Flavonoids enantiomer distribution in different parts of goldenrod (Solidago virgaurea L.), lucerne (Medicago sativa L.) and phacelia (Phacelia tanacetifolia Benth.)

Plant material is a rich source of valuable compounds such as flavanones. Their different forms influence bioavailability and biological activity, causing problems with the selection of plant material for specific purposes. The purpose of this research was to determine selected flavanone (eriodictyol, naringenin, liquiritigenin, and hesperetin) enantiomer contents in free form and bonded to glycosides by an RP‐UHPLC‐ESI‐MS/MS method. Different parts (stems, leaves, and flowers) of goldenrod (Solidago virgaurea L.), lucerne (Medicago sativa L.), and phacelia (Phacelia tanacetifolia Benth.) were used. The highest content of eriodictyol was found in goldenrod flowers (13.1 μg/g), where it occurred mainly as the (S)‐enantiomer, and the greatest proportion of the total amount was bonded to glycosides. The richest source of naringenin was found to be lucerne leaves (4.7 μg/g), where it was mainly bonded to glycosides and with the (S)‐enantiomer as the dominant form. Liquiritigenin was determined only in lucerne, where the flowers contained the highest amount (1.2 μg/g), with the (R)‐enantiomer as dominant aglycone form and the (S)‐enantiomer as the dominant glycosylated form. The highest hesperetin content was determined in phacelia leaves (0.38 μg/g), where it was present in the form of a glycoside and only as the (S)‐enantiomer. A comparison of the different analyte forms occurring in different plant parts was performed for the first time.     

Ethylene and IAA interactions in the inhibition of photoperiodic flower induction of <Emphasis Type="Italic">Pharbitis nil</Emphasis>

It has been shown that both IAA and ethylene application inhibit flower induction in the short-day plant Pharbitis nil. However application of IAA has elevated ethylene production in this plant, as well. Strong enhancement of ethylene production is also correlated with the night-break effect, which completely inhibits flowering. In order to determine what the role of IAA and ethylene is in the photoperiodic flower induction in Pharbitis nil, we measured changes in their levels during inductive and non-inductive photoperiods, and the effects of ethylene biosynthesis and action inhibitors on inhibition of flowering by IAA. Our results have shown that the inhibitory effect of IAA on Pharbitis nil flowering is not physiological but is connected with its effect on ethylene biosynthesis.     

Nerve Growth Factor Effect on Adenosine Transport in Cultured Chromaffin Cells

Chromaffin cells both recently isolated or in culture present a high-affinity adenosine transporter with a Km value of 1 microM. When cells were exposed to nerve growth factor (NGF; 10 ng/ml), the adenosine transporter affinity decreased to 3 microM. This value was maintained from 3 days after plating to the end of the culture period. A change in the transport capacity was observed, with a significant increase (approximately 200-260%) in NGF-cultured cells throughout the period studied.     

Initiation of primary cell cultures from human intracranial tumors on extracellular matrix from bovine corneal endothelial cells

Tissue specimens from 105 human gliomas and 57 human meningiomas were obtained at surgery, dissociated into single cells and small cell aggregates and then plated onto plain plastic tissue culture dishes and dishes which had been precoated with an extracellular matrix (ECM) derived from bovine corneal endothelium. In 80% of the glioma cases we observed a marked improvement in initial plating efficiency, colony formation and speed of attachment when cells were plated on ECM. In 5 cases cells attached only to the ECM-coated dishes but remained afloat in the untreated dishes. In addition it could be noted that over the first 2 days, those cells which had been initiated on ECM showed more signs of morphological differentiation, i.e., extension of cytoplasmic processes or formation of fiber networks between cell groups. If adaptation occurred and proliferation began in vitro, either immediately or after a several days' lag phase, both the ECM-cultured cells as well as those which slowly had adapted to culture on plastic could be passed on to untreated culture ware and perpetuated thereon. In the case of well-differentiated low-grade gliomas where no growth in culture took place, the cultures on ECM could at least be used for initial experiments in the primary cultures (P0). Meningiomas usually attached well to both, plastic or ECM. In 50% of our cases the plating efficiency was higher on ECM but after successful initial culture, the delay until the cells on plastic reached confluence in comparison with those on ECM was 1 or 2 days. Again there were 2 cases in which the cells would not plate on plastic. Here the cells which after 1 day were still afloat plated to more than 80% within the first 2 h after transfer to ECM. In all cases the cells from plastic and ECM cultures were indistinguishable and could be passed onto untreated dishes henceforth. In later culture stages ECM offers several advantages: It is easier to shift cells to serum-free defined culture conditions, the cells will grow at a faster rate on ECM when in higher passages and the maximal number of passages possible is higher on ECM.     

Hydrolysis of sunflower oil by means of hydrophobic membrane with lipolytic activity

Polytetrafluoroethylene (PTFE) membranes of different porosity were used for lipase from Rhizopus immobilization. The immobilized enzyme applied to sunflower oil hydrolysis achieved the activity of 1228 U/m² of the membrane area and the half-life time was calculated to be 7 days.     

The gene encoding dihydrolipoyl transacetylase from Azotobacter vinelandii. Expression in Escherichia coli and activation and isolation of the protein

The gene encoding the dihydrolipoyl transacetylase (E2) component from Azotobacter vinelandii has been cloned in Escherichia coli. High expression of the gene was found when the cells were grown for more than 14 h. The E2 produced was partially active, varying 10 and 90% in different experiments. By limited proteolysis of the protein it was shown that the catalytic domain was incorrectly folded, caused by formation of intermolecular or intramolecular S-S bridges. The enzyme was fully activated after unfolding in 2.5 M guanidine hydrochloride containing 2 mM dithiothreitol, followed by refolding by dialysis. Active E2 was isolated in a simple three-step procedure. It possessed a specific activity in the same order as that found after isolation of E2 from purified pyruvate dehydrogenase complex from A. vinelandii. Active E2 comprises about 7% of the total soluble cellular protein in the E. coli clone. By genetic manipulation, deletion mutants of E2 were created, one encoding the lipoyl domain and the N-terminal half of the pyruvate-dehydrogenase (E1)- and lipoamide-dehydrogenase (E3)-binding domain, the other encoding the catalytic domain and the C-terminal half of the E1- and E3-binding domain. In E. coli expression of both mutants was observed.