Reactivation of sarin-inhibited pig brain acetylcholinesterase using oxime antidotes

Introduction

Organophosphorus nerve agents inhibit the enzyme, acetylcholinesterase (AChE; EC 3.1.1.7). AChE reactivators (also known as oximes) are generally used for the reactivation of an inhibited enzyme.

Methods

Two new AChE reactivators—K033 and K027—were tested for theirin vitro reactivation of sarin-inhibited pig-brain AChE. Their reactivation potencies were compared with the commercially available AChE reactivators, pralidoxime, obidoxime, and HI-6.

Results

Of the oximes tested, the newly developed oxime K027 achieved the highest reactivation potency (100%; concentration of the oxime ?10^?2 M). However, oxime HI-6 (33%) and obidoxime (23%) seem to be the best AChE reactivators for human relevant doses (10^?4 M and lower).

Conclusion

For human relevant doses, newly developed oximes (K027 and K033) do not surpass the reactivation potency of the most promising oxime, HI-6.     

W/O/W multiple emulsions containing nitroimidazole derivates for vaginal delivery

The aim of our study was to formulate a stable multiple emulsions containing two nitroimidazole derivates, metronidazole (MT) and ornidazole (OR), for vaginal therapy. MT and OR were located internal and external phases of multiple emulsion, respectively, and the in vitro release studies were realized in phosphate (pH 7) and lactate buffer (pH 4.5) solutions to investigate better the effect of pH and location of active substance on the release. The imaging studies were realized in rabbits following labeling MT and OR with Technethium-99m (^99mTc) to evaluate the in vivo absorption characteristics. The percentage of MT and OR released from the multiple emulsions in alkaline media were 3.2- and 2.8-fold greater than that observed in acidic media, respectively, when they were introduced in the internal phase of the multiple emulsions. The absorption rate of MT from vaginal epithelium was faster than OR. We observed that especially in alkaline medium a high release was found that was convenient for the vaginal infections seen in the alkaline pH. We concluded that W/O/W multiple emulsions were locally effective in vagina and they could be introduced as a new drug carrier system for vaginal delivery.     

Excimer laser-assisted anterior lamellar keratoplasty for keratoconus, corneal problems after laser in situ keratomileusis, and corneal stromal opacities

PURPOSE: To evaluate excimer laser-assisted anterior lamellar keratoplasty to augment thin corneas as in keratoconus (<350 microm) and corneal ectasia after laser in situ keratomileusis (LASIK) and to treat anterior stromal opacities. SETTING: Ophthalmology Department, School of Medicine, Gazi University, Ankara, Turkey. METHODS: Thirteen eyes (5 keratoconus, 3 macular dystrophies, 1 post-LASIK ectasia, 1 post-LASIK interstitial keratitis, 3 post-herpetic keratitis sequelae) of 13 patients were included in this prospective study. The treatment group was divided into corneal ectasia and stromal opacity groups. A donor stromal button approximately 350 microm thick received a 100 microm excimer laser ablation on the endothelium. The remaining cornea (epithelium, Bowman's membrane, and stroma) was punched with a 7.5 or 7.7 mm trephine. After transepithelial ablation of the host cornea to 200 mum thickness, the corneal button was sutured with interrupted 10-0 monofilament nylon. Sutures were removed between 3 months and 6 months postoperatively. Preoperative and postoperative simulated keratometric cylinders and corneal thickness values were compared using the Wilcoxon signed rank test. The postoperative spherical equivalent refraction and best spectacle-corrected visual acuity (BSCVA) between the groups were compared using the Mann-Whitney U test. RESULTS: The mean follow-up was 27.6 months +/- 8.3 (SD). All patients gained 2 lines or more of BSCVA, and no patient lost a line. The mean corneal thickness was 381.2 +/- 88.2 microm preoperatively, which significantly increased to 534.9 +/- 96.6 microm postoperatively (P < .05). The mean preoperative simulated keratometric cylinder was 7.44 +/- 7.18 diopters (D); postoperatively, it decreased to 2.61 +/- 1.73 D (P < .05). There was no significant difference in postoperative spherical equivalent refraction or BSCVA between the groups (P > .05). CONCLUSIONS: This technique presents a different modality for the treatment of keratoconus, post-LASIK corneal problems, and other corneal stromal opacities with anterior lamellar keratoplasty. Additional studies with more patients and longer follow-up will help determine the role of this technique as a substitute for penetrating keratoplasty in these patients.     

Effects of diuretic-induced hypovolemia/isosmotic dehydration on cardiorespiratory responses to hyperthermia and its physical treatment in rabbits

Under conditions of heat stress and hyperosmotic dehydration, both animals and humans reduce thermoregulatory evaporation and regulate deep body temperature at elevated levels. Regarding the mechanisms, the main role in producing these thermoregulatory changes during dehydration is attributed to the increased osmolality of body fluids, although the role of the decreased plasma volume without changes in plasma osmolality (hypovolemia/isosmotic dehydration) has not been so far investigated. There are also controversial experimental results regarding the effects of dehydration on heat stress-induced cutaneous vasodilation. Therefore, this paper studied the effects of hypovolemia/isosmotic dehydration on cardiorespiratory responses to hyperthermia and its physical treatment in 17 anaesthetized adult rabbits. The animals were divided into two groups: normovolemic group (NV; n?=?10) and hypovolemic group (HV; n?=?7). In the HV group, hypovolemia/isosmotic dehydration (decrease in plasma volume by 16.1?±?1.2%) was induced by furosemide (5?mg?kg^?1 i.v.) without change in measured plasma Na⁺ concentration. Hyperthermia (the rise in body temperature (BT) to 42°C by a gradual body surface heating) caused significant increase in minute ventilation (V_E) in both groups. However, V_E values were significantly higher in the HV rabbits compared to the NV animals despite the lower breathing frequency (p?<?0.05). The panting was absent in the HV rabbits at the BT of 42°C, unlike the NV animals. From cardiovascular variables, the vasoconstrictor response in visceral (mesenteric) region during hyperthermia in hypovolemic/isosmotic animals was attenuated (p?<?0.05), whereas the heat stress-induced cutaneous vasodilation was not influenced by hypovolemia. Recovery of the BT by body surface cooling was accompanied by further increase in V_E in the NV group, whereas V_E decreased (p?<?0.05) in the HV animals. Cooling led to recovery of the cardiovascular parameters. There were found no significant cardiorespiratory differences between the groups (NV:HV) during cooling. The lower frequency of breathing and attenuation of the mesenteric vasoconstriction during exogenous hyperthermia are present not only during hyperosmotic dehydration induced by water deprivation, but they also occur under conditions of furosemide-induced isosmotic dehydration/hypovolemia in rabbits. The heat stress-induced cutaneous vasodilation regarding its biological importance was not influenced by hypovolemia/isosmotic dehydration. Therefore, it is suggested that hypovolemia alone is sufficient to produce described respiratory, thermoregulatory and cardiovascular changes in dehydrated rabbits during exogenous hyperthermia, whereas hyperosmolality is not a requisite.     

Possibilities of improving the parameters of hyperthermia in regional isolated limb perfusion using epidural bupivacaine and accurate temperature measurement of the three layers of limb tissue

The present study presents the author's modification of the method, which aims to create proper parameters of the treatment. The selected group consisted of 15 women and eight men, with a mean age of 57.2 years (range from 26 to 72 years). The patients were divided into two groups, depending on whether they were given epidural bupivacaine (group I - 13 patients treated between the years 2001 and 2004) or not [group II (control) - 10 patients treated earlier, between the years 1997 and 2000]. We observed a significant change in the temperature of thigh muscles (P=0.009) and shank muscles (P=0.006). In the control group II, there was a statistically significant difference (P=0.048) in the temperatures between the muscles and subcutaneous tissue on the one hand and the shank skin on the other. That difference was mean 0.67 degrees Celsius (from 0.4 to 0.9) during the perfusion after applying the cytostatic. The temperature of the skin was lower than the temperature of the deeper tissues of the shank and did not exceed 39.9 degrees Celsius. Such a difference in the temperatures was not observed in case of the group I patients who were given bupivacaine into the extrameningeal space before applying the cytostatic. The difference in the temperatures was on average 0.26 degrees Celsius and was not statistically significant (P=0.99), whereas the shank skin temperature was 40.0-40.6 degrees Celsius. The attained results imply that despite the noticeable improvement in the heating of the limb muscles after application of bupivacaine, the improvement in the heating of the skin and subcutaneous tissue is still not satisfactory, although the growing tendency implies such a possibility.     

Metabolic pathways of ochratoxin A

Ochratoxin A (OTA) as a carcinogenic of group 2B to humans is produced by various fungi strains as Aspergillus and Penicillium. It is one of the most common contaminant in foodstuff. OTA is nephrotoxic, hepatotoxic, teratogenic, and immunotoxic and is assumed to cause Balkan Endemic Nephropathy (BEN), a chronic kidney disease in humans when it is digested in combination with mycotoxin citrinin. The metabolism affects greatly the fates and the toxicity of a mycotoxins in humans, animals, and plants. The understanding of the metabolism of mycotoxins by the organism as fungi, yeast, bacteria and enzymes would be very helpful for the control of the contamination by the mycotoxins in foods and feeds, and understanding of the biotransformation of the mycotoxin in the body of humans, animals, plants, microorganisms would be beneficial to the risk assessment of food safety. In animals and humans, OTA can be metabolized in the kidney, liver and intestines. Hydrolysis, hydroxylation, lactone-opening and conjugation are the major metabolic pathways. OTalpha (OTα) formed by the cleavage of the peptidic bond in OTA is a major metabolite not only in animals and humans, but also in microorganisms and enzyme systems. It is considered as a nontoxic product. However, the lactone-opened product (OP-OTA), found in rodents, is higher toxic than its parent, OTA.. (4R)-4-OH-OTA is the major hydroxy product in rodents, whereas the 4S isomer is the major in pigs. 10-OH-OTA is currently found only in rabbits. Furthermore, OTA can lose the chlorine on C-5 to produce ochratoxin B (OTB), and OTB is further to 4-OH-OTB and ochratoxin β (OTβ). Ochratoxin quinine/hydroquinone (OTQ/OTHQ) is the metabolite of OTA in animals. In addition, the conjugates of OTA such as hexose and pentose conjugates can be found in animals. Such more polar metabolites make OTA to eliminate faster. Currently, a debate exits on the formation of OTA-DNA adducts. Plants can metabolize OTA as well. OH-OTA methyl ester and OH-OTA-β-glucoside are formed in many plants besides OTα and OH-OTA. OTA can be biotransformed into OTα by some yeast strains. Fungi can produce some of the same metabolites as animals. OTα, OTβ, 4-R-OH-OTA, 4-R-OH-OTB, and 10-OH-OTA are the metabolites in fungi. Several commercial enzymes are able to biodegrade OTA into the nontoxic OTα efficiently. This review on the metabolism of OTA helps to well understand the fate of OTA in different organisms, as well as provides very crucial information for toxicology and food safety assessments on human health.     

Asoxime (HI-6) impact on dogs after one and tenfold therapeutic doses: assessment of adverse effects, distribution, and oxidative stress

Asoxime (HI-6) is a well known oxime reactivator used for counteracting intoxication by nerve agents. It is able to reactivate acetylcholinesterase (AChE) inhibited even by sarin or soman. The present experiment was aimed to determine markers of oxidative stress represented by thiobarbituric acid reactive substances and antioxidants represented by ferric reducing antioxidant power, reduced and oxidized glutathione in a Beagle dog model. Two groups of dogs were intramuscularly exposed to single (11.4 mg/kg.b.wt.) or tenfold (114 mg/kg.b.wt.) human therapeutically doses of HI-6. HI-6 affinity for AChE in vitro was evaluated in a separate experiment. Complete serum biochemistry and pharmacokinetics were also performed with significant alteration in blood urea nitrogen, creatine phosphokinase, glucose and triglycerides. Blood samples were collected before HI-6 application and after 30, 60, and 120 min. The overall HI-6 impact on organism is discussed.     

Partition of bispyridinium oximes (trimedoxime and K074) administered in therapeutic doses into different parts of the rat brain

The penetration of acetylcholinesterase reactivators (oximes) into the central nervous system is typically restricted by the blood-brain barrier. Although oximes are highly hydrophilic compounds, some contradictory results confirming permeation into the brain exist. The aim of this study is to verify the penetration of oximes through the blood-brain barrier and to detect their levels achieved in different brain regions 60 min after the administration. It was confirmed that oximes are able to penetrate into the brain after injection of therapeutic doses corresponding with 5% of LD(50). The level in whole brain was 0.58% for trimedoxime and 0.85% for the experimental drug oxime K074 as the percentage of their plasma concentration. The highest concentration was found in frontal cortex (trimedoxime 2.27%; oxime K074 0.95%) and lowest in basal ganglia (trimedoxime 0.86%; oxime K074 0.42%). Entry of oximes into the brain is minimal, but some low reactivation effect should be expected. The reactivation potency of oximes might be higher or lower, depending on the real oxime concentration in a given area.     

Measurements of caffeine and plasma metabolite/caffeine ratios as a test for hepatic drug-oxidizing capacity in goats

The aim of this investigation was to determine the pharmacokinetics and demethylation of caffeine (CF) and the metabolite/CF ratios that correlated best with CF clearance, which were used to evaluate hepatic drug-oxidizing capacity of CF after a single intravenous dose (5?mg/kg) in hair goats (n?=?9). Pharmacokinetic parameters of CF and its metabolites, theobromine (TB), paraxanthine (PX) and theophylline (TP), were calculated. The plasma metabolic ratios TB/CF, PX/CF, TP/CF and TB+PX+TP/CF were determined at 6, 8 and 10?h after CF administration to evaluate their hepatic drug-oxidizing capacity. The plasma concentration-time data of CF were fit to a two-compartment model in all animals. The clearance of CF was 0.08?±?0.02?L/h/kg, and the volume of distribution was 0.91?±?0.16?L/kg. The demethylation fractions of CF to TB, PX and TP were 0.24, 0.37 and 0.39, respectively. Correlations between the metabolic ratios and CF clearance were quite high, except for the PX/CF ratio, particularly at 6?h (r?=?0.650-0.750, P?相似文献