Caraway (Carum carvi L.): Anther Culture and Production of DH Plants Caraway.

We describe the production of doubled haploids through anther culture in caraway. Induction conditions for the cultivation of donor plants, anther collection, composition of culture media, and physical induction conditions for embryogenesis have been described. As a result, responsive lines with numerous haploid embryo production were obtained, which after colchicine treatment became fertile. From a practical point of view, two doubled haploid populations are tested under field conditions.

Forming a Large-Scale Droplet Array in a Microcage Array Chip for High-Throughput Screening.

Forming a large-scale droplet array plays an important role for microfluidic droplet-based high-throughput screening and analysis. Herein, we describe a simple and rapid method to form a large-scale two-dimension (2D) droplet array by using a microcage array chip. Differing from the previous droplet array formation methods, microcages formed by being surrounded by multiple micropillars could rapidly spread the oil phase through the gaps between the micropillars and trap droplets with fast speed and convenient operation. We formed a large-scale 2D monolayer droplet array containing approximately 1 000 000 droplets on a 5.5 cm × 5.5 cm microcage array chip within 90 s. The droplets in the droplet array could be further incubated for performing biochemical reactions and detected by a fluorescence microscope in real time. Due to the exact trapping and positioning functions of the microcages to the droplets, single targeted fluorescent droplets in the array could be individually picked out and transferred to culture medium by a microfluidic droplet-handling robot with a success rate of 100% and a picking operation time of 2.0 s for one droplet under the optimized conditions. This system was validated in the screening of the bacterium expressing the esterase AFEST from a mixture of AFEST-expressing and phosphotriesterase-expressing E. coli cells, achieving a success rate of 100% for single-droplet picking while maintaining the bacterial cell viability. The present system has the potential to be applied in high-throughput screening and analysis, such as single cell analysis, directed evolution, and drug screening.

Ipomoea obscura (L.) enhances the functions of immunological effector cells, inhibits proinflammatory cytokines and nitric oxide production by LPS induced macrophages.

Most of the synthetic chemotherapeutic agents available today are immunosuppressant, cytotoxic and exerts variety of side effects. Botanical based immunomodulators are often employed as supportive or adjuvant therapy to overcome the undesired effects of cytotoxic chemotherapeutic agents and to restore normal health. The methanolic extract of traditionally important medicinal plant Ipomoea obscura exhibited immunomodulatory activity in BALB/c mice. Intraperitoneal administration of five doses of the extract (10 mg/kg body wt) was found to enhance the total WBC count (13912 cells/mm(3)) on the 12(th) day, bone marrow cellularity (28.9 x 10(6)cells/femur) and number of alpha-esterase positive cells (1246 cells/4000 cells). Treatment with the extract along with the antigen, sheep red blood cells (SRBC), produced an enhancement in the circulating antibody titer and the number of plaque forming cells (PFC) in the spleen. Maximum number of PFC (267.6 PFC/10(6) spleen cells) was obtained on the 6(th) day. At the same time administration of Ipomoea obscura extract significantly reduced the elevated levels of proinflammatory cytokines and nitric oxide production by lipopolysaccharide stimulated macrophages. These results indicate the immunomodulatory activity of the alcoholic extract of Ipomoea obscura.

[Study on isozymes in six species of Curcuma].

OBJECTIVE: To explore genetic relationships of the 39 materials in six species of Curcuma. METHOD: The peroxidase isozyme (POD) and esterase isozyme (EST) were studied using vertical slab polyacrylamide gel electrophoresis (PAGE) technique, and the zymograms were analyzed using the software of NTSYSpc2. 1. RESULT: The interspecific zymogramatic differences were obvious. Each species possessed its own specific zymogram distinguishing form the others. In the analysis of EST isozyme, C. phaeocaulis, C. wenyujin, C. kwangsiensis and C. chuanhuangjiang had their own specific zymogram. In the analysis of POD isozyme, just C. phaeocaulis and C. kwangsiensis had their specific zymogram. CONCLUSION: The genetic relationships are not associated with the geographical distributions and the genetic relationship between C. longa and C. sichuanensis are very close.

[Primary effects on Isatis indigotica after spaceflight].

Isatis indigotica carried by the Chinese first spaceship "Shenzhou" was studied in order to find the mutation after spaceflight. TLC differentiation experiments showed no distinct discrepancy among the samples of spaceflight and non-space-flight, and the same color spot appeared corresponding to the location of the arginine. Isoenzymes of esterase and peroxidase were studied with PAGE. Isoenzymes of esterase were difference among the samples. To peroxidases, little difference was found with them. The ratio of dry weight and extract contents showed out the mutation has emerged after spaceflight, but some characters were unstable. It is necessary for further study.

[Investigation on biologically characterst of wild species and herbal quality of Changium smyrnioides].

OBJECTIVE: To investigate the species resources of Changium smyrnioides. METHOD: Nine habitat populations of C. smyrnioides, included with population environment, time limit, morphological characters and so on were investigated. The phylogenetic relationship of different habitat populations of C. smyrnioides was studied by isoenzyme electrophoretic analysis of the EST. RESULT: The results showed that wild resources of C. smyrnioides were endangered. Seed germinating rate of cultivated plant was significantly higher than that of wild seed, but polysaccharide content in the root of wild species was significantly higher than that of cultivated plant. There existed a geographic phylogenetic relationship among different habitat populations. CONCLUSION: The wild C. smyrnioides resources are endangered. The wild C. smyrnioides around Nanjing city are better than the others, it could be used for the further selecting of species resources.

Vertical distribution of sediment enzyme activities involved in the cycling of carbon, nitrogen, phosphorus and sulphur in three boreal rural lakes.

Vertical distribution of eleven hydrolytic enzyme activities were investigated with fluorogenic model substrates in boreal rural lake sediments, Lake Uurainen (13 km(2)), Lake Jämijärvi (9 km(2)) and Lake Pyylampi (0.068 km(2)), in Finland. The eleven hydrolytic enzyme activities were high into deep sediment layers indicating potential for turnover of organic matter in the permanently anoxic zones. The activities of beta-glucosidase, sulphatase and N-acetyl-glucosaminidase were similar in the three lakes independent on the ecological status of the lake. Acetate- and butyrate-esterase were more active than the other enzymes in the three lakes. These unspecific esterases had sediment activities in forest Lake Pyylampi close to those reported for boreal coniferous forest soils in Finland. Similar beta-cellobiosidase activities throughout the sediment depths indicated constant depolymerisation potential for cellulose.

Utilization of monocrotophos as phosphorus source by Pseudomonas aeruginosa F10B and Clavibacter michiganense subsp. insidiosum SBL 11.

Monocrotophos (dimethyl (E)-1-methyl-2-(methylcarbamoyl) vinyl phosphate, or MCP), an organophosphorus insecticide, was used as a sole phosphorus source by the microorganisms isolated from the soil. None of the isolates could utilize MCP as a sole source of carbon. Two of the potential microbial isolates, Pseudomonas aeruginosa F10B and Clavibacter michiganense subsp. insidiosum SBL 11, could utilize MCP as a sole source of phosphorus. Pseudomonas aeruginosa F10B showed a lag phase of 4 h, while in the case of C. michiganense subsp. insidiosum SBL 11, it was 8 h when cultured in the presence of MCP. The generation time for both strains was increased in the medium containing MCP. It was 2.15 h for P. aeruginosa F10B in MCP medium as compared with 1.29 h in basal medium, while in case of C. michiganense subsp. insidiosum SBL 11 it was increased to 3.4 h in MCP medium as compared with 1.28 h in basal medium. These two strains were able to degrade technical MCP in shake-flask culture up to 98.9 and 86.9%, respectively, and pure MCP up to 79 and 80%, respectively, within 24 h at 37 degrees C. The optimal concentration of MCP required for the normal growth was 500 ppm. In the substrate preference study, Tris-p-nitrophenyl phosphate was the most preferred substrate followed by paraoxon. The enzyme responsible for the break down of MCP was phosphotriesterase, which was localized on the membrane-bound fraction of the disrupted cells. The gene responsible for the production of phosphotriesterase (opd) in P. aeruginosa F10B was plasmid-borne.

[Isozymes variations among the germplasm resources of Houttuynia in Sichuan].

The peroxidase and esterase isozymes of the germplasm resources of Houttuynia in Sichuan were analyzed. The results showed that it existed plentiful isozymes variations among the germplasm resources of Houttuynia in Sichuan. The cultivated populations had the same isozymes band types as the wild ones. There were 6 different types of peroxidase isozymes bands and 10 different esterase isozymes band totally. Each peroxidase band type was composed of 4 to 6 bands, and esterase isozymes had 5 to 8 bands. There was no significant difference between Houttuynia cordata and Houttuynia emeiensis populations in their isozymes bands types. The results also showed that the number of peroxidase isozymes bands in northern areas was somewhat higher than that in southern areas. However, esterase isozymes band types had no geographic differences.

[Study on the electrophoresis finger-prints of three groups of traditional Chinese drugs].

OBJECTIVE: To identify three groups of traditional Chinese drugs by electrophoresis finger-prints. METHODS: The 3 groups of drugs of different species or varieties were analyzed by protein, peroxidase(POD) isozyme and esterase (ES) isozyme polyacrylamide gel electrophoresis, respectively. And their finger-prints were established. RESULTS: Not less than one kind of electrophoresis finger-prints has significant difference between the drugs of each group, which can be used to distinguish all the species in this group. CONCLUSIONS: Different species even different varieties of traditional Chinese drugs can be identified accurately with proper electrophoresis finger-prints.

[Somatic hybridization between carrot and Bupleurum scorzonerifolium and esteras isoenzyme analysis].

OBJECTIVE: To transfer the effective elements of Bupleurum scorzonerifolium into carrot, and provide theoretical data for the exploitation, improvement and selection of the germplasm of Chinese medicinal plants. METHOD: The protoplasta of Bupleurum scorzonerifolium irradiated by ultraviolet light (UV) at an intensity of 300 microW.(cm2)-1 for 0, 1, 2 min respectively were fused with those of carrot Fisch by PEG method. The regenerated clones, derived form a single fused cell, were examined for their hybrid nature by phenotype and Esterase isoenzyme analysis. RESULT: Nine clones were identified as the somatic hybrids between B. scorzonerifolium and carrot. CONCLUSION: This provides a firm foundation for the further analysis of the main active components saikosaponin of somatic hybrids and the screening out of high-medicine-content hybrid cell lines.

Human salivary tea catechin levels and catechin esterase activities: implication in human cancer prevention studies.

Because of the possible application of tea in the prevention of oral and esophageal cancers, the salivary levels of tea catechins were determined in six human volunteers after drinking tea. Saliva samples were collected after thoroughly rinsing the mouth with water. After drinking green tea preparations equivalent to two to three cups of tea, peak saliva levels of (-)-epigallocatechin (EGC; 11.7-43.9 microg/ml), EGC-3-gallate (EGCG; 4.8-22 microg/ml), and (-)-epicatechin (EC; 1.8-7.5 microg/ml) were observed after a few minutes. These levels were 2 orders of magnitude higher than those in the plasma. The elimination half-life (t(1/2)) of the salivary catechins was 10-20 min, much shorter than that of the plasma. Holding a tea solution in the mouth for a few minutes without swallowing produced even higher salivary catechin levels, but taking tea solids in capsules resulted in no detectable salivary catechin level. Holding an EGCG solution in the mouth resulted in EGCG and EGC in the saliva and, subsequently, EGC in the urine. The results suggest that EGCG was converted to EGC in the oral cavity, and both catechins were absorbed through the oral mucosa. A catechin esterase activity that converts EGCG to EGC was found in the saliva. The enzyme was likely of human origin, but the activity was not inhibited by common human esterase inhibitor. The present results suggest that slowly drinking tea is a very effective way of delivering rather high concentrations of catechins to the oral cavity and then the esophagus.

[Isozyme electrophoresis of Fritillaria puqiensis during bulb organogenesis in vitro].

OBJECTIVE: Inquiring into the mechanism of bulb organogenesis of Fritillaria puqiensis. METHOD: Time course studies of the soluble protein, peroxidase and esterase were performed during the new bulb organogenesis by electrophoresis. RESULT: The atlas of electrophoresis changed greatly during the phase of differentiation and the formation of the new bulb. CONCLUSION: Different enzymes are associated with organogenesis and express activity at different times.

[Studies on gel electrophoresis of soluble protein and two kinds of isodynamic enzyme in hongqu from different producing areas].

OBJECTIVE: To investigate the consanguinity between 12 strains of Monascus(MS 01-12) separated and purified from different samples of Hongqu collected from 12 areas in China and the variant strain of Monascus(MS 18) mutated from M. purpureus. METHOD: Gel electrophoresis of soluble protein, esterase and superoxide dismutase(SOD) was used to study the consanguinity between strains MS 01-12 and the variant strain MS 18. RESULTS: The electrochromatophoreses of soluble protein, esterase and SOD in the 12 strains (MS 01-12) are identical, but in the variant strain MS 18 are remarkably different. The results are consistent with those of morphological studies. CONCLUSION: Strains MS 01-12 are the identical species M. purpureus, but strain MS 18 is different from strains of MS 01-12, or a variant of M. purpureus.

[Enzynology of infected part of red skin ginseng].

By means of polyacrylaminde gel eletrothoresis and thin layer scanning, the paper semi-quantitatively studies some relation between component and activity of isoenayme of Ginseng infecting red skin desease. The results show activity of esterase increases 100% than that of normal Ginseng; activity of polyphenol oxidase increases 60-30% and represent new isoenzyme bands in the early infection in juxtapose transolant experiment.

[Esterase isoenzyme of various types in rhizoma Gastrodiae].

By means of polyacrylamide gel slab electrophoresis, the esterase isoenzyme of various types in Gastrodia elata in different growing periods was studied and the relativity and difference in physiology and biochemistry were discussed. The result reveals that apparent variety of Rhizoma Gastrodiae is not decided by genetic factors.

Species identification of cyst and root-knot nematodes from potato by electrophoresis of individual females.

To carry out rapid and reliable species identification of Globodera and Meloidogyne specimens, two electrophoretic methods were modified and adapted for an automated electrophoresis system (PhastSystem, Pharmacia). Proteins of individual Globodera cysts were identified using isoelectric focusing and a sensitive silver stain. Proteins of young single Meloidogyne females were separated using polyacrylamide gradient gels and stained for the isozymes esterase and malate dehydrogenase.

Cytotoxic drug testing in head and neck cancer by multiparametric flow cytometry.

Individual chemosensitivity assessment of solid tumors has been hindered by the lack of a reliable and rapid test system for predicting inhibition of tumor cell growth in vitro of all malignant cells and for distinguishing between inflammatory and tumor cells. Six long-term cultured and recloned squamous cell carcinoma lines of the head and neck were analyzed using a newly developed multiparametric flow cytometric technique. After incubation with cytotoxic drugs at different time intervals, each cell line showed its own chemosensitive profile. With this multiparametric staining technique, it is possible to monitor the in vitro response of tumor cells and to compare their quantitative behavior with that of nonmalignant inflammatory cells in solid tumors of the head and neck.

Characteristics of acid esterase in Wolman's disease.

The characteristics of acid esterase from the patient with Wolman's disease, a rare familial lipidosis, were studied. Enzymatic analysis as well as mineral analysis were performed on the patient's liver, spleen, and adrenal glands. Acid esterase was low in the patient's leucocytes and other affected tissues. Further enzymatic study with subcellular fractions of the liver in both patient and control subject revealed that acid esterase was mostly localized in the membrane of lysosomes. The lysosomal esterase was unaffected by Ca2+, Mg2+, EDTA, E600 (microsomal esterase inhibitor), and it was less inhibited by NaCl than other fractions. Studies with those inhibitors showed that acid esterase has different properties compared to other lipases, such as lipoprotein lipase, adipose tissue lipase, and hepatic microsomal lipase. Studies with inhibitors also gave a negative view on a possible suppressive interaction of the high content of calcium in the target organs with acid esterase in Wolman's disease.

Bile salt-stimulated lipase and esterase activity in human milk after collection, storage, and heating: nutritional implications.

In a study of human milk obtained in the first month of lactation, lipase and esterase activity were assayed. Bile salt-stimulated lipase (BSSL) and bile salt-stimulated esterase (BSSE) activities in colostrum were similar to corresponding enzyme activities in transitional milk and in mature milk. BSSL and BSSE were significantly (P less than 0.001) correlated to one another, which suggests that lipase and esterase activities in milk are due to the same enzyme. When milk was allowed to stand at room temperature, in a refrigerator, or subjected to freezing and thawing, wide fluctuations were observed in lipase and esterase activities, but there was no systematic tendency for enzyme activity to increase or decrease. Heating milk to various temperatures between 40-55 degrees C resulted in progressive loss of enzyme activity. The activation energy for the process which inactivates the enzyme was found by linear regression to the Arrhenius plot to be 2 X 10(5) J X mole-1. Our findings suggest that lipase and esterase activity in human milk which is donated to hospitals and stored frozen can make a valuable contribution to fat digestion in the newborn infant, but pasteurization destroys the enzyme.