PRELIMINARY RESEARCH REGARDING THE THERAPEUTIC USES OF URTICA DIOICA L. NOTE I. THE POLYPHENOLS EVALUATION
The use of stinging nettle leaves for the treatment of diabetes mellitus without a scientific evaluation, persuaded us to study them, in order to obtain pharmacological active extracts. According to scientific data, polyphenols are known as HMG-CoA reductase inhibitors and have potent anti-glycation activity. These compounds are the main active principles in the Urticae folium. The aim of this study was to evaluate the polyphenols’content (polyphenolcarboxilic acids = PPCA, and flavonoids) of stinging nettle leaves, harvested from culture and wild flora, originated from Racari, Dambovita district, Romania. Spectrophotometric and chromatographic methods were used. A new HPLC method for the identification and quantification of caffeic and chlorogenic acids from Urticae folium was also developed. The highest contents of polyphenolcarboxilic acids (expressed as % of chlorogenic acid) and flavonoids (expressed as % of rutin) were found in the leaves of wild nettles harvested in the month of March (4.2295% chlorogenic acid and 0.6237% rutin). The polyphenols’ content is slightly lower than the ones mentioned in the scientific literature. The content of PPCA decreases from March till September, both in wild and cultivated samples. The variation of the flavonoids’content differs with the origin of raw material (culture or spontaneus flora). The new HPLC method for polyphenols’analysis is reproductible. Rezumat Utilizarea tradiţională a produsului Urticae folium in diabet, fără o justificare stiintifica a acţiunii precum si menţionarea flavonelor si acizilor fenolcarboxilici (compusii majoritari) ca inhibitori ai HMG-CoA reductazei si ai proceselor de glicozilare si lipoxidare avansată, ne‐au determinat să abordăm cercetarea acestora, in vederea valorificării terapeutice sub formă de extracte selective. In acest scop am derulat o cercetare spectrofotometrică si cromatografică (HPLC) pentru a pune in evidenţă polifenolii prezenţi in frunzele de urzică recoltate din flora spontană si de cultură, din localitatea Răcari, judeţul Dâmboviţa. De asemenea, ne-am propus să separăm acizii cafeic si clorogenic si să-i determinăm cantitativ printr-o nouă metodă HPLC/UV.
Anthocyan Glycosides from Urtica dioica
The aerial part of dioecious nettle (Urtica dioica L., Urticaceae) collected in October 2001 in Hob region (western Georgia) was investigated. The air-dried ground raw material was extracted with CHCl3 and then CH3OH containing HCl (1%) in the dark at room temperature. The CH3OH extract was concentrated in vacuum at 35-40°C and chromatographed over a column of cellulose powder with elution by aqueous HCl (0.5%). Fractions were collected as the anthocyan bands eluted. Eluates of pure compounds were combined, condensed in vacuum to a small volume, and treated with diethylether (10:1). The precipitate was separated, washed with ether, and dried in vacuum. Three compounds were obtained. Compound 1 was a crystalline dark-red powder, mp 260°C (dec.), PC Rf 0.06 (0.5% HCl). UV spectrum (MeOH containing 0.1% HCl, λmax, nm): 280, 530. Acid hydrolysis of 1 and 2 produced the aglycon with mp >300°C (dec.). The aglycon was identified as pelargonidin. This was confirmed by decomposing it with Ba(OH)2 solution (15%) to phloroglucinol and p-hydroxybenzoic acid and by chromatography with authentic pelargonidin, prepared by reduction of kaempferol. UV spectrum (MeOH containing 0.1% HCl, λmax, nm): 270, 520. UV spectra of the glycosides measured with methanolic AlCl3 (5%) showed that ring B had no dihydroxy group [1]. The carbohydrate part of 1 and 2 was D-xylose. The attachment site of the carbohydrates to the aglycon was determined by oxidation of the anthocyan glycosides 1 and 2 with H2O2 [2]. This revealed that the former contains D-xylose; the latter, biose, acid hydrolysis of which produced D-xylose. Stepwise acid hydrolysis of 2 formed 1 and D-xylose. These results indicate that 1 is pelargonidin monoxyloside; 2, pelargonidin xylobioside. These anthocyans are described by us for the first time. Compound 3, PC Rf 0.70 (0.5% HCl). UV spectrum (MeOH containing 0.1% HCl, λmax, nm): 280, 530. Acid hydrolysis produced the same aglycon, pelargonidin, and the sugars D-glucose and L-rhamnose. The structures of 1-3 are still under investigation. The total anthocyan content of dioecious nettle that was determined by a spectrophotometric method [3] calculated for cyanidin-3,5-diglucoside was 0.3%.
Clinical chemical values and variations in Rasa Aragonesa ewes and lambs
Reference values of biochemical parameters in serum of Rasa Aragonesa ewes and their physiological variations were determined. Total proteins were 53.0–80.0 g/l, glucose 2.17–5.13 mmol/l, cholesterol 0.83–2.31 mmol/l, total bilirubin 1.85–6.49 μmol/l, urea 3.15–10.39 mmol/l, GGT 36–93 U/l (37°C), ALAT 11–33 U/l (37°C) and ASAT 71–209 U/l (37°C). Ewes were divided into three groups: A, 1- and 2-year-old ewes; B, ewes 3 and 4 years old; C, older ewes. Age had significant influence (P<0.05) on ASAT only during pregnancy, with higher levels for younger ewes. Blood parameters in lambs were determined at 5–10 d of life (nursing), 25 to 30 d of life receiving their mother's milk and 80–90 g of feed; and 45 to 50 d of life, weaned, receiving only solid feed. Serum urea and ALAT tended to increase with age, whereas total protein, globulins, cholesterol and GGT decreased with age. These values are of practical importance in the diagnosis of diseases and metabolic disorders in Rasa Aragonesa ewes and lambs under field conditions.