Investigation of fractions present in the stem bark of Annickia kummeriae on their P-glycoprotein inhibitory pump activity.

Using MCF-7R cells and rhodamine 6G as the fluorescent probe, a bioassay-targeted purification process was pursued in order to isolate the active P-gp inhibitory fractions from Annickia kummeriae. Of 24 fractions obtained in the first preparative liquid chromatography (p-LC) run, only fraction 1 exhibited activity. Further p-LC fractionation led to the separation of fraction 1 into fractions 1.1-1.8. Fractions 1.4, 1.5 and 1.6 proved to be active by inducing a significant accumulation of rhodamine 6G by 3.3-, 4.5- and 4.9-fold at 10 microg/mL, and by 5.3-, 6.3- and 6.8-fold at 100 microg/mL, respectively. Fraction 1.6 was separated into several fractions by using an analytical liquid chromatography (a-LC) system. Fractions 1.6.18, 1.6.19 and 1.6.20 were active and they induced an accumulation of rhodamine 6G by 3.0-, 1.8- and 3.5-fold at 1x microg/mL and by 4.8-, 6.7- and 6.8-fold at 10x microg/mL, respectively. Afterwards, 28.3 mg of fraction 1.6 was processed by a-LC, and fractions 1.6.18, 1.6.19 and 1.6.20 were collected separately and dried. The amounts of materials recovered were 6.2, 7.4 and <1 mg, corresponding to 21.9%, 26.1% and <3.5% of fraction 1.6, respectively. From the total amount injected and the relative masses represented by these fractions, it can be calculated that the 1x microg/mL level corresponded to ca. 35, 42 and <5 microg/mL, respectively. Fluorescence microscopy revealed that incubation of the cells with rhodamine 6G alone did not show any fluorescence, whereas cells which were incubated in medium containing rhodamine 6G together with fraction 1.4, 1.6 or reserpine, clearly indicated accumulation of the dye intracellularly. This is an indication that the active compounds effected high intracellular fluorescence by inducing accumulation of the dye in the cells through inhibition of the P-gp pump.

Teratological studies in defatted jojoba meal-supplemented rats.

To look for possible developmental effects in the offspring of jojoba meal-treated Wistar rats, and to distinguish between the effects of reduced food intake and the specific developmental effects of jojoba meal itself, mated female rats were divided into three groups of 20 rats. They received during gestation: (a) normal rodent food (control group); (b) normal rodent food supplemented with 3% defatted jojoba meal (jojoba group); or (c) normal rodent food pair-fed with the jojoba group (pair-fed group). The jojoba meal group showed approximately 30% inhibition of food intake. Ten rats from each group were killed on gestation day 21. Compared to the control group, foetal body weight was reduced in both the jojoba and pair-fed groups, with a greater reduction in the jojoba group. Skeletal ossification was retarded to the same extent in both the jojoba and pair-fed groups. The other 10 rats from each group were left to produce litters. Compared with controls, the body weight of the pups was lower in both the jojoba and pair-fed groups; the reduction was slightly greater in the jojoba group, but this difference disappeared after 1 week. The offspring showed no other abnormalities and reproduced normally. We conclude that, at the dose used, the retardation in foetal skeletal ossification, induced by jojoba meal supplementation during gestation, is due to food intake inhibition. Moreover, the lower birth weight of the young of jojoba-treated dams compared with the pair-fed group is merely due to a lower body weight gain during gestation.

Reproductive performance of rats treated with defatted jojoba meal or simmondsin before or during gestation.

The effects on food intake, growth and reproductive performance parameters of defatted jojoba meal and pure simmondsin, an extract from jojoba meal, were compared in female Wistar rats. Rats fed 0.15% simmondsin or 3% defatted jojoba meal (equivalent to 0.15% simmondsin) for 8 weeks before conception showed a similar reduction in food intake (about 20%) and a similar growth retardation compared with controls. Both treatments induced a reduction in the number of corpora lutea on gestation day 16: this effect could be ascribed to the lower food intake before conception because it was also observed in rats pair-fed to the treated ones. Rats given feed containing 0.15% simmondsin or 3% defatted jojoba meal during days 1-16 of gestation showed a similar reduction in food intake relative to controls. Foetal and placental weights were reduced, relative to controls, to a similar extent in both groups, and the reductions were slightly greater than in the corresponding pair-fed groups. We conclude that the effects on food intake, growth and reproductive performance that were seen after feeding rats defatted jojoba meal were due to the simmondsin content of the meal. The simmondsin induced reduction in food intake and probably also a relative protein shortage.

Devazepide reverses the anorexic effect of simmondsin in the rat.

Simmondsin, a glycoside extracted from jojoba meal (Simmondsia chinensis), causes a reduction in food intake after oral administration. To investigate the mechanism by which simmondsin reduces food intake, fasted and free-feeding rats were given simmondsin-supplemented food and simultaneously injected with devazepide, a specific antagonist of peripheral-type cholecystokinin receptors (CCKA receptors). In free-feeding rats, supplementation of food with 0.5% simmondsin caused a reduction in food intake of +/- 40% in the period of 4 h following food presentation. Intraperitoneal injection of 100 micrograms devazepide/kg body weight prevented this effect. In rats fasted for 20 h, the food intake in the 30 min after presentation of food supplemented with 0.15% or 0.50% simmondsin was reduced in a dose-related manner; this was also inhibited by simultaneous application of devazepide. It is suggested that peripheral CCKA receptors are involved in the effect of simmondsin on food intake. However, a direct effect of simmondsin on CCKA receptors has been excluded, since simmondsin was unable to cause contraction of the guinea-pig gallbladder in vitro.

Standardizing 8-methoxypsoralen plasma profiles by using an emulsion form.

Uniform 8-methoxypsoralen (8-MOP) absorption from the gastrointestinal tract is necessary to avoid day-to-day variations in 8-MOP plasma levels when treating patients with psoriasis by photochemotherapy. Because of its low water solubility, particle size and crystal form of the 8-MOP can significantly influence its bioavailability. The presentation form is also important, as is shown by the present study in which 8-MOP plasma levels were compared in thirty patients after oral administration in three different forms: formulation A consisted of gelatin capsules containing 8-MOP with a mean particle size of 200 mu; formulation B consisted of gelatin capsules containing 8-MOP in microcrystalline form with particle size between 20 mu and 30 mu; formulation C contained the same microcrystalline 8-MOP but in an emulsion base. Significantly higher plasma levels were found with formulations B and C than with formulation A. Furthermore, the individual differences in plasma profiles were markedly less with the emulsion base than with the capsule forms. Therefore, the clinical use of 8-MOP in emulsion form would constitute a major step in 8-MOP dose standardization and could lead to better control of subjective side effects and better therapeutic results.

Dietary influences on 8-MOP plasma levels in PUVA patients with psoriasis.

To study the effect of diet on 8-MOP plasma levels, these levels were measured three times in twenty PUVA patients with psoriasis; while fasting, after a low-fat breakfast, and after a fat-rich breakfast. No significant differences were found between the maximal 8-MOP plasma levels in the fasting condition and the levels after the low-fat breakfast, both peaking 2 h after ingestion. After the fat-rich breakfast, however, the 8-MOP plasma levels were not only significantly lower at 2 h but also peaked later, reaching their maximum at 3-4 h after intake. The drug should therefore be given in a dietetically standardized way. This raises the question whether some cases of therapy resistance can be dietetically influenced.