Identification of self-growth-inhibiting compounds lauric acid and 7-(Z)-tetradecenoic acid from Helicobacter pylori.

Helicobacter pylori growth medium is usually supplemented with horse serum (HS) or FCS. However, cyclodextrin derivatives or activated charcoal can replace serum. In this study, we purified self-growth-inhibiting (SGI) compounds from H. pylori growth medium. The compounds were recovered from porous resin, Diaion HP-20, which was added to the H. pylori growth medium instead of known supplements. These SGI compounds were also identified from 2,6-di-O-methyl-ß-cyclodextrin, which was supplemented in a pleuropneumonia-like organisms broth. The growth-inhibiting compounds were identified as lauric acid (LA) and 7-(Z)-tetradecenoic acid [7-(Z)-TDA]. Although several fatty acids had been identified in H. pylori, these specific compounds were not previously found in this species. However, we confirmed that these fatty acids were universally present in the cultivation medium of the H. pylori strains examined in this study. A live/dead assay carried out without HS indicated that these compounds were bacteriostatic; however, no significant growth-inhibiting effect was observed against other tested bacterial species that constituted the indigenous bacterial flora. These findings suggested that LA and 7-(Z)-TDA might play important roles in the survival of H. pylori in human stomach epithelial cells.

Bioactive metabolites isolated from Penicillium sp. YY-20, the endophytic fungus from Ginkgo biloba.

Six known metabolites, adenosine (1), methyl ß-D-ribofuranoside (2), adenine (3), 2'-deoxyadenosine (4), 3-methylpiperazine-2,5-dione (5) and 2'-deoxyuridine (6), were isolated from the extracts of the endophytic fungus Penicillium sp. YY-20 isolated from the root of Ginkgo biloba, and their structures were elucidated by spectroscopic methods. The antioxidant and growth-promoting activities of these compounds were first evaluated. The results indicated that compounds 1, 3 and 4 exhibited potential DPPH-scavenging activities compared with positive control. In addition, all the compounds (except 5) stimulated seed germination of Raphanus sativus, Brassica napus and Brassica chinensis but had weak stimulating effect on their root and hypocotyl growth.

Sports doping: emerging designer and therapeutic ß2-agonists.

Beta2-adrenergic agonists, or ß2-agonists, are considered essential bronchodilator drugs in the treatment of bronchial asthma, both as symptom-relievers and, in combination with inhaled corticosteroids, as disease-controllers. The use of ß2-agonists is prohibited in sports by the World Anti-Doping Agency (WADA) due to claimed anabolic effects, and also, is prohibited as growth promoters in cattle fattening in the European Union. This paper reviews the last seven-year (2006-2012) literature concerning the development of novel ß2-agonists molecules either by modifying the molecule of known ß2-agonists or by introducing moieties producing indole-, adamantyl- or phenyl urea derivatives. New emerging ß2-agonists molecules for future therapeutic use are also presented, intending to emphasize their potential use for doping purposes or as growth promoters in the near future.

Dummy-template molecularly imprinted solid phase extraction for selective analysis of ractopamine in pork.

Molecularly imprinted polymers (MIPs) for selective adsorption of ractopamine hydrochloride (RAC) were synthesised by an in situ method, in which salbutamol (SAL) was used as the dummy-template to avoid the template leakage. Scanning electron microscopy (SEM), mercury porosimerty and Fourier transform infrared spectroscopy (FTIR) were used to investigate the physical and morphological characteristics of the dummy-template MIPs. The test of adsorption selectivity indicated that the dummy-template MIPs exhibited high selectivity to RAC. The saturated adsorption capacity for RAC on dummy-template MIPs was 90.9 µg g(-1). Based on the dummy-template polymers, a liquid chromatography-mass spectrometry (LC-MS) method was developed for the selective analysis of RAC in real pork samples. The averages of intra- and inter-day accuracy ranged from 78.9% to 92.2% and from 90.7% to 93.1%, respectively. The RSD% of repeatability ranged from 1.9% to 6.3%, and the RSD% of intermediate precision ranged from 3.5% to 9.2%, while the limit of detection (LOD) was 0.02 µg kg(-1).

A sensitive and selective imprinted solid phase extraction coupled to HPLC for simultaneous detection of trace quinoxaline-2-carboxylic acid and methyl-3-quinoxaline-2-carboxylic acid in animal muscles.

A new molecularly imprinted polymer (MIP), selective for major metabolites of quinoxaline-1,4-dioxides, was prepared through bulk polymerisation using quinoxaline-2-carboxylic acid (QCA) as template, diethylaminoethylmethacrylate as functional monomer and ethylene glycol dimethacrylate as cross-linker in tetrahydrofuran. The synthesised MIP was characterised by Fourier transform infrared and adsorption experiments. MIP exhibited high affinity, fast kinetics for QCA and good selectivity for QCA and methyl-3-quinoxaline-2-carboxylic acid (MQCA). MIP obtained was used as a selective sorbent for molecularly imprinted solid phase extraction (MISPE) coupled with HPLC to detect QCA and MQCA. Under the optimal conditions, the limits of detection (S/N=3) of porcine, chicken and fish muscles were 0.1, 0.3, 0.1 µg/kg for QCA and 0.2, 0.3, 0.1 µg/kg for MQCA, respectively and good recoveries were obtained in the range from 60.0 to 119.4%. These results indicated the MISPE-HPLC procedure could be successfully used for the determination QCA and MQCA in animal muscles.

Use of plasma rich in growth factor for schneiderian membrane management during maxillary sinus augmentation procedure.

The aim of this pilot study was to present a novel technique for the management of the Schneiderian membrane during maxillary sinus lift surgery using plasma rich in growth factors (PRGF). Eight maxillary sinuses were augmented in 8 patients. Two small perforations of the Schneiderian membrane occurred during the lifting procedure, which were solved using the PRGF clot before grafting the site with PRGF and anorganic bovine bone. With the exception of 1 patient who experienced pain following an acute sinus infection after 3 days of uneventful healing, the patients' postoperative quality of life was generally good. The most common complication (50% of cases) was hematoma, which disappeared after 1 week. Despite the limitations of this study concerning the sample size and the study design, the use of PRGF may be helpful in reducing complications following sinus lift surgery. More well-designed studies, with larger sample size, are needed to validate this protocol.

Wound healing by a 3.2 kDa recombinant polypeptide from velvet antler of Cervus nippon Temminck.

Velvet antler (VA) is used in traditional Chinese medicine to treat a wide range of ailments including the enhancement of wound healing. A 3.2 kDa recombinant polypeptide of VA from sika deer was purified and compared to native polypeptides stimulation growth of NIH3T3 cells. Both stimulated growth in a dose-dependent manner (10-100 µg/ml). To study its wound healing properties, burn-wounded rats were topically administered with recombinant VA polypeptide or native polypeptide. Rats treated with 0.05 and 0.1% (w/w) polypeptides exhibited significant wound healing. As the yield of recombinant polypeptide was 40-fold higher than that of the native polypeptide, it may therefore be a useful biopharmaceutical.

Preliminary pharmacological evaluation of Martynia annua Linn leaves for wound healing.

OBJECTIVE: To evaluate the wound healing potential of fractions from ethanol extract of Martynia annua (M. annua) Linn leaves. METHODS: Ethanol extract of M. annua Linn leaves was fractionate into three different fractions (MAF-A, MAF-B and MAF-C) which were screened for wound healing potential using two models: excision and incision on rats. The thin layer chromatography (TLC) profile of all fractions were analyzed and TLC of luteolin was also done. The Povidone-Iodine Ointment was used as reference for comparision. Excision and incision wounds were created on dorsal portion of rats for study. Wound contraction, biochemical parameters (protein level and hydroxyproline level) and histopathological study were performed in excision wound model whereas incision model was used for determination of tensile strength. RESULTS: The wound contraction and tensile strength of skin tissues were observed significantly greater in MAF-C fraction treated group than other two fractions (P<0.01). In excision wound method (on day 18) protein content and hydroxyproline were found significantly higher in MAF-C group than control group (P<0.01). Histopathological study also showed better angiogenesis, matured collagen fibres and fibroblast cells as compared with the control group. CONCLUSIONS: In conclusion, our findings suggest that fraction MAF-C from ethanol extract of M. annua leaves is found most effective in wound healing.

Effect of bioregulators isolated from the liver, blood serum, and bile of mammals on the state of newt liver tissue in organotypic culture.

We developed models of in vitro organotypic culturing of newt liver tissue with and without adhesion to the substrate. The effects of bioregulators isolated from mammalian liver, blood serum, and bile were studied on the developed models and their specificity was demonstrated. The state of the liver was evaluated by the area of clusters of pigmented cells and by the number of mitoses in the connective tissue cells of the cortical layer. These bioregulators exhibited their biological effects only under conditions of roller organotypic culturing of newt liver tissue.

Plant-growth regulator, imidazole-4-carboxamide, produced by the fairy ring forming fungus Lepista sordida.

Rings or arcs of fungus-regulated plant growth occur often on the floor of woodlands, in agricultural areas, and in grasslands worldwide. These rings are commonly called "fairy rings". A plant-growth regulating compound was isolated from a fairy ring forming fungus, Lepista sordida , and its chemical structure was identified as imidazole-4-carboxamide (ICA) by spectroscopic analyses including single-crystal X-ray diffraction techniques. ICA inhibited the growth of turfgrass and rice seedling. On the other hand, in a greenhouse experiment, this compound increased rice grain yield by 26% compared with control.

Use of coated capillaries for the electrophoretic separation of stereoisomers of a growth hormone secretagogue.

The diastereoisomeric separation of peptidomimetics of hexarelin, a strong growth hormone secretagogue, in CE has been studied. Highly sulfated-gamma-CD was found to be an appropriate selector for the separation of the stereoisomers. However, non-repeatable analyses were obtained on bare fused silica capillary due to the progressive adsorption of the analytes on the capillary wall. Two types of polyelectrolyte coating agents were tested to prevent this phenomenon. Coating with neutral polyethylene oxide was found to be efficient but resulted in a very long analysis time (about 40 min). Coating with cationic poly(diallyldimethylammonium) chloride was found both to prevent analyte adsorption, reduce analysis time and alter separation selectivity. EOF measurement revealed that the highly sulfated-gamma-CDs were strongly adsorbed on the poly(diallyldimethylammonium) chloride coating surface yielding a stable strong cathodic EOF, which considerably reduced analysis time (about 12 min). Very good repeatability of analysis was obtained (RSD(migration time)<1%).

Isolation and characterization of brassinosteroids from algal cultures of Chlorella vulgaris Beijerinck (Trebouxiophyceae).

The brassinosteroids (BRs) occur ubiquitously in the plant kingdom. The occurrence of BRs has been demonstrated in almost every part of higher plants, such as pollen, flower buds, fruits, seeds, vascular cambium, leaves, shoots and roots. In this study, BRs were isolated and identified in the culture of wild-type Chlorella vulgaris. Seven BRs, including teasterone, typhasterol, 6-deoxoteasterone, 6-deoxotyphasterol, 6-deoxocastasterone, castasterone and brassinolide, were identified by GC-MS. All compounds belong to the BR biosynthetic pathway. The results suggest that early and late C6 oxidation pathways are operating in C. vulgaris. This study represents the first isolation of BRs from C. vulgaris cultures.

Platelet-rich plasma and platelet gel preparation using Plateltex.

BACKGROUND: The platelet gel is made by embedding concentrate platelets within a semisolid (gel) network of polymerized fibrin. It is believed that this blood component will be used more and more in the treatment of several clinical conditions and as an adjunctive material in tissue engineering. Several systems are available to produce platelet-rich plasma (PRP) for topical therapy. Recently, a new system became commercially available, Plateltex. Here we report the technical performance of this system in comparison with the performance of other commercially available systems: PRGF, PRP-Landesber, Curasan, PCCS, Harvest, Vivostat, Regen and Fibrinet. MATERIAL AND METHODS: Both the PRP and the gel were prepared according to the manufacturer's directions. The blood samples of 20 donors were used. The yield, the efficiency, and the amount of platelet-derived growth factor AB (PDGF-AB), transforming growth factor beta, vascular endothelial growth factor and fibroblast growth factor were measured in the resulting PRP. The feature of the batroxobin-induced gelation was evaluated. RESULTS: The yield, the collection efficiency and the growth factor content of Plateltex were comparable to those of most of the other available systems. The gelation time was not dependent on the fibrinogen concentration; however, it was strongly influenced by the contact surface area of the container where the clotting reaction took place (P < 0.0001). CONCLUSIONS: Plateltex provided platelet recovery, collection efficiency and PDGF-AB availability close to those provided by other systems marketed with the same intended use. Batroxobin, the enzyme provided to induce gelation, acts differently from thrombin, which is used by most other systems. Platelets treated with thrombin become activated; they release their growth factors quickly. Furthermore, thrombin-platelet interaction is a physiological mechanism that hastens the clot-retraction rate. On the contrary, platelets treated with batroxobin do not become activated; they are passively entrapped within the fibrin network, and their growth factor release occurs slowly. In these conditions, the clot retraction takes longer to occur. According to these differences between thrombin and batroxobin, it is expected that batroxobin-induced PRP activation will tailor slow release of the platelet content, thus, providing longer in loco availability of trophic factors. In selected clinical conditions, this durable anabolic factor availability might be preferable to quick thrombin-induced growth factor release.

Isolation and identification of F-spondin in the boar testis and its production during testis growth.

F-spondin/vascular smooth muscle cell growth-promoting factor (VSGP), purified from the follicular fluid of adult bovine ovaries, has been identified as a promoter of neuronal differentiation and vascular smooth muscle growth. The objectives of the present study were (1) to clarify whether F-spondin is also produced in the testis, which is ontogenically equivalent to the ovary, and (2) to examine whether production of this protein changes with testicular growth. To isolate F-spondin from the testis, testicular homogenates obtained from 8-week-old boars were sequentially subjected to heparin-Sepharose chromatography, diethylaminoethyl (DEAE)-Sepharose chromatography, and reverse-phase high-performance liquid chromatography (RP-HPLC). The isolated protein had a molecular mass of approximately 110 kDa and was cross-reactive with anti-F-spondin antibody by Western blotting. The purified protein was further characterized by amino acid sequence analysis of its internal peptide. The sequence obtained was GEQCNIVPDN VD, and a homology search indicated that the purified protein is a homologue of rat, human, and bovine F-spondin. By fractionation of the same amounts of testis tissue obtained from 1-, 8-, 16-, and 40-week-old boars, we analyzed age-related production of F-spondin in the testis. Western blotting of the fractions obtained from RP-HPLC revealed the presence of a band at approximately 110 kDa, corresponding to F-spondin, in the testes obtained from boars between 1 and 16 weeks old, but this band was not detected at 40 weeks. These results clearly indicate that (1) the porcine testis produces F-spondin and that (2) production of this protein is evident in the immature porcine testis, but not the adult testis.

Quantification of growth factors in allogenic bone grafts extracted with three different methods.

BACKGROUND: Bony allografts are used for defect filling. A reliable sterilization method is the peracetic acid-ethanol sterilization procedure (PES). Several studies showed the antimicrobiological efficacy of this method. Aim of this study was the quantification of growth factors necessary for bone formation in PES sterilized allografts (n = 9). METHODS: To extract the growth factors from the tissue three different methods were used: (a) use of collagenase 1 for extraction, (b) incubation of the material in a proteinase inhibitor cocktail (Complete), and (c) extraction with guanidine HCl. The supernatants from the different methods were analyzed for the total protein concentration and different growth factors. RESULTS: The extraction with guanidine HCl resulted in the highest amount of protein measurable in the supernatants of the samples. For comparison of the individual growth factor values the results were normalized to the protein content. The highest growth factor amount/protein was detectable for BMP-2 using the GndHCL method followed by FGFa, IGF-I, TGF-beta1, VEGF, and PDGF. Comparing the three extraction methods, significant differences were measured for the individual growth factor content. CONCLUSION: PES sterilized bony allografts contain several growth factors. Depending on the extraction method, the quantity of the analyzed growth factors varies.

Neurite outgrowth activity of cyathane diterpenes from Sarcodon cyrneus, cyrneines A and B.

Two new cyathane diterpenes, cyrneines A (1) and B (2), were isolated from the mushroom Sarcodon cyrneus. The structures of the novel diterpenoids were determined by analysis of their spectroscopic data. Neither cyrneine A nor cyrneine B at 100 microM showed cytotoxicity as determined by LDH analysis. The stimulating activity on neurite outgrowth of cyrneines was evaluated. Rat pheochromocytoma cells (PC12), used as a model system of neuronal differentiation, were cultured with cyrneine A (100 microM), cyrneine B (100 microM) or NGF (100 ng/ml) for 24 h. Interestingly, cyrneines A and B significantly promoted neurite outgrowth in addition to NGF as a positive control.

Growth promoting substance in yeast extract for methylotrophic growth of Candida boidinii.

A growth-promoting substance (GPS) for methylotrophic yeasts was purified from yeast extract. The purified GPS showed growth promotion by reducing the lag time of the growth of methylotrophic Candida boidinii, while the growth rate at the exponential phase was the same as that of growth without GPS. GPS was finally identified to be L-proline by the criteria of instrumental analysis and chemical shift assignments.

A ceramide and cerebroside from the starfish asterias amurensis Lütken and their plant-growth promotion activities.

The new phytosphingosine-type ceramide asteriaceramide A (1) and glucocerebroside asteriacerebroside G (2), together with two known cerebrosides, asteriacerebrosides A and B, were isolated from lipophilic fractions of the whole bodies of the Northern Pacific starfish Asterias amurensis Lütken. The water-soluble fraction afforded two known asterosaponins, glycoside B(2) and asterosaponin-1. The structures of 1 and 2 were determined on the basis of chemical and spectroscopic evidence as (2S,3S,4R,13Z)-2-[(2'R)-2-hydroxyhexadecanoylamino]-13-docosene-1,3,4-triol (1) and 1-O-(beta-d-glucopyranosyl)-(2S,3R,4E,13Z)-2-[(2'R)-2-hydroxytetradecanoylamino]-4,13-docosadiene-1,3-diol (2). Compounds 1, 2, and asteriacerebrosides A and B promoted plant growth in sprouts of Brassica campestris.

Fibroblast growth stimulation by extracts and compounds of Onosma argentatum roots.

The roots of Onosma argentatum are used traditionally in Turkey for wound healing and burns. The n-hexane-dichloromethane extract of the roots, and four shikonin derivatives (deoxyshikonin, acetyl shikonin, 3-hydroxy-isovaleryl shikonin and 5,8-O-dimethyl acetyl shikonin) isolated from the n-hexane-dichloromethane extract were investigated for their ability to stimulate the growth of human amnion fibroblasts. A range of concentrations was studied and the extract found to stimulate the growth of human amnion fibroblasts in vitro at 0.1 microg/mL whilst 5,8-O-dimethyl acetyl shikonin had the same effect at 0.05-5 microg/mL, although cytotoxicity was observed at 50 microg/mL for all samples. The extract and all the other isolated compounds showed cytotoxicity at 10 microg/mL with the extract and 3-hydroxy-isovaleryl shikonin showing cytotoxicity at 5 microg/mL. It is suggested that any wound healing effect of the roots of Onosma argentatum might be partly due to an additive effect of the shikonin derivatives present.

Intact growth factors are conserved in the extracellular matrix of ancient human bone and teeth: a storehouse for the study of human evolution in health and disease.

For the first time we have extracted, solubilized and identified growth factors, such as insulin growth factor II (IGF-II), bone morphogenetic protein-2 (BMP-2), and transforming growth factor-beta (TGF-beta), from archaeological compact human bone and tooth dentin dating from the late pre-ceramic pottery Neolithic (late PPNB) and the early Middle Ages. These factors are typical of special physiological or pathological situations in the metabolism of bone. The extracellular matrix proteins from bone and teeth of individuals from the late PPNB and early Middle Ages were separated by 2-D electrophoresis and more than 300 different protein spots were detected by silver staining. The matrix protein patterns of compact bone and tooth from the same individual (early Middle Ages) are very different and only 16% of the protein spots were detected in both compact bone and tooth dentin.