• Fisheries and Aquatic Sciences
• /
• v.12 no.1
• /
• pp.16-23
• /
• 2009

Lipids were extracted from marine rotifer, Brachionus rotundiformis in order to examine the functionality of lipid enzymatic modification. The fatty acids, palmitic, linoleic, oleic and stearic acids were the dominant forms accounting for approximately 35.8%, 21.5%, 15.9% and 7.7% of the total lipid content, respectively. Lipid fractions were categorized as neutral lipids (38.5%), glycolipids (45.9%) and phospholipids (17.6%), and after extraction from the rotifer were isolated by thin-layer chromatography (TLC) as free fatty acids (FFA), monoacylglycerol (MAG), diacylglycerol (DAG) and triacylglycerol (TAG). The production of polyunsaturated fatty acid (PUFA) concentrate from rotifer lipids was studied using lipase-catalyzed hydrolysis. In addition, rotifer lipids were modified by hydrolysis using lipases such as porcine pancreas, Candida rugosa and Rhizomucor miehei. The lipase from Rhizomucor miehei was effective in extracting linoleic acid (C 18:2), while the lipase from Candida rugosa was effective in palmitic acid (C16:0) extraction.

• The Korean Journal of Physiology and Pharmacology
• /
• v.2 no.2
• /
• pp.251-260
• /
• 1998

Cromakalim (BRL 34915), known as an airway smooth muscle relaxant, inhibited the releases of mediators in the antigen-induced mast cell activation. It has been suggested that cromakalim, in part, inhibited mediator releases by inhibiting the initial increase of 1,2-diacylglycerol (DAG) produced by the activation of the other phospholipase system which is different from phosphatidylcholine-phospholipase D pathway. The aim of this study is to further examine the inhibitory mechanism of cromakalim on the mediator release in the mast cell activation. Guinea pig lung mast cells were purified by using enzyme digestion and percoll density gradient. In purified mast cells prelabeled with $[^3H]PIP_2$, phospholipase C (PLC) activity was assessed by the production of $[^3H]$insitol phosphates. Protein kinase C (PKC) activity was assessed by measuring the protein phosphorylated from mast cells prelabeled with $[{\gamma}-32P]ATP$, and Phospholipase $A_2\;(PLA_2)$ activity by measuring the lyso-phosphatidylcholine produced from mast cell prelabeled with 1-palmitoyl-2-arachidonyl $phosphatidyl-[^{14}C]choline$. Histamine was assayed by fluorometric analyzer, and leukotrienes by radioimmunoassay. The PLC activity was increased by activation of the passively sensitized mast cells. This increased PLC activity was decreased by cromakalim pretreatment. The PKC activity increased by the activation of the passively sensitized mast cells was decreased by calphostin C, staurosporine and cromakalim, respectively. The $PLA_2$ activity was increased in the activated mast cells. The pretreatment of cromakalim did not significantly decrease $PLA_2$ activity. These data show that cromakalim inhibits histamine release by continuously inhibiting signal transduction processes which is mediated via PLC pathway during mast cell activation, but that cromakalim does not affect $PLA_2$ activity related to leukotriene release.

• Applied Biological Chemistry
• /
• v.41 no.6
• /
• pp.414-420
• /
• 1998

We studied the difference effects of dietary ${\alpha}-linolenic\;acid\;({\alpha}-LA,\;18:3\;n-3)$, eicosapentaenoic acid (EPA, 20:5 n-3) and docosahexaenoic acid (DHA, 22:6 n-3) on the lowering of triacylglycerol in the liver and serum on lipid metabolism in rats. Rats were fed semipurified diets containing 10% fat with constant polyunsaturated/monounsaturated/saturated fatty acids (1:1:1) and n-6/n-3 ratio (1:2). EPA (98%) and DHA (98%) were added in diets as the ethyl esters. The concentration of liver triacylglycerol was significantly lower in rats fed both EPA and DHA than in those fed ${\alpha}-LA$. The concentration of liver phospholipid was significantly higher in rats fed DHA than in those fed ${\alpha}-LA$ and EPA. Both EPA and DHA reduced serum triacylglycerol concentration compared with ${\alpha}-LA$, but this effect was more pronounced in the EPA diet. The activity of phophatidate phosphohydrolase in the liver microsome was significantly lower in rats fed both EPA and DHA than in those fed ${\alpha}-LA$. but, there was no significant difference on the activities of diacylglycerol acyltransferase among the three groups. The concentration of liver triacylglycerol were correlated with changes in the microsomal phosphatidate phosphohydrolase activity (r=0.84). Hepatic NADPH generating enzyme, glucose-6-phosphate dehydrogenase was more effective to reduce the activity in rats fed both EPA and DHA than in those fed ${\alpha}-LA$. In conclusion, EPA or DHA reduced the hepatic triacylglycerol concentration by inhibiting microsomal phosphatidate phosphohydrolase, thereby inhibiting synthesis of triacylglycerol in the liver.

• The Journal of Korean Physical Therapy
• /
• v.20 no.3
• /
• pp.61-68
• /
• 2008

Purpose: Protein kinase C (PKC) is a member of a family of serine/threonine kinases that are activated by diacylglycerol (DG) and PKC stimulants. PKC play a key role in signal transduction, including muscle contraction, cell migration, apoptosis, cell proliferation and differentiation. However, the mechanism relating mitogen-activated protein kinases (MAPKs) and PKC, especially in the volume-dependent hypertensive state, remains unclear. Methods: In the present study, I investigated the relationship between PKC and MAPKs for isometric contraction, PKC translocation, and enzymatic activity from normotensive sham-operated rats (NSR) and aldosterone-analogue deoxycorticosterone acetate (DOCA) hypertensive rats (ADHR). Results: Systolic blood pressure was significantly increased in ADHR than in NSR. Physiological salt solution (PSS)-induced resting tension and the intracellular $Ca^{2+}$ concentration ([$Ca^{2+}{_i}$]) were different in the ADHR and NSR. The expression of PKC$\alpha$, PKC$\beta$II, PKC$\delta$, PKC$\varepsilon$ and PKC$\xi$ were different between the cytoplasmic and membranous fractions. However, expression of the PKC isoforms did not differ for the ADHR and NSR. The use of 12-deoxyphorbol 13-isobutyrate (DPB, a PKC stimulant) induced isometric contraction in $Ca^{2+}$-free medium, which was diminished in muscle strips from ADHR as compared to NSR. Increased vasoconstriction and phosphorylation induced by the use of 1 ${\mu}$M DPB were inhibited by treatment with 10 ${\mu}$M PD098059 and 10 ${\mu}$M SB203580, inhibitors of extracellular-regulated protein kinase 1/2 (ERK1/2) and p38 MAPK from ADHR, respectively. Conclusion: These results suggest that the development of aldosterone analogue-induced hypertension is associated with an altered blood pressure, resting tension, [$Ca^{2+}{_i}$], and that the $Ca^{2+}$-independent contraction evoked by PKC stimulants is due to the activation of ERK1/2 and p38 MAPK in volume-dependent hypertension. Therefore, it is suggested that PKC activity affects volume-dependent hypertension and the need to develop cardiovascular disease-specialized physical therapy.

• Applied Biological Chemistry
• /
• v.47 no.2
• /
• pp.251-257
• /
• 2004

The root of lxeris dentata forma albiflora was extracted with 80% aqueous MeOH and solvent fractionated with EtOAc, n-BuOH and water, successively. From the EtOAc and n-BuOH fractions, four sesquiterpene compounds were isolated through the repeated silica gel and ODS column chromatographies. The chemical structures were determined as zaluzanin C (1), $9{\alpha}-hydroxyguaian-4(l5),10(14),11(13)-triene-6,12-olide$ (2), $3{\beta}-O-{\beta}-D-glucopyranosyl-8{\alpha}-hydroxyguaian-4(15),10(14 )-diene-6,12-olide$ (3), and $3{\beta}-O-{\beta}- D-glucopyranosyl-8{\beta}hydroxyguaian-10(14)-ene-6,12-olide$ (4) through the interpretation of several spectral data including 2D-NMR. Some showed the inhibitory effects on DGAT (Diacylglycerol acyltransferase), ($IC_{50}$ values of 1, 2: 0.13, 0.10 mM), the catalyzing enzymes of the intracellular esterification of diacylglycerol and FPTase (Famesyl-protein transferase), ($IC_{50}$ values of 1, 2: 0.15, 0.18 mM), the farnesylation enzyme for Ras protein charge of cancer promotion.

• Korean Journal of Plant Tissue Culture
• /
• v.27 no.5
• /
• pp.375-377
• /
• 2000

It is now generally accepted that a phosphoinositide cycle is involved in the transduction of a variety of signals in plant cells. In animal cells, the hydrolysis of phosphatidyl-4,5-bisphosphate catalysed by phosphatidylinositol - specific phospholipase C yields to D-myo-inositol - 1,4,5-trisphosphate and diacylglycerol, which are well known second messengers. The binding of InsP$_3$to a receptor located on the endoplasmic reticulum triggers a calcium release from the endoplasmic reticulum. We have detected and partially characterised key components of phosphoinositide signaling. First, tobacco microsomal fraction and plasma membrane PI-PLC. Consecutively, using a radioligand binding assay we have identified a $Ca^{2+}$ -dependent high affinity InsP$_3$binding site in microsomal membrane fraction vesicle preparation and then we have measured inositol-1,4,5-trisphosphate induced calcium release from tobacco microsomal fraction. These findings suggest that phosphoinositide signaling system is present and operates in the tobacco suspension culture.e.

• Journal of the Korean Society of Food Science and Nutrition
• /
• v.32 no.8
• /
• pp.1200-1205
• /
• 2003

Structured lipids (SL) were synthesized by transesterification of corn oil and conjugated linoleic acid (CLA) in the continuous type reactor using sn-1,3 specific Rhizomucor miehei lipase. The parameters of reaction were observed in terms of flow rate, temperature, and substrate molar ratios. The highest incorporation of CLA was obtained with 1 mL/min flow rate, 55$^{\circ}C$ and 1 : 3 (corn oil/CLA) molar ratio, showing 10.26 ㏖%. When different reaction temperatures and substrate ratios were studied, the highest incorporation was obtained at $65^{\circ}C$ (17.33 ㏖%) and 1 : 5 (corn oil/CLA) ratio (17.50 ㏖%), respectively. After pancreatic lipase analysis, most of all CLA were found at sn-1,3 position. The iodine values of obtained SLs ranged from 110 to 120. From the neutral lipid analysis by normal-phase HPLC, produced SLs composed of 99.35 ∼ 99.89% triacylglycerols, 0.11 ∼ 0.51% 1,2- and 1,3-diacylglycerols, and 0.06 ∼ 0.22% monoacylglycerols.

• Microbiology and Biotechnology Letters
• /
• v.32 no.1
• /
• pp.84-90
• /
• 2004

Diglyceride (DG) was prepared by reaction of soybean oil and glycerol in the presence of lipase. The initial rate of DG production was greatly affected by the amount of lipase. However the DG content at equilibrium was hardly affected by the amount of lipase added to the reaction mixture. The initial rate of FFA formation was highly affected by the moisture content between 0.5 and 2.3%, but at higher water content (3.3-5.2%), there was a small increase in the rate. And DG content at equilibrium slowly increased with the increase of the water content in glycerol up to 4.4%. However, there was a sharp decrease in DG content at higher water content (5.2-6.4%) due to higher free fatty acid production. The highest yield of DC was obtained at the temperature ranges of 30-5$0^{\circ}C$. The final yield of DG was not dependent on the glycerol (GL) to triglyceride (TG) molar ratio. However, at the molar ratio of 0.75:1 (GL/TG), the enzyme-catalyzed reaction was highly efficient and utilized all the glycerol. In optimized conditions for glycerolysis a yield of approximately 45% DG was obtained. 66% of total DG was 1,3-DG.

• Journal of Microbiology and Biotechnology
• /
• v.25 no.11
• /
• pp.1827-1834
• /
• 2015

The SMG1 lipase from Malassezia globosa is a newly found mono- and diacylglycerol (DAG) lipase that has a unique lid in the loop conformation that differs from the common alpha-helix lid. In the present study, we characterized the contribution of three residues, L103 and F104 in the lid and F278 in the rim of the binding site groove, on the function of SMG1 lipase. Site-directed mutagenesis was conducted at these sites, and each of the mutants was expressed in the yeast Pichia pastoris, purified, and characterized for their activity toward DAG and p-nitrophenol (pNP) ester. Compared with wild-type SMG1, F278A retained approximately 78% of its activity toward DAG, but only 11% activity toward pNP octanoate (pNP-C8). L103G increased its activity on pNP-C8 by approximately 2-fold, whereas F104G showed an approximate 40% decrease in pNP-C8 activity, and they both showed decreased activity on the DAG emulsion. The deletion of 103-104 retained approximately 30% of its activity toward the DAG emulsion, with an almost complete loss of pNP-C8 activity. The deletion of 103-104 showed a weaker penetration ability to a soybean phosphocholine monolayer than wild-type SMG1. Based on the modulation of the specificity and activity observed, a pNP-C8 binding model for the ester (pNP-C8, N102, and F278 form a flexible bridge) and a specific lipid-anchoring mechanism for DAG (L103 and F104 serve as "anchors" to the lipid interface) were proposed.

• Biomedical Science Letters
• /
• v.14 no.2
• /
• pp.69-76
• /
• 2008

${\gamma}$-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the central nervous system, and its actions are mediated by subtypes of GABA receptors named as $GABA_A$, $GABA_B,\;and\;GABA_C,\;GABA_A$, receptor consisting of ${\alpha},\;{\beta},\;{\gamma}\;and\;{\delta}$ subunits is a heterooligomeric ligand-gated chloride channel. This study was performed to investigate regulation of $GABA_A$ receptor by protein kinase C(PKC). Ion currents were recorded using gramicidine-perforated patch and whole cell patch clamp. mRNA encoding the subunits of PKC expressed in major pelvic ganglion (MPG) neurons was detected by using RT-PCR. The GABA-induced inward current was increased by PKC activators and decreased by PKC inhibitors, respectively. These effects were not associated with intracellular $Ca^{2+}$ and GAG (1-oleoyl-2-acetyl-sn-glycerol), a membrane permeable diacylglycerol (DAG) analogue. These results mean that the subfamily of PKC participating in activation of $GABA_A$ receptor would be an atypical PKC (aPKC). Among theses, ${\xi}$ isoform of aPKC was detected by RT-PCR. Taking together, we suggest that excitable $GABA_A$ receptor in sympathetic MPG neuron seemed to be regulated by aPKC, particular in ${\xi}$ isoform. The regulatory roles of PKC on excitatory $GABA_A$ receptors in sympathetic neurons of MPG may be an important factor to control the functional activity of various pelvic organs such as bowel movement, micturition and erection.