Bryophyllum pinnatum - metabolite profiling and "in vitro" effects on porcine detrusor contractility

Bryophyllum pinnatum is a succulent perennial plant that belongs to the family of Crassulaceae and is originally from Madagascar. B. pinnatum is widespread in tropical areas worldwide and has been used in traditional medicine. In Europe, Rudolf Steiner introduced B. pinnatum in anthroposophical medicine for the first time in 1921. Nowadays, B. pinnatum is a phytotherapeutic that is available in the form of leaf press juice and leaf extracts. Besides other potential indications, B. pinnatum is used and examined for the treatment of hyperactive conditions. Since 1970, B. pinnatum has been used successfully as a tocolytic agent in various hospitals. The efficacy could be confirmed by several empirical as well as by clinical studies. In vitro experiments demonstrated a relaxant effect of the aqueous leaf extract, leaf press juice, and the flavonoid fraction of B. pinnatum on spontanous human myometrial contractions. An additional experiment also showed a relaxant effect of the aqueous leaf extract on oxytocin-induced contractions.
Based on the fact that B. pinnatum has a relaxant effect on uterine smooth muscle, the inhibitory effect on urinary bladder smooth muscle was investigated. Overactive bladder (OAB) is a symptomatic syndrome, which affects many individuals and is defined as urinary urgency, with or without urge incontinence, usually with frequency and nocturia. Antimuscarinic drugs are the first-line pharmacotherapy prescribed to OAB patients. Due to anticholinergic side effects and insufficient therapeutic effects, an alternative phytopharmaceutical treatment is increasingly desired. Recently, a randomised, double-blind placebo-controlled study demonstrated a positive impact of B. pinnatum 50% chewable tablets against placebo by reducing the micturition frequency over 24 hours. In initial experiments, the B. pinnatum leaf press juice (5 - 10%) demonstrated a maximum relaxant effect on carbachol pre-contracted detrusor of 18.7% and an inhibition of electrically induced contractility by 74.6%.
For the administration of a phytotherapeutic, it is essential to have extensive knowledge about the constituents and their therapeutic effects and toxicity. This thesis describes the isolation and identification of B. pinnatum constituents in a methanolic leaf extract. Two phenolic acid derivatives and nine known flavonoid glycosides were isolated, including two new natural products belonging to the flavonoids, namely quercetin 3-O-α-L-arabinopyranosyl-(1 → 2)-α-L-rhamnopyranoside 7-O-β-D-glucopyranoside and myricetin 3-O-α-L-arabinopyranosyl-(1 → 2)-α-L-rhamnopyranoside. Bufadienolides, a substance group with sedative, positive inotropic and CNS-related activities is known in other Bryophyllum species, e.g. B. daigremontianum, and has been detected in the methanolic extract of B. pinnatum only in trace amounts. For this reason, four reference substances were isolated from B. daigremontianum and subsequently, bersaldegenin-1-acetate, bryophyllin A, bersaldegenin-3-acetate, and bersaldegenin-1,3,5-orthoacetate were detected in B. pinnatum.
Based on these findings, the methanolic leaf extract of B. pinnatum has been fractionated in a flavonoid, bufadienolide, and polar fraction. The effect of these three fractions on electrically stimulated porcine detrusor contractions was evaluated and compared with oxybutynin. The flavonoid fraction demonstrated a dose- and time-dependent inhibition of detrusor muscle contractility and a significant maximum inhibition of 78.7% using a 1 mg/mL concentration. The inhibitory effect of the flavonoid fraction was comparable with the maximum inhibition produced by the reference substance oxybutynin (10-6 M) at 78.1%. Bladder strips treated with the bufadienolide fraction (0.1 - 40 μg/mL) did not produce any inhibitory effect on detrusor contractility. The polar fraction demonstrated an unexpected dose-dependent inhibition. However, the observed effect may be explained by decreased pH caused by L-malic acid present in this fraction. These results suggest that flavonoids are important for the inhibition of detrusor contractility.
To examine the effect of B. pinnatum on human detrusor muscles, human bladder wall samples were obtained from patients after radical cystectomy. The in vitro experimental results confirmed the observed inhibitory effect of the leaf press juice and the flavonoid fraction; the effect seems to be more potent as the inhibition is observed at lower concentrations than with porcine detrusor strips.
The insights gained from the intensive inhibition of porcine and human detrusor contractility by Bryophyllum pinnatum are promising and support further investigations for the treatment of hyperactive conditions.