Localization and Organization of Scopolamine Biosynthesis in Duboisia myoporoides R. Br.

Tropane alkaloids (TAs), especially hyoscyamine and scopolamine, are important precursors for anticholinergic and antispasmodic drugs. Hyoscyamine and scopolamine are currently obtained at commercial scale from hybrid crosses of Duboisia myoporoides × Duboisia leichhardtii plants. In this study, we present a global investigation of the localization and organization of TA biosynthesis in a Duboisia myoporoides R. Br. wild-type line. The tissue-specific spatial distribution of TAs within D. myoporoides is presented, including quantification of the TAs littorine, 6-hydroxy hyoscyamine, hyoscyamine, scopolamine and, additionally, hyoscyamine aldehyde as well as scopolamine glucoside. Scopolamine (14.77 ± 5.03 mg g-1), and to a lesser extent hyoscyamine (3.01 ± 1.54 mg g-1) as well as 6-hydroxy hyoscyamine (4.35 ± 1.18 mg g-1), are accumulated in leaves during plant development, with the highest concentration of total TAs detected in 6-month-old plants. Littorine, an early precursor in TA biosynthesis, was present only in the roots (0.46 ± 0.07 mg g-1). During development, the spatial distribution of all investigated alkaloids changed due to secondary growth in the roots. Transcripts of pmt, tr-I and cyp80f1 genes, involved in early stages of TA biosynthesis, were found to be most abundant in the roots. In contrast, the transcript encoding hyoscyamine 6ß-hydroxylase (h6h) was highest in the leaves of 3-month-old plants. This investigation presents the spatial distribution of biochemical components as well as gene expression profiles of genetic factors known to participate in TA biosynthesis in D. myoporoides. The results of this investigation may aid in future breeding or genetic enhancement strategies aimed at increasing the yields of TAs in these medicinally valuable plant species.

Influence of Light, Temperature, and Macronutrients on Growth and Scopolamine Biosynthesis in Duboisia species.

Scopolamine is used in the pharmaceutical industry as a precursor in the organic synthesis of different classes of important active substances and is extracted in large scale from field grown Duboisia plants. Previous research revealed that plant growth as well as production of scopolamine and its derivatives varies strongly depending on abiotic factors. However, only a small amount of systematic research has been done on the influence of environmental conditions on scopolamine and biomass production, so far. In order to extend knowledge in this field, plants of three different genotypes (wild type Duboisia myoporoides and hybrids of D. myoporoides and Duboisia leichhardtii) were grown in climate chambers under controlled conditions in order to systematically analyse the influence of temperature (20, 24, 28 °C), light (50-300 µmol/m2 × s, 12, 18, 24 h per day) and macronutrients (nitrogen, calcium, potassium) on growth and scopolamine biosynthesis. The data indicate that light intensity and daily exposure to light have a major impact on scopolamine production and plant development, whereas temperature only shows a minor influence. Nitrogen (N) positively affects biomass production with increasing levels up to 4 mM, but is negatively correlated with scopolamine content. Calcium (Ca) shows a negative influence on scopolamine biosynthesis at increased levels above 1 mM as well. Potassium (K) neither affects biomass nor scopolamine production within the tested concentration range (0.05-4 mM). All in all, it can be concluded that light intensity and nitrogen supply are especially important regulating variables that can be applied in a targeted manner for influencing scopolamine and biomass production.

Influence of blister package design on usability among older adults.

BACKGROUND: Blisters packs are commonly used as packaging for oral drug products. Utilization problems among older adults with pharmaceutical packaging are well known from investigations of multi-unit dose containers, but there is a lack of studies focusing on blister packaging. This study was performed to identify design parameters that should be considered when developing blister packaging for drug products intended to be used by older adults. OBJECTIVES: To investigate the relationship between blister pack design and utilization problems among older adults. SETTING: Community-based study including 141 volunteers (age 80+) living at home and in assisted-living facilities. METHOD: In random order 5 blister packs of uniform shape but with different opening characteristics (variation of the force required to open the blisters and different mechanisms of opening) were tested by each participant. MAIN OUTCOME MEASURE: Opening was evaluated (difficulty, pain perception and overall assessment) by questioning of the participants. Additionally, data were collected as to whether participants managed to take out 4 tablets within 4 min test duration and whether they gave up opening. Reasons for utilization problems were examined by subgroup analysis, frequently made observations, and technological characterization of blisters. Results Blisters with thicker push-through foils were assessed as being inferior. Anyhow, most of participants (>84 %) were capable of opening each type of push-through blister within 4 min. By contrast, many of the participants were unable to open peel (30 %) and child-resistant peel-off push-through blisters (44 %) or gave up trying before the end of the test. In addition to medical conditions, vision and age of participants correlated with utilization problems. CONCLUSION: Blister pack design, including opening force and opening mechanism, can have significant impact on the usability of blister packs by older adults. The study identified several parameters that should be considered with respect to older adults when developing blister packaging for drug products.

Influences of heat seal lacquer thickness on the quality of blister packages.

A sealability of aluminium lidding foils against formable polymer materials of blister packages is usually achieved by a coating of aluminium with certain grammages of heat seal lacquers. To investigate influences of their thickness on quality of blister packages, lidding foils with different grammages of two lacquer types were manufactured. Sealing experiments (variation of temperature, pressure and sealing time) were performed. Sealed seam strengths were determined with mechanical tensile tests, tightness of cold form blisters were analysed by means of helium leakage tests. Time-dependent moisture uptake of stored blisters was monitored with micro-gas chromatography. By means of a simple calculation model the permeability coefficients of the heat seal lacquers were determined. Lidding foils with higher lacquer grammages showed significantly greater sealed seam strengths. Helium leakage tests showed only slight effects of heat seal lacquer grammage on tightness of blisters. But cold form blisters with lidding foils of higher lacquer grammages showed a significantly greater moisture uptake. Since the heat seal lacquers and the rigid polyvinyl chloride of the formable aluminium compound foils had similar permeability coefficients, the contribution of the lacquers to the total permeability of the investigated cold form blisters was only slightly.

Comparison of quantitative analysis techniques for the determination of heat seal lacquer layers on aluminum blister foils.

For decades a gravimetric method has been common standard for the determination of heat seal lacquers on aluminum blister foils. With the availability of appropriate techniques such as interferometric, infrared reflection absorption spectroscopic (IRRAS), beta backscatter, impedance spectroscopic and eddy current techniques respectively, more efficient determinations can be foreseen which are subject of the present communication. The different methods were compared to each other regarding parameters required for validation of analytical procedures according to the ICH guidelines Q2 (R1) such as linearity, precision, accuracy, robustness and quantitation limits. The interferometric, IRRAS and beta backscatter techniques were well suitable for the measurements. Using these techniques novel procedures applicable for routine quality control of pharmaceutical packaging materials are suggested.

The phytochemical profile and identification of main phenolic compounds from the leaf exudate of Aloe secundiflora by high-performance liquid chromatography-mass spectroscopy.

The phytochemical profile of Aloe secundiflora (Aloeaceae) and the identity of eight major compounds, including the two main constituents, have been determined from the leaf exudate of this ethnoveterinary used species from Kenya and Tanzania. Analytical HPLC-MS studies of the exudate have revealed that it comprises a mixture of phenolic compounds, mainly anthrones (aloenin, aloenin B, isobarbaloin, barbaloin and other aloin derivatives), chromones and phenylpyrones with a low content of polysaccharides and aliphatic compounds. The high percentage of anthrones in the exudate could provide a first line of evidence for the use of the plant in ethnoveterinary practices.