clearly to be significantly more effective and less toxic than control flat FU therapy. Future research must focus on easy-to-obtain markers of specific rhythms to individualize the chronomodulated FU delivery.