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  • Coleus (cell cultures)  (2)
  • [Ca2+] i  (2)
  • Animals
  • METEOROLOGY AND CLIMATOLOGY
  • Springer  (4)
  • 1990-1994  (4)
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  • 1990-1994  (4)
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  • 1
    Electronic Resource
    Electronic Resource
    Springer
    Planta 191 (1993), S. 18-22 
    ISSN: 1432-2048
    Keywords: Coleus (cell cultures) ; Hydroxycinnamic acid ester ; Rosmarinic acid ; Rosmarinic acid synthase (purification)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Rosmarinic acid synthase from cell cultures of Coleus blumei Benth. was purified to apparent homogeneity by fractionated ammonium sulfate precipitation (60–80% saturation), hydrophobic interaction chromatography, affinity chromatography and gel filtration. This purification procedure resulted in a 225-fold-enriched specific enzyme activity with a yield of 9%. The protein preparation was apparently pure according to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and two-dimensional gel electrophoresis. The apparent molecular mass determined by gel filtration and SDS-PAGE was 77 kDa, indicating that rosmarinic acid synthase is a monomeric enzyme.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1432-2048
    Keywords: Coleus (cell cultures) ; Hydroxycinnamic acid ester ; Phenylpropanoid metabolism ; Rosmarinic acid (biosynthetic pathway)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A biosynthetic pathway for rosmarinic acid is proposed. This pathway is deduced from studies of the enzymes detectable in preparations from suspension cells of Coleus blumei. Phenylalanine is transformed to 4-coumaroyl-CoA by the enzymes of the general phenylpropanoid pathway: phenylalanine ammonia-lyase (EC 4.3.1.5), cinnamic acid 4-hydroxylase (EC 1.14.13.11) and hydroxycinnamic acid:CoA ligase (EC 6.2.1.12). Tyrosine is metabolized to 4-hydroxyphenyllactate by tyrosine aminotransferase (EC 2.6.1.5) and hydroxyphenylpyruvate reductase. The ester can be formed from 4-coumaroyl-CoA and 4-hydroxyphenyllactate by the catalytic activity of rosmarinic acid synthase with concomitant release of CoA. Microsomal hydroxylase activities introduce the hydroxyl groups at positions 3 and 3′ of the aromatic rings of the ester 4-coumaroyl-4′-hydroxyphenyllactate giving rise to rosmarinic acid.
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  • 3
    ISSN: 1432-1424
    Keywords: patch clamp ; [Ca2+] i ; Na+ dependency ; RINm5F cell ; fura-2 ; whole cell
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Summary The patch-clamp technique and measurements of single cell [Ca2+] i have been used to investigate the importance of extracellular Na+ for carbohydrate-induced stimulation of RINm5F insulin-secreting cells. Using patch-clamp whole-cell (current-clamp) recordings the average cellular transmembrane potential was estimated to be −60±1 mV (n=83) and the average basal [Ca2+] i 102±6nm (n=37). When challenged with either glucose (2.5–10mm) ord-glyceraldehyde (10mm) the cells depolarized, which led to the initiation of Ca2+ spike potentials and a sharp rise in [Ca2+] i . Similar effects were also observed with the sulphonylurea compound tolbutamide (0.01–0.1mm). Both the generation of the spike potentials and the increase in [Ca2+] i were abolished when Ca2+ was removed from the bathing media. When all external Na+ was replaced with N-methyl-d-glucamine, in the continued presence of either glucose,d-glyceraldehyde or tolbutamide, a membrane repolarization resulted, which terminated Ca2+ spike potentials and attenuated the rise in [Ca2+] i . Tetrodotoxin (TTX) (1–2 μm) was also found to both repolarize the membrane and abolish secretagogue-induced rises in [Ca2+] i .
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  • 4
    ISSN: 1432-1424
    Keywords: patch-clamp ; fura-2 ; KATP channels ; [Ca2+] i ; insulin-secreting cell ; RINm5F cell ; diazoxide ; cromakalim (BRL 34915) ; tolbutamide
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Summary Patch-clamp and single cell [Ca2+] i measurements have been used to investigate the effects of the potassium channel modulators cromakalim, diazoxide and tolbutamide on the insulin-secreting cell line RINm5F. In intact cells, with an average cellular transmembrane potential of −62±2 mV (n=42) and an average basal [Ca2+] i of 102±6nm (n=37), glucose (2.5–10mm): (i) depolarized the membrane, through a decrease in the outward KATP current, (ii) evoked Ca2+ spike potentials, and (iii) caused a sharp rise in [Ca2+] i . In the continued presence of glucose both cromakalim (100–200 μm) and diazoxide (100 μm) repolarized the membrane, terminated Ca2+ spike potentials and attenuated the secretagogue-induced rise in [Ca2+] i . In whole cells (voltage-clamp records) and excised outside-out membrane patches, both cromakalim and diazoxide enhanced the current by opening ATP-sensitive K+ channels. Diazoxide was consistently found to be more potent than cromakalim. Tolbutamide, a specific inhibitor of ATP-sensitive K+ channels, reversed the effects of cromakalim on membrane potential and KATP currents.
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