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[Biological standardization of Ginkgo extracts].

The determination of the inhibition of PAF (platelet-activating factor)-induced platelet aggregation has been proposed as a biological standardization method for commercially available Ginkgo biloba extracts by measuring the characteristic pharmacological effect of ginkgolides in vitro. The determination is specific for ginkgolides A, B, C, and J and is not influenced by other constituents present in Ginkgo biloba extracts. IC50 values of ginkgolide B can be used to standardize various Ginkgo extracts produced by special extraction methods with respect to equi-effective ginkgolide B contents. In order to compare values obtained by a chemical-analytical procedure with those obtained by the biological assay, the equi-effective total ginkgolide content of each Ginkgo extract had to be calculated. Accordingly, the concentrations of the individual ginkgolides in the various Ginkgo extracts were determined chromatographically by assaying ginkgolides as trime-thylsilyl derivatives. Their individual contributions towards the measured in vitro effects were derived from their respective IC50 values. The calculated equi-effective total ginkgolide contents of the Ginkgo extracts were in good agreement with those obtained by gas chromatography. The results demonstrate that, in addition to a chemical standardization, the biological standardization of Ginkgo extract preparations is also feasible.

Lactones

[Preparation and definition of Ginkgo biloba extract].

Ginkgo biloba extract is a well-defined and complex product prepared from green leaves of Ginkgo biloba. The leaves are harvested from trees growing in plantations in South Corea, Japan and France. The mode of culture, harvesting and extraction are perfectly standardized and controlled. Analysis of Ginkgo biloba extract makes it possible to confirm that undesirable substances have been eliminated and to measure the amount of active principles. The extract contains flavonoid substances, such as the Ginkgo-flavone glycosides and terponoids which are characteristic of Ginkgo and have a unique structure (ginkgolides, bilobalide).

Chemical Phenomena

Induction of superoxide dismutase and catalase activity in different rat tissues and protection from UVB irradiation after topical application of Ginkgo biloba extracts.

Ginkgo biloba extract (GBE) prepared from the leaves of Ginkgo biloba with 50% diluted alcohol was found to locally induce superoxide dismutase (SOD) and catalase (CAT) enzyme activity in epidermis after topical application, and also to systemically increase the activity of both enzymes in the liver, heart and kidney of Sprague Dawley rats. Skin pretreated with 50% diluted alcohol-extracted liquid formulation was protected from exacerbation of UVB damage. Changes in the lipid structure of the skin of rats determined by ATR/FT-IR spectroscopy demonstrated penetration of active components from GBE dosage formulations.

Administration, Topical

Superoxide anion scavenging effect and superoxide dismutase activity of Ginkgo biloba extract.

Ginkgo biloba extract is known to be efficient in diseases associated with free radical generation. The purpose of this work was to study, under in vitro conditions, the action of Ginkgo biloba extract (Gbe) against superoxide anion (O2-.), which is directly or indirectly implicated in cell damage. Gbe appears to have both an O2-. scavenging effect and also a superoxide dismutase activity. Its antiradical effect was demonstrated by low temperature electron spin resonance and in a non-enzymatic system (phenazine methosulfate-NADH), and its enzymatic activity was shown by polarographic determination.

Animals

Influence of the specific antagonist of PAF-acether, BN 52021, and of Ginkgo extract on the rejection of murine tail skin allografts and the PAF-acether mortality in mice in particular consideration of the role of TXB2.

An oral application of 2 x 10 mg/kg daily of the PAF antagonist, BN 52021, significantly prolonged the distal allograft survival in the murine tail skin model of cell-mediated allograft rejection. BN 52021 diminished the TXB2 content in the skin at and near to the transplantation site by two thirds whereas PAF-acether significantly enhanced the TXB2 content in murine tail skin. Similar to the tail skin BN 52021 also reduced by more than 70% the TXB2 release from human granulocytes stimulated by calcium ionophore A 23187. The PAF-acether mortality (LD75) in NMRI mice was decreased to 20% by BN 52021 with a lower dose than by BN 52020. Comparing Ginkgo total extract in doses which significantly diminished the 75% mortality by PAF-acether in mice with BN 52021 and BN 52020 in doses that produced the same inhibitory effect, a potentiated effect of Ginkgo extract resulted with respect to the share of BN 52021 and BN 52020 in Ginkgo extract.

Animals

[Neuromediator changes during cerebral aging. The effect of Ginkgo biloba extract].

Ginkgo biloba extract exerts a specific effect on the noradrenergic system and on beta-receptors. No variation was found in alpha 2-receptors and serotonin uptake. These findings provide the first evidence of central effects of a drug acting on cerebral ageing, connected specifically to reactivation of the noradrenergic system in the cerebral cortex.

Aging

[Ginkgo extract EGb 761 (tenobin)/HAES versus naftidrofuryl (Dusodril)/HAES. A randomized study of therapy of sudden deafness].

80 patients with idiopathic sudden hearing loss existing no longer than 10 days were included in a randomised reference-controlled study. The therapeutic value of Ginkgo EGb 761 (Tebonin) + HAES was compared to that of Naftidrofuryl (Dusodril)+HAES. The main mechanisms of action of EGb 761 are a vasoregulating activity (increased blood flow), the platelet activating factor antagonism and a prevention of membrane damage caused by free radicals. Naftidrofuryl has antiserotonergic and therefore vasodilatory properties. The statistical analysis of the audiometric data was performed in measuring the relative hearing gain as described by Eibach 1979. After one week of observation, 40% of the patients in each group showed a complete remission of hearing loss. This was also observed by other authors who had compared other drugs. Therefore, in these cases, it is most likely that spontaneous recovery is the most important factor. After two and three weeks of observation, measuring the relative hearing gain, there was a significant borderline benefit of EGb 761 (p = 0.06) without any side effects. Some patients of the reference group developed side effects such as orthostatic dysregulation or headache or sleep disturbances. Minimising side effects should be one of the most important goals in therapy of sudden hearing loss until the efficiency of infusion therapy is proved.

Adult

[Superoxide dismutase activity of Ginkgo biloba extract].

The Ginkgo biloba extract is obtained from green leaves of the Ginkgo biloba tree. Preparations with this active substance are among others used for the treatment of disturbances of the cerebral function and arteriosclerotic diseases. In in-vitro and in-vivo studies antagonistic effects of radical scavenger and PAF (platelet activating factor) were described. In this study a concentration-depending superoxide dismutase activity of the Ginkgo biloba extract rökan liquid could be made evident.

Diterpenes

Effect of two doses of ginkgo biloba extract (EGb 761) on the dual-coding test in elderly subjects.

The subjects of this double-blind study were 18 elderly men and women (mean age, 69.3 years) with slight age-related memory impairment. In a crossover-study design, each subject received placebo or an extract of Ginkgo biloba (EGb 761) (320 mg or 600 mg) 1 hour before performing a dual-coding test that measures the speed of information processing; the test consists of several coding series of drawings and words presented at decreasing times of 1920, 960, 480, 240, and 120 ms. The dual-coding phenomenon (a break point between coding verbal material and images) was demonstrated in all the tests. After placebo, the break point was observed at 960 ms and dual coding beginning at 1920 ms. After each dose of the ginkgo extract, the break point (at 480 ms) and dual coding (at 960 ms) were significantly shifted toward a shorter presentation time, indicating an improvement in the speed of information processing.

Aged

The nitric oxide-scavenging properties of Ginkgo biloba extract EGb 761.

Ginkgo biloba extract EGb 761 was found to be a scavenger of nitric oxide in in vitro acellular systems, under physiological conditions. EGb 761 competed with oxyhemoglobin for reaction with nitric oxide generated during the interaction of hydroxylamine with Complex I of catalase. An EGb 761 dose-dependent decrease in the amount of nitrite formed in the reaction of oxygen with nitric oxide produced from solution of 5 mM sodium nitroprusside was also observed. These data implicate it as a potential therapeutic agent in conditions of altered production of nitric oxide.

Animals

[Efficacy of an extract of Ginkgo biloba in the treatment of chronic obliterating arteriopathies of the lower limbs in stage III of Fontaine's classification].

UNLABELLED: A controlled trial of Ginkgo biloba extract in injectable form (Tanakan 50 mg, a lyophilizate for parenteral use) was carried out versus a placebo as a preoperative medical treatment of stage III (Fontaine classification) chronic occlusive arterial disease of the lower limbs (with pain in decubitus). The 64 men and women patients in this multicenter study (32 in each group) were over 18 years of age and had a cultural and intellectual level as well as a physical condition allowing them to play an active role in the experiment (self-evaluation of pain). During 8 days they received two daily infusions of 500 cc of normal saline solution containing either 100 mg of Ginkgo biloba extract or a placebo of identical appearance. During this period, anticoagulants were authorized; hemodilution, vasoactive drugs and platelet anti-aggregates were forbidden; pentazocine (Fortal, 50-mg tablets) was allowed at the patient's request. Pain was rated according to a visual scale, with each patient marking a point between two extremes ("maximum imaginable pain" and "total absence of pain") 100 mm apart. A questionnaire based on that of Melzack (McGill Pain Questionnaire) completed this qualitative as well as quantitative self-evaluation of pain. The results of these questionnaires were assessed on the basis of 4 scores, each determined by the patient's choice among 3 evaluation figures. The chi square, Student and Wilcoxon tests were used for statistical analysis. RESULTS: The two randomly-composed groups with Ginkgo biloba extract and a placebo were comparable (Table I). Analysis was based on 55 observations (26 in the extract group and 29 in the placebo group).(ABSTRACT TRUNCATED AT 250 WORDS)

Arterial Occlusive Diseases

[Drug therapy of disorders of cerebral performance. Randomized comparative study of dihydroergotoxine and Ginkgo biloba extract].

In a randomized trial lasting six weeks and involving 80 elderly patients with cerebrovascular disorders, the effectiveness and tolerance of dihydroergotoxine was compared with an extract of Ginkgo biloba. On the basis of psychometric tests and assessment scales, it was shown that treatment with either substance improved the condition of the patients. While, for the most part, intergroup comparison revealed no major statistically significant differences, such changes, affecting various parameters, were found during follow-up--more frequently within the dihydroergotoxine group than within the group treated with Ginkgo biloba extract.

Aged

Attenuation of salicylate-induced tinnitus by Ginkgo biloba extract in rats.

The effects of an extract from Ginkgo biloba, EGb 761, on tinnitus were tested using an animal model of tinnitus. Daily oral administration of EGb 761 in doses from 10 to 100 mg/ kg/day began 2 weeks before behavioral procedures and continued until the end of the experiment. Tinnitus was induced by daily administration of 321 mg/kg sodium salicylate s.c. (corresponding to 275 mg/kg/day of salicylate acid) in fourteen groups of pigmented rats, 6 animals/group. The results from salicylate- and EGb-761-treated animals were compared to control groups receiving either salicylate, saline, or EGb 761 only in doses of 100 mg/kg. Administration of EGb 761 resulted in a statistically significant decrease of the behavioral manifestation of tinnitus for doses of 25, 50 and 100 mg/kg/ day.

Administration, Oral

Lipid peroxide, phospholipids, glutathione levels and superoxide dismutase activity in rat brain after ischaemia: effect of ginkgo biloba extract.

The influence of ginkgo biloba extract on the lipid peroxide product (malondialdehyde, MDA), glutathione (GSH) and phospholipids levels as well as superoxide dismutase (SOD, 1.15.1.1) and lactate dehydrogenase (LDH, 1.1.1.27) activities in rat brain after occlusion of common carotid arteries was investigated. Two experimental models were studied: 60 min ischaemia without reperfusion and 60 min ischaemia followed by 60 min reperfusion. Compared to sham-operated animals, ischaemia followed by reperfusion increased cytosolic LDH activity and mitochondrial lipid peroxide content and decreased the superoxide dismutase activity and mitochondrial total phospholipids level. Preischaemic administration of ginkgo biloba extract (150 mg kg-1, p.o.) could normalize the SOD activity of the rat brain. The extract was also able to reduce the lipid peroxide and phospholipids contents of the mitochondrial rat brain. These effects could be explained on the basis of the antioxidant property of ginkgo biloba extract and suggests its beneficial role in the protection against post-ischaemic injury.

Animals

Ex vivo regulation of adrenal cortical cell steroid and protein synthesis, in response to adrenocorticotropic hormone stimulation, by the Ginkgo biloba extract EGb 761 and isolated ginkgolide B.

We previously demonstrated that repeated treatment of rats with the standardized extract of Ginkgo biloba leaves, EGb 761, and its bioactive component ginkgolide B (GKB), specifically reduces the ligand binding, and protein and messenger RNA expression of the adrenal mitochondrial peripheral benzodiazepine receptor (PBR), a key element in the regulation of cholesterol transport, resulting in decreased circulating corticosterone levels. Adrenocortical cells were isolated from rats treated with EGb 761 or GKB and cultured for 2 and 12 days. The effect of ACTH on normal and metabolically labeled cells was examined. Corticosterone levels were measured by RIA, and protein synthesis was analyzed by two-dimensional gel electrophoresis. Ex vivo treatment with EGb 761 and GKB resulted, respectively, in 50% and 80% reductions of ACTH-stimulated corticosterone production by adrenocortical cells cultured for 2 days compared with that by cells isolated from saline-treated rats. Two-dimensional gel electrophoresis analysis revealed that in cells from both control and drug-treated animals, ACTH induced the synthesis, at the same level, of a 29-kDa and pI 6.4-6.7 protein identified as the adrenal steroidogenic acute regulatory protein (StAR). In addition, treatment with EGb 761 and GKB specifically altered the synthesis of seven proteins, including inhibition of synthesis of a 17-kDa, identified as PBR. After 12 days in culture, ACTH-stimulated adrenocortical cell steroid synthesis was maintained, and it was identical among the cells isolated from animals treated with GKB or saline. Under the same conditions, the expression of PBR was recovered, whereas no effect of ACTH on the 29-kDa and 6.4-6.7 pI protein (StAR) or other protein synthesis could be seen. A comparative analysis of the effects of GKB and EGb 761 on adrenocortical steroidogenesis and protein synthesis identified, in addition to the 17-kDa PBR, target proteins of 32 kDa (pI 6.7) and 40 kDa (pI 5.7-6.0) as potential mediators of the effect of EGb 761 and GKB on ACTH-stimulated glucocorticoid synthesis. In conclusion, these results 1) validate and extend our previous in vivo findings on the effect of EGb 761 and GKB on ACTH-stimulated adrenocortical steroidogenesis, 2) demonstrate the specificity and reversibility of EGb 761 and GKB treatment, 3) question the role of the 29-kDa, 6.4-6.7 pI protein (mature StAR) as the sole mediator of ACTH-stimulated steroid production, and 4) demonstrate the obligatory role of PBR in hormone-regulated steroidogenesis.

Adrenal Cortex

Protection of polyunsaturated fatty acids against iron-dependent lipid peroxidation by a Ginkgo biloba extract (EGb 761).

Iron can induce a peroxidative degradation of the membrane polyunsaturated fatty acids by the well-known Fenton reaction. Chelated iron can also form a complex with oxygen, called perferryl ion, which is able to induce lipoperoxidation without a detectable production of hydroxyl radicals. The antioxidant properties of a titrated and standardized extract of Ginkgo biloba leaves (EGb 761) against iron-dependent peroxidative degradation of the membrane polyunsaturated fatty acids were studied. Rat liver microsomes were exposed to a mixture of NADPH, ADP and FeCl3 hypothesized to produce iron-oxygen complexes. The results of the analysis of the microsomal polyunsaturated fatty acids by gas chromatography show that the susceptibility to peroxidation of the different polyunsaturated fatty acids increases with their unsaturation level, and that EGb 761 protects these membrane polyunsaturated fatty acids regardless of their susceptibility to peroxidation. This protective effect is correlated with the decrease in the thiobarbituric acid-reactive substances concentration, but the calculation of the antioxidant potency of EGb 761 as an IC50 value using results of the thiobarbituric acid reaction leads to an underestimated evaluation when the reaction is carried out in the presence of iron ions.

Animals

Efficiency of Ginkgo biloba extract (EGb 761) in antioxidant protection against myocardial ischemia and reperfusion injury.

The cardio-protective mechanisms of EGb 761, an extract of Ginkgo biloba leaves, on myocardial ischemia-reperfusion injury were investigated using rabbits subjected to 30 minutes of regional cardiac ischemia and 120 min of reperfusion under anesthesia. Compared to the saline perfused group, EGb 761 treatment (10 mg/kg, injected into the coronary artery) significantly inhibited the increase in lipid peroxidation and maintained total and CuZn-SOD levels in both plasma and tissue during and at the end of reperfusion. Both the decrease in tissue type plasminogen activator (t-PA) and the increase in plasminogen activator inhibitor-1 (PAI-1) caused by ischemia-reperfusion were also significantly suppressed by EGb 761 treatment. Furthermore, the ultrastructure of the myocytes of the EGb 761 treated heart was slightly damaged after ischemia-reperfusion, while the control ischemic-reperfused hearts demonstrated severe histological damages such as swelling and vacuolization of the mitochondria. These results suggest that EGb 761 protects hearts by its antioxidant properties and by its ability to adjust fibrinolytic activity.

Animals

The bioavailability of ginkgolides in Ginkgo biloba extracts.

A new Ginkgo biloba leaf extract, BioGinkgo 27/7, was prepared using a method that enriches ginkgolide B. The bioavailability of ginkgolides in these extracts was assessed in rabbits in comparison with a commercially available standardized 24/6 extract. It was found that, after a single dose, a higher concentration of ginkgolides was maintained for a longer period of time with these extracts than was found with commercial extract prepared by existing methods.

Animals