Optimizing the lyophilization cycle and the consequences of collapse on the pharmaceutical acceptability of Erwinia L-asparaginase.

The antileukemia enzyme, Erwinia L-asparaginase, occurs as a tetramer which can be dissociated by the stresses of lyophilization into four subunits (subunit M(r) 34 000 Da). Dissociation can be reduced by adding protectants to the formulation to stabilize the biopolymer, while the product should dry to form a pharmaceutically elegant, shelf-stable cake which is readily soluble. Using analytical ultracentrifugation, HPLC, and circular dichroism we have related structural dissociation of the enzyme during lyophilization to biological activity. Additives such as mannitol prevent ablation loss of vial contents and dry to form cosmetically elegant cakes but provide little biological protection, since during freezing they crystallize and are removed from the preparation. Excipients persisting throughout the cycle in the amorphous state provide improved biological protection, although high molecular weight compounds such as Dextran (M(r) 70000 Da) are most effective only during product freezing or storage. Low molecular weight sugars are protective throughout the cycle although formulations containing monosaccharides often exhibit low collapse temperatures (Tc) measured using a freeze-drying microscope or glass transition temperatures (Tg') measured by thermal analysis, but these formulations distort as drying progresses to form a collapsed, cosmetically unacceptable cake, with reduced activity, poor stability, a high moisture content, and reduced solubility. Collapse can be avoided by formulating with disaccharides, which display higher Tc temperatures than monosaccharides, or drying below Tc. Dried samples which persist in the amorphous state can also collapse when stored above their solid-state collapse temperatures when they decay at a faster rate than predicted by Arrhenius kinetics. The solid-state collapse temperature can be significantly decreased by the diffusion of moisture from the stopper into the dry product resulting in an increase in sample water content. Lyophilization cycle times can be reduced by analyzing collapse characteristics so that the relationship between product temperature and chamber pressure can be controlled so that drying rates can be optimized while ensuring that the product does not melt or collapse during sublimation.

Alpha 2-adrenoceptors in the guinea-pig uterus: heterogeneity in the circular and longitudinal smooth muscle layers.

Homogenate binding and functional studies have been undertaken to investigate the role of alpha 2-adrenoceptors on the circular and longitudinal myometrial layers of the dioestrous guinea-pig uterus. Each myometrial layer contained a single population of [3H]rauwolscine binding sites (KD values approximately 3 nM) for which yohimbine exhibited a higher affinity than prazosin, and xylazine a higher affinity than phenylephrine, indicating the presence of alpha 2-adrenoceptor binding sites. In circular myometrium, xylazine enhanced contractile responses, was more potent than either noradrenaline or phenylephrine in inhibiting forskolin-stimulated accumulation of cyclic AMP, and reduced the inhibitory effect of forskolin on phenylephrine-induced phosphatidyl inositol hydrolysis. In longitudinal myometrium xylazine enhanced contractile responses to phenylephrine but did not inhibit forskolin-stimulated accumulation of cyclic AMP. We conclude that alpha 2-adrenoceptor binding sites are present in both uterine layers and mediate uterine contractility possibly through different mechanisms.

Effects of inert fat on energy balance, plasma concentrations of hormones, and reproduction in dairy cows.

The objective was to determine the effects of dietary inert fat on estimated energy balance, hormones in plasma, and reproduction during early lactation. From wk 0 to 12 postpartum, 14 pluriparous Holstein cows were fed individually a TMR, and blood samples were taken twice weekly for quantification of IGF-I, progesterone, and cholesterol. During wk 5 to 12, one-half of the cows remained on the TMR, and the other half were fed the TMR containing inert fat at 1.8% of dietary DM. Estrous behavior was monitored twice daily, and body condition scores were recorded every 4 wk. Cows fed inert fat between wk 5 and 12 postpartum had similar concentrations of IGF-I in plasma but greater luteal phase progesterone secretions than cows fed the control diet. Total cholesterol in plasma also was greater in cows fed inert fat than in cows fed the control diet. Intervals to first, second, and third ovulation or estrus did not differ among cows fed control or inert fat diets. Body condition scores, daily DMI, and milk production were not affected by dietary inert fat. Inert fat fed to cows between wk 5 and 12 postpartum did not affect ovulatory activity but may enhance luteal activity.

Insulin-like growth factor-I in dairy cows: relationships among energy balance, body condition, ovarian activity, and estrous behavior.

The objective was to determine relationships among concentrations of insulin-like growth factor-I in serum, energy balance, body condition, and ovarian activity during early lactation. From 0 to 12 wk postpartum, 11 pluriparous Holstein cows were individually fed a total mixed diet and bled twice weekly for quantification of insulin-like growth factor-I and progesterone. Estrous behavior was monitored twice daily and body condition scores were recorded weekly. Individual feed intake and milk production were recorded daily and milk composition and BW were measured weekly. Cows with positive (3.43 Mcal/d; n = 5) energy balance during the first 12 wk postpartum had greater concentrations of insulin-like growth factor-I in serum and greater luteal-phase progesterone secretion than in cows with negative (-1.69 Mcal/d; n = 6) energy balance. Interval to first ovulation or first estrus did not differ between cows with positive versus negative energy balance. Correlation coefficients among weekly averages revealed positive correlations between energy balance and serum progesterone, and energy balance and serum insulin-like growth factor-I. Body condition scores were not correlated with energy balance or serum insulin-like growth factor-I. We conclude that reduced luteal activity that accompanies negative energy balance may be associated with reduced concentrations of insulin-like growth factor-I in serum.

Human adjuvant disease. A new cause of chylous effusions.

Connective-tissue disease occurring after cosmetic surgery with silicone injections or implants has been reported. This disorder has been called human adjuvant disease. One patient is described in whom a chylous effusion and systemic lupus erythematosus-like disorder developed after mammary augmentation with silicone gel-filled prostheses. This patient represents still another example of human adjuvant disease. A brief review of the literature regarding human adjuvant disease is also presented.

Optimal radiotherapy treatment planning for carcinoma of the cervix uteri.

A study is described in which an optimal time-dose-fractionation (TDF) value for radiotherapy of carcinoma of the cervix uteri is found using clinical data for points 2 cm from the cervical os proximally along the direction of the cervical canal and 2 cm laterally to the canal. The optimal TDF value was found to be 127 +/- 10 for regimens utilizing external beam cobalt 60 irradiation (or external irradiation of equivalent quality) in combination with intracavitary radium irradiation.

Oral inhalation system for exposing rabbits to cigarette smoke.

An oral inhalation system for exposing rabbits to cigarette smoke has been developed. The essential components of the system comprise a smoke delivery device that adapts to the oral cavity, a molded face mask, and a sealable body enclosure. The functional capability of the system has been validated by detecting the presence of nicotine in plasma and by measuring the deposition of dotricontane labeled with radioactive carbon (14C) in lungs after exposure to cigarette smoke.