Age-related changes in the metabolism and excretion of allyl isothiocyanate. A model compound for glutathione conjugation

The objective of this study was to evaluate age-related changes in the in vivo metabolism and excretion of allyl isothiocyanate (AITC), which is used as a model compound to assess glutathione conjugation. AITC is a constituent of oil of mustard and a food additive that has been shown to induce transitional cell papillomas in the urinary bladder of male Fischer rats. Male Fischer rats, ages 3, 16, and 27 months, were dosed orally with 14C-AITC (25 mg/kg). Urine, feces, volatiles, and expired air were collected for 72 hr. Biliary excretion was also examined after iv administration of 10 mg/kg of 14C-AITC. There was no significant difference between these age groups in the percentage of the total dose of AITC eliminated in urine, which is the major route of excretion. The percentage of the dose excreted as volatiles increased in the 27-month animals. These animals also showed a decrease in the production of 14CO2. An age-related decrease in fecal excretion of AITC-derived radioactivity was observed. However, the percentage of the dose excreted in bile increased at 16 months and then decreased in the senescent rats as compared to the 3-month age group. The age-related decrease in 14CO2 production suggests a change in the oxidative metabolism of AITC in senescent animals. However, there was no change in the relative amounts of mercapturic acid excreted in urine, suggesting that glutathione conjugation was unchanged with age.(ABSTRACT TRUNCATED AT 250 WORDS)     

Studies on benzyl acetate. III. The percutaneous absorption and disposition of [methylene-14C]benzyl acetate in the rat

[methylene-14C]Benzyl acetate was applied over an area of 6.25, 12 or 18 cm2 to the shaved backs of male Fischer 344 rats under an occlusive dressing at dose levels of 100, 250 and 500 mg/kg. The compound was administered either as the neat substance or as a 50% (v/v) solution in ethanol. After 6 hr the dressing was removed, the shaven area was washed with ethanol and the dressing and washings were counted for 14C. Urine and faeces were collected for 72 hr from the start of treatment and urinary metabolites were assayed by radio-TLC and HPLC. Following administration of the neat compound, a significant proportion of the dose was recovered from the application site (28-48%) and a similar proportion (28-46%) was absorbed and excreted in the 0-24-hr urine. Excretion of 14C in the urine over 0-24 hr accounted for c. 95% of absorbed 14C in all cases, and total recovery of radioactivity was 79-84% with less than 2% of the dose present in the carcass at the end of the experiments. The extent of absorption of benzyl acetate per unit area of skin, as assessed by the recovery of its metabolites in urine, rose with increasing concentration (mg/cm2) of the test compound on the skin. The absorption of topically applied benzyl acetate was essentially the same when the dose was administered in a 50% ethanolic solution. In all cases, the major urinary metabolite was hippuric acid (c. 95% of urinary 14C), together with much smaller amounts of benzoyl glucuronide, benzoic acid and benzylmercapturic acid. The distribution of 14C in the tissues was examined 6 and 24 hr after the topical application of 5 mg [methylene-14C]benzyl acetate/kg as a 1% (v/v) solution in ethanol to rats. Radioactivity in all carcasses was less than 4% of the administered dose and levels in all the organs examined were lower at 24 than at 6 hr.     

Availability of glycine and coenzyme A limits glycine conjugation in vivo.

Using benzoic acid as substrate, this study tested the hypothesis that capacity limitation of glycine conjugation in vivo is due to substrate-induced depletion of hepatic cosubstrates (i.e., ATP, coenzyme A, and glycine) utilized in the conjugation reaction. Benzolyglycine formation was investigated by following the disappearance of benzoic acid from blood and appearance of benzoylglycine in blood and urine after administration of sodium benzoate (0.2-2 mmol/kg, iv) to anesthetized rats whose urine formation was stimulated by mannitol administration. Capacity limitation of glycine conjugation is indicated by (a) the gradual dose-dependent reduction of benzoate blood clearance from 39 ml/min/kg at 0.2 mmol/kg benzoate to 3.7 ml/min/kg at 2 mmol/kg, and (b) the tendency to attain maximal blood levels and urinary excretion rates of benzoylglycine after administration of 0.5-1 mmol/kg benzoate. The maximal urinary excretion rate of benzoylglycine after benzoylglycine administration exceeded the maximal excretion rate of endogenously formed benzoylglycine (approximately 5 mumol/kg/min) 5-fold. This suggests that the urinary excretion rate of endogenously formed benzoylglycine reflects the rate of its formation. Benzoate depleted hepatic glycine (to 40%) and coenzyme A (to 14%) in a dose-dependent fashion; however, it did not change ATP levels in liver. The pattern of this dose-dependent cosubstrate depletion suggests that benzoate primarily causes consumption of hepatic glycine which, at high substrate dosage, leads to marked depletion of coenzyme A in the liver. Thus, these observations indicate that capacity-limited glycine conjugation may be due to limited availability of glycine and coenzyme A for the conjugation process.     

A linear recirculation model for drug disposition.

A new approach to the modeling of drug disposition is described. Disposition is regarded as the result of repetitive passes of the drug around the circulation. Mathematical analysis of experimental blood concentration data yields an expression describing the kinetics of a single pass through the tissues. In physicochemical terms the single-pass behavior depends to a large extent on the interaction of the drug with individual tissues, which greatly simplifies interpretation. The method may reveal features of disposition not apparent from experimental blood concentration data.     

Age-related changes in disposition and metabolism of benzene in male C57BL/6N mice.

Benzene disposition and metabolism were examined as a function of age in male C57BL/6N mice aged 3 and 18 months. Mice received a single oral dose of either 10 or 200 mg/kg 14C-benzene (approximately 25 microCi/kg). Excretion of 14C-derived benzene radioactivity (RA) was monitored in urine, feces, and as exhaled 14CO2 from 0 to 72 hr, and as exhaled unmetabolized benzene from 0 to 6 hr. At 10 mg/kg 14C-benzene, urinary elimination was the major route of excretion in both 3- and 18-month mice. Urinary excretion of 14C-derived benzene RA was significantly decreased in 18- vs. 3-month mice at 4, 6, 24, and 48 hr, while fecal excretion was significantly increased at 72 hr. Elimination of 14C-benzene as 14CO2 and unmetabolized 14C-benzene was also increased in 18- vs. 3-month mice at this dose. Hydroquinone glucuronide (HQG), phenylsulfate (PS), and muconic acid (MUC) were the major urinary metabolites at 10 mg/kg 14C-benzene in both 3- and 18-month mice, representing approximately 40, 28, and 15% of an administered dose of 14C-benzene. Smaller amounts of phenyl glucuronide (4.0%), pre-phenyl mercapturic acid (1.2%), and catechol glucuronide (0.5%) were also detected. No significant differences were found with age in the percentage of an administered dose of benzene excreted as the various metabolites at 10 mg/kg. At 200 mg/kg 14C-benzene, the total percentage of 14C-derived benzene RA eliminated in urine within 72 hr was not significantly different with age, but elimination at early time points (4, 6, and 8 hr) was significantly decreased in 18- vs. 3-month mice.(ABSTRACT TRUNCATED AT 250 WORDS)     

Drug-protein conjugates--X. The role of protein conjugation in the disposition of dinitrofluorobenzene

The metabolism and irreversible protein binding of 2,4-[3,5-3H]dinitrofluorobenzene (3H-DNFB), a model chemically reactive compound, were studied in the rat. 3H-DNFB given intravenously (5 micrograms, 5 mg or 25 mg per kg) to anaesthetized cannulated rats was rapidly metabolized via the mercapturic acid pathway. The metabolites were extensively eliminated in bile and urine: predominantly as the glutathione conjugate and mercapturate in bile, and as the mercapturate in urine. Only ca. 3-10% of the doses remained in the liver, kidneys, spleen, heart and lungs at 3 hr. Dinitrophenyl mercapturate was the principal urinary metabolite in conscious rats dosed i.v. (5 mg or 25 mg per kg). Only 15-25% of the radiolabelled material in liver and kidney at 3 hr was irreversibly bound to protein, but 45-99% of that in the other organs and 49-88% in plasma was irreversibly bound. Preliminary evidence for the metabolism of 3H-DNFB (5 mg/kg and 25 mg/kg doses) to N2-acetyl-N6-DNP-lysine, a novel conjugate and metabolite of dinitrophenylated proteins in vivo, is presented.     

Age-related changes in the manifestations of tuberculosis. Implications for drug therapy.

Although the overall number of tuberculosis cases reported in developed countries has decreased markedly since about the middle of the twentieth century, at present active tuberculosis in the geriatric population is highly prevalent even in developed countries. With an increase in the aged population and striking progress in medicine during the past two decades, the development of tuberculosis infections and the active disease has been gradually changed; namely, the number of older people at risk of infection has been increasing. In the elderly, postprimary pulmonary tuberculosis and miliary tuberculosis are particularly difficult to diagnose. A high index of suspicion, a tuberculin skin test, and appropriate laboratory studies are essential for early diagnosis. In choosing the proper therapy for tuberculosis in the elderly, efficacy, patient compliance, and toxicity must be considered. During this decade, more so than ever before, much time and work will be required to more completely understand the factors that place a patient at risk for infection and to accomplish eradication of tuberculosis.     

A linear recirculation model for drug disposition

A new approach to the modeling of drug disposition is described. Disposition is regarded as the result of repetitive passes of the drug around the circulation. Mathematical analysis of experimental blood concentration data yields an expression describing the kinetics of a single pass through the tissues. In physicochemical terms the single-pass behavior depends to a large extent on the interaction of the drug with individual tissues, which greatly simplifies interpretation. The method may reveal features of disposition not apparent from experimental blood concentration data.     

Studies on benzyl acetate. I. Effect of dose size and vehicle on the plasma pharmacokinetics and metabolism of [methylene-14C]benzyl acetate in the rat

Male Fischer 344 rats received [methylene-¹⁴C]benzyl acetate by gavage in a dose of 5, 250 or 500 mg/kg, as the neat substance, in corn oil or in propylene glycol. Urine and faeces were collected and urinary metabolites were assayed by radio-TLC and HPLC. Other animals were killed at various times and exsanguinated, and plasma levels of ¹⁴C in plasma occurred earliest and were highest when benzyl acetate was given neat. Peak levels were lower and absorption was delayed with the propylene glycol vehicle. The use of corn oil as the dose vehicle at the higher doses (250 and 500 mg/kg) led to the maintenance of plateau plasma levels, at about one half of the peak levels seen with the neat compound, for up to 8 hr after administration. At the 5 mg/kg dose, the plasma levels of ¹⁴C were essentially the same whether the dose was given in corn oil or propylene glycol. At the 250- and 500-mg/kg doses, at all time points, the major metabolite in plasma was benzoic acid, accompanied by smaller amounts of hippuric acid. Benzyl alcohol was also detected in some plasma samples. At the 5-mg/kg dose, the major plasma metabolite was hippuric acid, together with a smaller amount of benzoic acid. When propylene glycol was used as the vehicle at this dose level, benzylmercapturic acid was also present in the plasma. The major urinary metabolite was hippuric acid (c. 66% of the dose), with benzoic acid (2%) and benzylmercapturic acid (1%) also present. The elimination of benzoyl glucuronide increased with increasing dose, from c. 3 to 11% of the dose.     

Age-related changes in hepatic function. Implications for drug therapy.

An age-related decrease in the hepatic clearance of many drugs has been reported. Several mechanisms have been proposed, but only some are supported by hard evidence. Liver volume declines with age, as does hepatic blood flow--changes which may largely account for the reduced clearance of capacity- and flow-limited drugs, respectively. Age-related histological changes in the liver are minor and of uncertain significance; standard liver function tests do not change significantly with aging. There is, as yet, no direct evidence of a generalised fall in hepatic drug-metabolising enzyme activities in aging humans measured in vitro, but some in vivo studies suggest that certain very specific cytochrome P450 isoenzymes may be affected by aging, especially in men. Finally, there may be an age-related decline in the response to environmental influences.     

Age-related changes of vitamin A status

Ageing is an independent risk factor for the development of cardiovascular disease. The ageing process is known to be associated with increased oxidative stress and an increased risk for cardiovascular and other diseases, such as cancer. To delay this process, therapeutic strategies involving the use of naturally occurring antioxidants, such as vitamin A, have gained considerable interest. Therefore, we wanted to investigate in a model of mammalian ageing whether changes in tissue and plasma levels of vitamin A occur with increasing age. This would constitute a prime rationale for its dietary supplementation. Experiments were performed in three different age groups (4-6 months old, 19 months old, 32-35 months old) of F1 (F344 x BN) healthy male rats that were fed a normal diet without any additional supplementation. Vitamin A and carotenoids in plasma and major organs were measured by reverse-phase high-performance liquid chromatography. In 3-year-old rats, vitamin A levels were found to be decreased in plasma (P < 0.0001) as compared with young and middle-aged animals. However, they were markedly increased in the main storage organ (ie, the liver) (P < 0.01-0.0001), and also in the aortic vessel wall. They were undetectable in the heart, irrespective of age. Increased tissue levels of vitamin A, especially in the vasculature, may be part of an age-associated self-regulatory process of adaptation, possibly as a counter-regulation against oxidative tissue damage. Based upon the assumption that in elderly humans, as in our animal model, a similar demand-regulated mechanism may work independently of additional dietary vitamin A supplementation, one may question the strategy of large clinical interventional trials using vitamin A or its derivatives beyond normal dietary intake.     

Correlation of thermodynamic activity and vapour diffusion through human skin for the model compound, benzyl alcohol

This work tested the potential for predicting percutaneous absorption rates of a volatile penetrant from any vehicle by using thermodynamic activity measurements. Benzyl alcohol was chosen as a non-ideal, hydrogen bonding, volatile model penetrant. A manual headspace gas chromatography method measured benzyl alcohol vapour concentrations and thermodynamic activities above binary mixtures with vehicles: butanol, butyl acetate, isopropyl myristate, isophorone, toluene and propylene carbonate. Benzyl alcohol vapour diffusion through human, abdominal skin was also measured in-vitro for these mixtures. The benzyl alcohol vapour flux was linearly related to the activity, suggesting that percutaneous absorption is controlled by thermodynamic activity when the vehicle has no effect on the stratum corneum barrier.     

Age-related changes in trimethadione oxidizing capacity.

The metabolism of trimethadione (TMO) following oral administration (4 mg kg-1) has been studied in 11 young male and 11 elderly male patients. The elimination half-life (h) of TMO was 11.8 +/- 1.4 (mean +/- s.e. mean) in the young and 23.5 +/- 2.17 in the elderly (P less than 0.01). Total body clearance (1 h-1 kg-1) was 41.4 +/- 2.8 in the young and 29.0 +/- 2.4 in the elderly (P less than 0.01). The apparent volume of distribution (1 kg-1) was 0.67 +/- 0.03 in the young and 0.65 +/- 0.04 in the elderly. Serum dimethadione (DMO)/TMO ratios at 4 h were 0.65 +/- 0.03 in the young and 0.46 +/- 0.03 in the elderly (P less than 0.01). These results suggest that N-demethylation of TMO is inversely related to age.     

A physiological pharmacokinetic model for morphine disposition in the pregnant rat

A physiological model was used to examine the disposition of morphine in the pregnant rat. In the model was incorporated an expression of both a linear and a nonlinear binding term of morphine to the maternal muscular tissue. Furthermore, the experimental data suggested that a diffusion-limited transport of morphine occurred across the placenta. Morphine showed a relatively high partition into the maternal kidney and muscle tissues. The concentration of morphine in the foetus was about 1.5 times higher than that of the maternal plasma, whereas the foetal brain concentration was about 4 times higher than that of the maternal plasma. The influence on morphine disposition by changes in both the tissue binding of the maternal muscle and the placental plasma flow was explored by model simulations. Due to the diffusionlimited transport of morphine across the placenta, a change in the placental plasma flow would only have an effect on the concentration-time profile of morphine in the foetal tissues if the plasma flow approached and became less than the diffusion clearance across the placenta. An increase in the partition of morphine into the maternal muscle produced an increase in the terminal half-life in all tissues including the foetus.     

Effect of chlorophenoxyacetic acid herbicides on glycine conjugation of benzoic acid.

1. 2,4-Dichlorophenoxyacetic acid (2,4-D) and 2,4,5-trichlorophenoxyacetic acid (2,4,5-T) (0.1-0.5 mmol/kg i.p.) delayed the disappearance of injected benzoate from blood and diminished the urinary excretion of the formed benzoylglycine, but elevated the blood levels of benzoylglycine in rat, suggesting that these herbicides interfere with both the formation and the renal transport of benzoylglycine. 2. Inhibition of the renal excretion of benzoylglycine by 2,4-D or 2,4,5-T (0.5 mmol/kg i.p.) was directly demonstrated in rat injected with benzoylglycine. 3. Inhibition of benzoylglycine formation from benzoic acid by 2,4-D or 2,4,5-T (0.5 mmol/kg i.p.) was directly demonstrated in renal pedicles-ligated rats injected with benzoate. 4. Neither 2,4-D nor 2,4,5-T influenced the hepatic concentrations of ATP, coenzyme A (CoA) or glycine; therefore, it is unlikely that they inhibit glycine conjugation of benzoic acid by diminishing the availability of co-substrates. 5. Although the chlorophenoxyacetic acids did not appear to be a substrate for the mitochondrial acyl-CoA synthetases, both 2,4-D and 2,4,5-T diminished the activity of benzoyl-CoA synthetase (but not that of benzoyl-CoA:glycine N-acyltransferase) in solubilized hepatic mitochondria. These findings suggest that 2,4-D and 2,4,5-T impair benzoylglycine formation in rat by inhibiting benzoyl-CoA synthetase.     

Age-related differences in ophthalmic drug disposition III. Corneal permeability of pilocarpine in rabbits

In the two previous papers in this series, age-related differences in ocular tissue size and protein binding were investigated relative to the distribution of drug within the eye. The present paper deals with another potentially important determinant of ocular drug disposition, namely corneal permeability. An in vitro technique was utilized to study the corneal permeability of pilocarpine in rabbits of two different ages. The effect of pH on pilocarpine transport was also examined. Results indicate that the transport of ionized pilocarpine contributes significantly to the overall corneal permeation of pilocarpine. There appears to be no difference in the transport characteristics in corneas of the two ages as far as ionized pilocarpine is concerned. However, the corneas of the younger animals are more permeable to non-ionized pilocarpine than those of 60-day-old rabbits. These results suggest that different pathways or layers in the cornea are rate-limiting for the transport of ionized and unionized pilocarpine. Furthermore, the significant transport of ionized species observed in these studies indicates that the pH-partition hypothesis is not totally adequate to explain the corneal transport of pilocarpine in this particular system.     

Age-related changes in the pharmacokinetics and pharmacodynamics of nifedipine.

1. The effects of age on the pharmacology of nifedipine were investigated in 11 young and six elderly normotensive volunteers. 2. Following 2.5 mg of nifedipine i.v. the plasma clearance of nifedipine was 348 +/- 83 (s.d.) ml min-1 in the elderly compared with 519 +/- 125 ml min-1 in the young (P less than 0.05) and the AUC in the elderly was significantly greater at 125 +/- 28 ng ml-1 h compared with 83.9 +/- 19 ng ml-1 h (P less than 0.05). The Vss was similar in both age groups. 3. Following 10 mg oral sustained release nifedipine the AUC was 281 +/- 64 ng ml-1 h in the elderly compared with 136 +/- 56 ng ml-1 h in the young (P less than 0.002) and Cmax in the elderly was significantly greater at 36.8 +/- 11.8 ng ml-1 compared with 22.3 +/- 5.8 ng ml-1 (P less than 0.05). The trend towards an increased bioavailability in elderly subjects (36%) was supported by a significantly lower nitropyridine metabolite/nifedipine ratio in the elderly. 4. Absorption rate limited kinetics of the sustained release formulation were indicated by the prolonged t1/2 compared with i.v. administration. In the elderly t1/2 (oral) was significantly greater than in the young (elderly 6.7 +/- 2.2 h, young 3.8 +/- 1.4 h, P less than 0.05). 5. Haemodynamic changes in the young were confined to a tachycardia following i.v. administration. In the elderly, supine BP fell significantly following both oral and i.v. nifedipine while the heart rate remained unchanged.(ABSTRACT TRUNCATED AT 250 WORDS)