Control strategy and methods for continuous direct compression processes.

We presented a control strategy for tablet manufacturing processes based on continuous direct compression. The work was conducted by the experts of pharmaceutical companies, machine suppliers, academia, and regulatory authority in Japan. Among different items in the process, the component ratio and blended powder content were selected as the items requiring the control method specific to continuous manufacturing different from the conventional batch manufacturing. The control and management of the Loss in Weight (LIW) feeder were deemed the most important, and the Residence Time Distribution (RTD) model were regarded effective for setting the control range and for controlling of the LIW feeder. Based on these ideas, the concept of process control using RTD was summarized. The presented contents can serve as a solid fundament for adopting a new control method of continuous direct compression processes in and beyond the Japanese market.

Selenocysteine beta-lyase and methylselenol demethylase in the metabolism of Se-methylated selenocompounds into selenide.

The lyase activity toward Se-methylated selenoamino acids and the demethylase activity toward methylselenol in the metabolism of selenium were characterized in vitro. The beta- and gamma-lyase activities toward selenomethionine (SeMet) and Se-methylselenocysteine (MeSeCys), respectively, were compared under exactly identical conditions by incubating 77Se-SeMet and 76Se-MeSeCys simultaneously in a liver supernatant, and then estimated by the decreases in the labeled starting selenoamino acids (MeSeCys and SeMet), and also by the increases in the labeled enzyme products (methylselenol and selenide) after oxidation to methylseleninic acid (MSA(IV)) and selenite, respectively, by HPLC-inductively coupled plasma-mass spectrometry (ICP-MS). Only 76Se-MeSeCys was decreased and only 76Se-selenite was produced, suggesting that conversion of MeSeCys to methylselenol by beta-lyase followed by that of methylselenol to selenide by demethylase actively occurred in the liver supernatant. The demethylase activity was characterized by incubating 77Se-methylselenol produced in situ from 77Se-MSA(IV) and glutathione in a partially purified enzyme preparation. It was found that demethylation takes place directly through an attack by a hydroxide anion on the methyl group of methylselenol producing selenide and methanol, selenide being detected on HPLC-ICP-MS after oxidation to selenite, and methanol on GC-MS. It was concluded that beta- but not gamma-lyase activity could be detected in a liver supernatant, and that the resulting methylselenol product is demethylated through hydrolysis, with methanol and selenide being produced (MeSeCys-->CH3SeH-->HSeH + CH3OH).

Simultaneous tracing of 76Se-selenite and 77Se-selenomethionine by absolute labeling and speciation.

Nutritional selenocompounds are transformed into the assumed common intermediate selenide, which is utilized for the synthesis of selenoenzymes or transformed into methylated metabolites for excretion. Hence, selenocompound metabolites can be traced only with labeled selenium. Here we applied a new tracer method for the metallomics of biometals using simultaneous speciation of each metallome labeled with different homo-elemental isotopes to metabolism and availability of selenium. Rats were depleted of endogenous natural abundance selenium by feeding a single selenium stable isotope ((82)Se-selenite) and then administered (76)Se-selenite and (77)Se-selenomethionine ((77)Se-SeMet)simultaneously. Biological samples were subjected to quantification and speciation analysis by HPLC-ICPMS. Metabolites of the labeled (76)Se and (77)Se and interaction with endogenous selenium were traced and examined without interference from the corresponding endogenous natural abundance isotopes. Differences in the distribution and metabolism among organs and between the two nutritional selenocompounds were compared under exactly identical biological and analytical conditions: (1) selenite was distributed more efficiently than SeMet in organs and body fluids except the pancreas. (2) SeMet was taken up by organs in its intact form. (3) Selenium of SeMet origin was distributed selectively in the pancreas and mostly bound to a protein together with intact SeMet. (4) Selenosugars A and B but not trimethylselenonium (TMSe) were detected in the liver. (5) Selenosugar B and TMSe were detected in the kidneys.

Metabolic transformation of methylseleninic acid through key selenium intermediate selenide.

Methylseleninic acid (MSA(IV)) [CH(3)Se(O)OH] is readily reducible to methylselenol [CH(3)SeH], the assumed lyase metabolite and the proposed biologically active form of methylated selenoamino acids. At the same time, MSA(IV) is an oxidation product of the major urinary metabolite selenosugar. (77)Se-Enriched MSA(IV) was injected intravenously into rats (25 microg Se/kg body weight), and urine, blood and liver were obtained at five time points after the injection. Time-related changes in the concentration of (77)Se were determined together with speciation analysis of the labeled metabolites. (77)Se was mostly moved into red blood cells (RBCs) within 10 min, and then redistributed into organs within 30 min. Excessive (77)Se taken up by the liver was first detected as selenosugar A and then as B, suggesting that MSA(IV) was transformed to selenide, and then to selenosugar A followed by methylation to selenosugar B (urinary metabolite). (77)Se was incorporated also into selenoproteins (most efficiently to plasma selenoprotein P that is synthesized in liver), suggesting that MSA(IV) is utilized for the synthesis of selenosugar (for excretion) and selenoproteins (for utilization) through selenide. In vitro experiments with simultaneous incubation of (77)Se-MSA(IV) and (82)Se-selenite in a RBC suspension revealed the precise difference in the metabolism between MSA(IV) and selenite in RBCs. (77)Se excreted into the urine was mostly detected as selenosugar but with a distinct amount of trimethylselenonium, suggesting that selenosugar and trimethylselenonium are produced depending on the capacity to transform methylselenol to selenide. MSA(IV) was suggested to be reduced to methylselenol (allowing the production of a proposed active form of selenium), and then transformed (demethylated) to selenide for utilization and excretion.

Selenosugar and trimethylselenonium among urinary Se metabolites: dose- and age-related changes.

Once selenium (Se) is absorbed by the body, it is excreted mostly into the urine and the major metabolite is 1beta-methylseleno-N-acetyl-d-galactosamine (selenosugar) within the required to low-toxic range. Selenosugar plateaus with a dose higher than 2.0 microg Se/ml water or g diet, and trimethylselenonium (TMSe) starts to increase, indicating that TMSe can be a biomarker of excessive and toxic doses of Se. Here, we show dose-related changes in the two urinary Se metabolites to clarify the relationship between the dose and urinary metabolites by feeding selenite to rats. It was also examined whether the metabolites are related to age, and further whether a possible exogenous source of the N-acetyl-d-galactosamine moiety, chondroitin 4-sulfate, affects the urinary metabolites. Selenite in drinking water was fed ad libitum to male Wistar rats of 36 and 5 weeks of age, and the concentrations of Se in the urine and organs were determined together with speciation of the urinary Se metabolites. In young rats, selenosugar was always the major urinary metabolite and TMSe increased with a dose higher than 2.0 microg Se/ml drinking water. On the other hand, in adult rats, TMSe increased only marginally despite that the rats suffered much more greatly from the Se toxicity, suggesting that TMSe cannot be a biomarker of Se toxicity. The results suggest that sources of the sugar moiety of selenosugar are more abundant in adult rats than in young rats. Chondroitin 4-sulfate did not affect the ratio of the two urinary metabolites, suggesting that the sugar source is of endogenous origin and that it increases with age.