Biological Properties of Extracts Obtained from In Vitro Culture of Plectranthus scutellarioides in a Cell Model.

Plectranthus scutellarioides (L.) R.Br. is a medicinal plant that has long been used in traditional medicine to treat conditions such as abscesses, ulcers, and ear and eye infections. It is known to have a wide range of biological properties, such as antibacterial, antioxidant, antifungal, anti-inflammatory, anti-diabetic and anti-cancer effects. In this study, we established in vitro cultures from both the aerial parts and roots of Plectranthus scutellarioides. Subsequently, we compared the basic phytochemical profile of the obtained extracts and conducted a biological analysis to assess their potential for inducing apoptosis in breast (MCF-7) and lung (A549) cancer cells. Phytochemical analysis by HPLC-MS revealed the presence of compounds belonging to phenolic acids (ferulic, syringic, vanillic, rosmarinic, chlorogenic, caffeic, coumaric, dihydroxybenzoic acids), flavonoids (eriodyctiol and cirsimaritin), and terpenes such as 6,11,12,14,16-Pentahydroxy-3,17diacetyl-8,11,13-abietatrien-7-one, 6,11,12,14,16-Pentahydroxy-3,17-diacetyl5,8,11,13-abietatetraen-7-one, and 3,6,12-Trihydroxy-2-acetyl-8,12-abietadien7,11,14-trione. The results show that both extracts have a cytotoxic and genotoxic effect against MCF-7 and A549 cancer cells, with a different degree of sensitivity. It was also shown that both extracts can induce apoptosis by altering the expression of apoptotic genes (Bax, Bcl-2, TP53, Fas, and TNFSF10), reducing mitochondrial membrane potential, increasing ROS levels, and increasing DNA damage. In addition, it has been shown that the tested extracts can alter blood coagulation parameters. Our results indicate that extracts from in vitro cultures of Plectranthus scutellarioides aerial parts and roots have promising therapeutic application, but further research is needed to better understand the mechanisms of their action in the in vitro model.

Taxodione and Extracts from Salvia austriaca Roots as Human Cholinesterase Inhibitors.

Taxodione, an abietane diterpenoid, was isolated from Salvia austriaca transformed roots grown in in vitro conditions. The compound is known to have antibacterial, cytotoxic and anti-tumour properties. This study evaluates the ability of pure taxodione and extracts obtained from the S. austriaca hairy roots and roots from field-grown plants to inhibit human acetylcholinesterase and butyrylcholinesterase. Both extracts were found to have similar actions against acetylcholinesterase. The IC50 for extracts from transformed and untransformed roots were 142.5 and 139.5 µg ml(-1), respectively. The highest activity towards human acetylcholinesterase was demonstrated by taxodione (IC50 = 54.84 µg ml(-1)). With respect to BChE inhibition, the root extracts demonstrated stronger activity (IC50 = 23.6 µg ml(-1): field-grown plants and 41.6 µg ml(-1): transformed roots) than taxodione (IC50 = 195.9 µg ml(-1)). Taxodione showed significant cytotoxicity against A549 cell line (IC50 = 9.1 µg ml(-1)), whereas the activities for the extracts from S. austriaca roots of field-grown plants (IC50 = 75.7 µg ml(-1)) and hairy roots (IC50 = 86.2 µg ml(-1)) were lower. Computer modelling suggests that taxodione should not demonstrate cardiotoxic or genotoxic activity. It also indicates that taxodione should demonstrate very rapid transport from the body with very good blood-brain barrier penetration, but with no cumulative effect on the human body. The obtained results indicate that taxodione is a safe compound and may be used for further investigations in pharmacological activities.

Aronia melanocarpa Elliot reduces the activity of angiotensin i-converting enzyme-in vitro and ex vivo studies.

PURPOSE: The aim of the study was to analyze the effects of two-month supplementation with chokeberry preparation on the activity of angiotensin I-converting enzyme (ACE) in patients with metabolic syndrome (MS). During the in vitro stage of the study, we determined the concentration of chokeberry extract, which inhibited the activity of ACE by 50% (IC50). METHODS: The participants (n = 70) were divided into three groups: I-patients with MS who received chokeberry extract supplements, II-healthy controls, and III-patients with MS treated with ACE inhibitors. RESULTS: After one and two months of the experiment, a decrease in ACE activity corresponded to 25% and 30%, respectively. We documented significant positive correlations between the ACE activity and the systolic (r = 0.459, P = 0.048) and diastolic blood pressure, (r = 0.603, P = 0.005) and CRP. The IC50 of chokeberry extract and captopril amounted to 155.4 ± 12.1 µg/mL and 0.52 ± 0.18 µg/mL, respectively. CONCLUSIONS: Our in vitro study revealed that chokeberry extract is a relatively weak ACE inhibitor. However, the results of clinical observations suggest that the favorable hypotensive action of chokeberry polyphenols may be an outcome of both ACE inhibition and other pleotropic effects, for example, antioxidative effect.

Extract of Aronia melanocarpa-modified hemostasis: in vitro studies.

PURPOSE: Aronia melanocarpa has an extremely high content of procyanidins and anthocyanins. The multidirectional benefits of consumption of these berries are widely reported. Although numerous studies confirmed the influence of polyphenols on various stages of hemostasis, the exact mechanism of this phenomenon is not understood. The aim of our study was to evaluate the in vitro effect of A. melanocarpa extract on various parameters of hemostasis. METHODS: Adenosine 5'-diphosphate (ADP)-induced aggregation was measured with turbidimetric method. Spontaneous and ADP-activated platelet adhesion were investigated using a colorimetric method. The global assay of coagulation and fibrinolysis was performed with the use of optical clotting and lysis (CL) test. Thrombin (0.5 IU/mL) and tissue plasminogen activator (60 ng/mL) were used to obtain a CL curve. The activity of thrombin and plasmin was determined by means of chromogenic substrate (S-2238, S-2251) RESULTS: The aronia extract contributed to the reduction in spontaneous and ADP-activated platelet adhesion. A significant increase in overall potential of CL as well as significant changes in key parameters of these processes (T t-thrombin time, F vo-initial plasma clotting velocity, and L max-maximum lysis) was reported. Chokeberry extract significantly inhibited the amidolytic activity of thrombin and plasmin. CONCLUSION: Our in vitro findings indicate a complex mechanism of influence of chokeberry polyphenols on platelet activity and the overall potential of CL. We confirmed that chokeberry inhibits the amidolytic activity of thrombin. It was demonstrated for the first time that chokeberry polyphenols inhibit the amidolytic activity of another serine protease, i.e., plasmin, which is the main fibrinolytic enzyme. Furthermore, our research points out a significant contribution of other plasma components and fibrinogen in the modulation of hemostasis by polyphenols.

Short-term supplementation with Aronia melanocarpa extract improves platelet aggregation, clotting, and fibrinolysis in patients with metabolic syndrome.

PURPOSE: A diet rich in berries is believed to play a distinct role in the prevention of metabolic diseases associated with obesity. So far, there have been no published clinical observations evaluating the influence of Aronia melanocarpa on hemostasis. The aim of our study was to investigate the effects of A. melanocarpa extract (AM) supplementation on platelet aggregation, clot formation, and lysis in patients with metabolic syndrome (MS). METHODS: Middle-aged non-medicated subjects with MS (n = 38) and 14 healthy volunteers were included in this study. Patients with MS were treated with 100 mg of AM three times daily for 2 months. RESULTS: We observed a significant reduction in the concentration of TC, LDL-C, and TG after AM supplementation. Beneficial changes in coagulation parameters were also observed. After 1 month of AM administration, we noticed significant inhibition of platelet aggregation. However, this effect became less pronounced after 2 months of supplementation. In the case of coagulation induced by endogenic thrombin, a significant decrease in the overall potential for coagulation was induced after 1 or 2 months of supplementation. Moreover, after 1 month of AM extract supplementation, we observed a beneficial reduction in the overall potential for clot formation and fibrinolysis. CONCLUSIONS: We observed the normalization of hemostasis parameters in MS patients after both 1 and 2 months of AM administration. After 1 month of AM supplementation, we found favorable changes in regards to the overall potential for plasma clotting, clot formation, and lysis, as well as in the lipid profiles of subjects.

Melatonin treatment improves blood pressure, lipid profile, and parameters of oxidative stress in patients with metabolic syndrome.

Experimental studies have proven that melatonin has many beneficial pleiotropic actions. The aim of this study was to assess melatonin efficacy in patients with metabolic syndrome (MS). The study included 33 healthy volunteers (who were not treated with melatonin) and 30 patients with MS, who did not respond to 3-month lifestyle modification. Patients with MS were treated with melatonin (5 mg/day, 2 hr before bedtime) for 2 months. The following parameters were studied: systolic and diastolic blood pressure (SBP, DBP), levels of glucose, serum lipids, C-reactive protein, fibrinogen, activities of antioxidative enzymes: catalase (CAT), glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), thiobarbituric acid reactive substrates (TBARS). After 2-month therapy in comparison with baseline, the following significant changes were measured: systolic blood pressure (132.8±9.8 versus 120.5±11.0 mmHg, P<0.001), DBP (81.7±8.8 versus 75±7.4 mmHg, P<0.01), low-density lipoprotein cholesterol (LDL-C) (149.7±26.4 versus 139.9±30.2 mg/dL, P<0.05), TBARS (0.5±0.2 versus 0.4±0.1 µm/gHb, P<0.01), and CAT (245.9±46.9 versus 276.8±39.4 U/gHb). Melatonin administered for 2 months significantly improved antioxidative defense (increase in CAT activity, decrease in TBARS level) and lipid profile (decrease in LDL-C), and lowered blood pressure. We conclude that melatonin therapy may be of benefit for patients with MS, particularly with arterial hypertension. Further studies with higher doses of melatonin or prolonged supplementation are awaited.

Aronia melanocarpa extract reduces blood pressure, serum endothelin, lipid, and oxidative stress marker levels in patients with metabolic syndrome.

BACKGROUND: Experimental studies have shown that anthocyanins may exert pleiotropic effects. The aim of the study was to determine the influence of Aronia melanocarpa extract on blood pressure and serum concentration of endothelin-1 (ET-1), lipids, glucose, uric acid, C-reactive protein (CRP), fibrinogen, the antioxidant enzymes catalase (CAT), superoxide dysmutase (SOD), and glutathione peroxidase (GSH-Px), and lipid peroxidation (thiobarbituric acid-reacting substrates, TBARS) in erythrocytes of patients with metabolic syndrome (MS). MATERIAL/METHODS: The study comprised 22 healthy volunteers and 25 patients with MS. Patients with MS were treated with aronia extract (3 x 100 mg/day) for two months. The above parameters were measured. RESULTS: After two months of therapy, statistically significant decreases were observed in SBP (143.40+/-7.87 vs. 131.83+/-12.24 mmHg, p<0.001), DBP (87.20+/-9.9 vs. 82.13+/-10.33 mmHg, p<0.05), ET-1 (2.44+/-0.51 vs. 1.74+/-0.42 pg/ml, p<0.001), TC (242.80+/-34.48 vs. 227.96+/-33.07 mg/dl, p<0.001), LDL-C (158.71+/-35.78 vs. 146.21+/-34.63 mg/dl, p<0.01), TG (215.92+/-63.61 vs. 187.58+/-90 mg/dl, p<0.05), TBARS (0.0712+/-0.0191 vs. 0.0362+/-0.0135 micromol/g-Hb, p<0.001), and CAT (261.30+/-59.78 vs. 213.34+/-47.36 U/mg-Hb) and significant increases in SOD (2380.63+/-419.91 vs. 3066.53+/-542.24 U/g-Hb, p<0.001), GSH-Px (12.60+/-5.97 vs. 19.18+/-9.09 U/g-Hb, p<0.01), and fibrinogen levels (249.20+/-27.17 vs. 276.67+/-57.41 mg/dl, p<0.05) compared with the baseline values. CONCLUSIONS: Aronia extract may be of benefit to patients with MS. This seems to result from the influence of anthocyanins and possibly other flavonoids on blood pressure, serum level of ET-1, lipids, and oxidative status (GSH-Px, SOD, TBARS).

A multiparameter test of clot formation and fibrinolysis for in-vitro drug screening.

A large spectrum of methods has been used in both routine and scientific studies of the hemostatic system. The particular interest of the investigators has been focused on methods simultaneously evaluating clotting and fibrinolysis processes. The aim of the present study was to develop an optical method for overall evaluation of clot formation and lysis (CL-test) that could be used in drug screening. The CL-test was performed in citrate plasma diluted with Tris-buffered saline. Thrombin was applied for plasma clotting (0.5 IU/ml), while tissue plasminogen activator (60 ng/ml) was used for fibrinolysis activation. Clot formation and lysis were monitored in thermostatic conditions (37 degrees C) as a continuous record of transmittance change. By means of own computer program, kinetic parameters of the processes studied and plasma overall clot formation and fibrinolysis potential, expressed as the area under the clotgeneration and fibrinolysis curves, were calculated. The CL-test was developed and checked by evaluation of the effect, on clot formation and lysis, of various concentrations of acetylsalicylic acid (a drug that affects hemostasis), aprotinin (fibrinolysis activator) and venoruton (fibrinolysis inhibitor). The obtained results confirmed that the test we propose for monitoring clot formation, stabilization and lysis is sensitive and enables precise estimation of the processes studied. In our opinion, it can be a useful tool in drug screening investigations.

A new nitrate derivative of piperazine: its influence on platelet activity.

Nitric oxide (NO) is a potent vasodilator and inhibitor of platelet activation. Its donors, organic nitrates, are still a main group of drugs administered in ischaemic heart disease. The aim of this study was to investigate the effect of a new NO-donor analogue, 1-(3-piperidinepropionyl)-4-(2-nitrooxy-3-piperidinepropyl) piperazine trihydrochloride (NO-P), on platelet activity. Its influence on the main mechanisms of human platelet activation (adhesion, shape change, secretion and aggregation) was evaluated with the use of a pharmacological model produced on the basis of known platelet activation measuring methods and our computer program. Our experiments revealed that the new NO derivative of piperazine favourably influences platelet activity, and decreases adhesion (spontaneous and induced by ADP) and aggregation. NO-P shows the same direction of action as nitroglycerin (used as a model compound), and is even stronger in the case of ADP-induced and collagen-induced aggregation. These findings broaden the possibility of using NO-P in cardiovascular diseases. Furthermore, our computer program, used to evaluate kinetic parameters of platelet aggregation, shape change, and the adhesion measuring method, provides a simple and accessible experimental model. This model can be useful in in-vitro screening studies, estimating the influence of new compounds (potential drugs) on platelet activity.