We studied the effects of several medicinal insects on biosynthesis of polysaccharides from Ganoderma lucidum in submerged culture. The results showed that the medicinal insect, Catharsius molossus at 5 g/L significantly promoted the biosynthesis of intracellular polysaccharides (IPS) and extracellular polysaccharides (EPS) of G. lucidum, and compared with control, IPS and EPS yields markedly enhanced from (1.93 +/- 0.09) g/L to (2.41 +/- 0.12) g/L and (520.3 +/- 20.2) mg/L to (608.9 +/- 20.2) mg/L, respectively (P < 0.05). Both IPS and EPS consisted of five kinds of components, and IPS-1 and EPS-1 were the major components of IPS and EPS, respectively. Further separation studies showed that IPS-1 was made up of three single compounds, while EPS-1 was made up of two single compounds. There were no new components in both IPS and EPS obtained from G lucidum in submerged culture by the addition of the insect, C. molossus, suggesting the biosynthetic pathways of the major components of IPS and EPS had not been changed.
Animals ; Cockroaches ; chemistry ; Culture Media ; chemistry ; Fermentation ; Materia Medica ; pharmacology ; Polysaccharides ; biosynthesis ; chemistry ; Reishi ; growth & development ; metabolism

The contamination of aflatoxin B_1,B_2,G_1,G_2,M_1 and M_2 in Eupolyphaga Steleophaga was determined by immunoaffinity column clean-up and HPLC-FLD with post-column photochemical derivatization. Chromatographic separations were carried out using a Cloversil C_(18) column( 4. 6 mm×250 mm,5 μm) that were eluted in isocratic with methanol-acetonitrile-water( 20 ∶ 20 ∶ 60) as the mobile phase. The excitation wavelength and the emission wavelength of fluorescence detector were maintained at 360 nm and 450 nm,respectively. The flow rate was 0. 8 m L·min~(-1),and the column temperature was 30 ℃ ． The sample was prepared using the immunoaffinity column,then the recovery was measured with 75. 47%-101. 8% with RSD values lower than 6. 7%. A total of 20 batches of Eupolyphaga Steleophaga samples were assayed. According to the Chinese Pharmacopoeia( 2015 edition,part 1),the aflatoxin B_1 limit should be less than 5 μg·kg~(-1),and the sum of aflatoxins( AFB_1,AFB2,AFG_1,AFG_2) should be less than 10 μg·kg-1. Therefore,the positive rate of the 20 samples was 50. 0%,and 7 batches of samples exceeded the standard,and the over-standard rate was as high as 70. 0%. Among them,aflatoxins B_1,B_2,G_1,G_2,M_1,and M_2 were detected in three batches( SD-1,AH-1,AH-3),and aflatoxins B_1,B2,G1,G2,and M1 were detected in one batch( AH-7). The results showed that the newly developed method in this work is suitable for the simultaneous determination of six aflatoxins in Eupolyphaga Steleophaga,and also suggested that it should be of high values to take the contamination with aflatoxins into concerns.
Aflatoxins/analysis* ; Animals ; Chromatography, Affinity ; Chromatography, High Pressure Liquid ; Cockroaches/chemistry*

OBJECTIVEChinese allergic subjects have high levels of sensitization to house dust mite (HDM) and other indoor allergens. This study quantifies common indoor allergen levels in Chinese households.

METHODSDust samples were collected from nine cities. Major allergens Der p 1 and Der f 1 from Dermatophagoides pteronyssinus and D. farinae, and specific antigens of Blomia tropicalis, Tyrophagus putrescentiae, Acarus siro, and cockroach species Blattella germanica and Periplaneta americana were measured by ELISA.

RESULTSHDM allergens were found in dust samples from bedding in 95% of the Chinese households. The median levels varied from <0.006 to 9.2 µg/g of dust, depending on the city. The percentages of households having HDM allergen levels associated with the risk of developing allergy sensitization and asthma were 65% and 25%, respectively. Specific antigens of the storage mite and cockroach were only found in samples from the southern and tropical regions of China. Levels of mite allergens were generally higher in samples from bedding compared to samples from the living room, even for storage mites, whereas levels of cockroach antigens were higher in the living room samples.

CONCLUSIONHDM allergens are present in bedding dust samples from most Chinese households. Cities in southern and central China have relatively high levels of HDM major allergens compared to cities in northern and western China. Antigens of storage mites and cockroaches are not as common as HDM allergens.

Air Pollution, Indoor ; statistics & numerical data ; Allergens ; chemistry ; Animals ; Bedding and Linens ; China ; Cockroaches ; Dust ; analysis ; Housing ; Pyroglyphidae ; Seasons

It is widely known that the cockroach is an inhalant allergen in atopic asthma and allergic rhinitis. Even though Bla g I and Bla g II are considered as the major allergens, several relatively high-molecular weight (MW) cockroach allergens have also been recently identified by IgE-immunoblot in western countries. However, the environmental control and diagnostic tests mainly focussed on Bla g I and Bla g II. Furthermore there is no data about major IgE-binding cockroach antigens in Korea. We performed this study to identify the major German cockroach allergens in Korean atopic children. By the results of allergy skin tests, 14 children with atopic asthma (9 were cockroach-sensitive and 5 were cockroach-nonsensitive atopics) were enrolled in this study. We conducted IgE immunoblot and autoradiographic analysis using Yonsei-extract of German cockroach antigen produced in our laboratory, individual sera from 9 cockroach- sensitive children, and the pooled sera of 5 house-dust-mites-only-sensitive children. We performed an allergic skin test to cockroach mix, and a radioallergosorbent test (RAST) using German cockroach crude extract on all subjects. German cockroach-specific IgE was detected in 6 out of 9 subjects by RAST. We identified at least 15 IgE-binding protein bands, and among them, the components of MWs of 76, 64, 50, 38, and <14 kilodaltons (kDa) were the major German cockroach allergens in study subjects. Therefore, Bla g I (25-30 kDa) and Bla g II (36 kDa) could not be the absolute indicators of German cockroach sensitization and parameters of environmental control.
Adolescence ; Allergens/analysis ; Animal ; Asthma/metabolism* ; Asthma/immunology ; Asthma/complications ; Child ; Child, Preschool ; Cockroaches/immunology ; Cockroaches/chemistry* ; Female ; Human ; Hypersensitivity/metabolism* ; Hypersensitivity/immunology ; Hypersensitivity/complications ; IgE/metabolism* ; Korea ; Male ; Tissue Extracts/metabolism*

Cockroaches have been implicated as a cause of respiratory allergy in urban areas worldwide. IgE-reactive German cockroach proteins were identified with molecular weights (MWs) of 90, 66, 50, 43 and 36 KD by immunoblot analysis in both immune BALB/c mice and sensitized humans. Prominent IgE-reactive proteins were purified using FPLC by ion-exchange chromatography, gel filtration and hydrophobic chromatography. The N-terminal amino acid sequence of a purified protein with a MW of 66 KD on SDS-PAGE was Val-Thr-Leu-Lys-Lys(Val)-Met-Ile-Lys-Thr-Phe-Tyr. No homologous protein was found through a search of GenBank that indicated a novel IgE-reactive protein in German cockroach extract. Another purified protein with a MW of 36 KD reacted strongly with a monoclonal antibody against Bla g 2.
Amino Acid Sequence/genetics ; Animal ; Cockroaches/chemistry* ; Human ; IgE/immunology* ; Insect Proteins/isolation & purification* ; Insect Proteins/immunology* ; Insect Proteins/genetics ; Mice ; Mice, Inbred BALB C ; Tissue Extracts/chemistry*

Bla g 2 is a cockroach allergen of great importance. This study was conducted to identify IgE-binding epitope(s) of Bla g 2 using the recombinant protein technique. Approximately 50% of tested sera showed IgE reactivity to Pichiaexpressed Bla g 2 (PrBla g 2) and E. coli-expressed Bla g 2 (ErBla g 2). Only 5.3% of serum samples showed stronger reactivity to PrBla g 2 than ErBla g 2, indicating that serum was reactive to conformational or carbohydrate epitopes. The full-length and 5 peptide fragments of Bla g 2 were produced in E. coli. All fragments showed IgE-binding activity to the cockroach-allergy patients' sera. Specifically, peptide fragments of amino acid residue 1-75 and 146-225 appeared to be important for IgE-binding. The information about the IgE-binding epitope of Bla g 2 can aid in the diagnosis and treatment for cockroach allergies.
Adolescent ; Adult ; Aged ; Amino Acid Sequence ; Animals ; Antibodies ; Aspartic Endopeptidases/chemistry/genetics/*immunology ; Child ; Cockroaches/immunology ; Epitopes ; Gene Expression Regulation ; Humans ; Immunoglobulin E/*immunology ; Middle Aged ; Young Adult

OBJECTIVETo obtain the cDNA sequence encoding fibrinolytic enzyme from Eupolyphaga sinensis and express it in prokaryotic and eukaryotic expression system.

METHODThe primers were designed according to the cDNA of other animals'fibrinolytic enzyme. The cDNA sequence was cloned by RT-PCR and 3 RACE.

RESULTSequence analysis revealed that the length of the cDNA fragment was 672 bp and encoded a protein of 224 amino acid residues, the N end amino acid sequence residues was IVGG in accordance with other fibrinolytic enzyme. The cDNA sequence was expressed in E. coli, inactive protein was obtained. While expressed in Pichia pastoris, recombinant protein had fibrinolytic activity.

CONCLUSIONThe cDNA sequence of fibrinolytic enzyme from E. sinensis Walker was cloned and expressed for the first time and it proved a good basis for further functional study of the enzyme.

Amino Acid Sequence ; Animals ; Base Sequence ; Cloning, Molecular ; Cockroaches ; chemistry ; enzymology ; genetics ; DNA, Complementary ; chemistry ; genetics ; Fibrinolysin ; chemistry ; genetics ; metabolism ; Gene Expression ; Insect Proteins ; chemistry ; genetics ; metabolism ; Molecular Sequence Data ; Sequence Alignment

OBJECTIVETo explore the effect of compound decoction on notoginsenosides in Panax notoginseng.

METHODNotoginsenoside R1, Rg1, Re, Rb1 and pH were used as the parameters to investigate the changes on the content of notoginsenosides in different compound extractions by heating for two hours and their correlation with pH.

RESULTWhen the pH values of solution of P. notoginseng with Fructus ligustri, P. notoginseng with Eupolyphaga seu steleophaga, P. notoginseng with Pheretima asiatica, and Zhitangjiang Fang (free of Hirudo) were rept higher than 5.7, the reserved rate (RR) of notoginsenside were higher than 90%; When the pH values of decoetion of P. notoginseng with Salvia miltiorrhiza, P. notoginseng with Paeonia lactiflora, P. notoginseng with Platycodon grandiflorum, P. notoginseng with Arctium lappa were kept 4.5-5.5, their RR of notoginsenside were 60% - 85%; When the pH values of the decotction of P. notoginseng with Hirudo nipponica was decreased to 3.4, its RR of of notoginsenside was 38.4%; When the pH values of Zhitangjiang Fang extraction was regulated by 0.1% NaOH solution to pH 6. 3, and the RR of notoginsenside increased to 97%.

CONCLUSIONThe pH of other Chinese herbal medicines extraction with P. notoginseng compound is a critical effect on the stability and yields of notoginsensides.

Animals ; Arctium ; chemistry ; Cockroaches ; chemistry ; Drug Combinations ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Ginsenosides ; analysis ; Hirudo medicinalis ; chemistry ; Hot Temperature ; Hydrogen-Ion Concentration ; Ligustrum ; chemistry ; Materia Medica ; chemistry ; isolation & purification ; Oligochaeta ; chemistry ; Paeonia ; chemistry ; Panax ; chemistry ; Platycodon ; chemistry ; Salvia miltiorrhiza ; chemistry

Many allergists are currently focusing on the development of new diagnostic tools, and are attempting to improve both the sensitivity and specificity. A multiple allergen simultaneous test-chemiluminescent assay (MAST-CLA) is one of the most popular diagnostic tools used in the Republic of Korea. However, there remains controversy among allergists with regard to the cut-off point for a positive result. The present study was conducted in order to determine the validity of MAST-CLA as compared with that of the skin prick test, with particular emphasis on arthropod allergens, on the basis of percentage agreement rates and k-values, and also to suggest the optimal positive cutoff points using receiver operating characteristic (ROC) curves. The study was conducted with 97 subjects (54 men, 43 women). Optimal individual cut-off points were calculated as follows; class II for Dermatophagoides farinae, class I for Dermatophagoides pteronyssinus, and trace for a cockroach mix. These findings suggest that attempting to apply optimal individual cut-off points will be a good way of improving diagnostic tests, particularly MAST-CLA.
Adult ; Allergens/*immunology ; Animals ; Antigens, Dermatophagoides/*immunology ; Chemiluminescent Measurements/*methods/standards ; Cockroaches/chemistry ; Dermatophagoides farinae/chemistry ; Dermatophagoides pteronyssinus/chemistry ; Female ; Humans ; Hypersensitivity/*diagnosis/immunology ; Insect Proteins/*immunology ; Male ; *ROC Curve ; Skin Tests/methods

We studied the effects of Catharsius molossus (a Chinese medicinal insect) on the cell growth, fermentation kinetics of key bioactive substances and anti-cancer activity of Ganoderma lucidum in submerged fermentation. The results showed that C. molossus at all the tested concentrations had no stimulatory effect on the cell growth. However, addition of C. molossus at 5 g/L lead to significant effects on the fermentation kinetics of polysaccharides and triterpenoids of G lucidum, and at 7th day in fermentation process, the yields of polysaccharides and triterpenoids reached 2.81 g/L and 539.0 mg/L, respectively, while they were 2.25 g/L and 428.2 mg/L in control. In vivo anti-cancer studies showed that the inhibitory rates of control fermented G lucidum (CFG) and a combination of water extract from C. molossus and CFG on the developed tumor (Heps) in mice were 41.61% and 42.24%, respectively. Moreover, the inhibitory rate of the G lucidum fermented with C. molossus (GFC) reached 57.21%, which was enhanced 37.49%, compared to the inhibitory rate of the control fermented G lucidum. These results suggest that supplementation of C. molossus in submerged fermentation of G lucidum lead to a significant enhancement of the anti-cancer activity of cultured G lucidum.
Animals ; Antineoplastic Agents, Phytogenic ; biosynthesis ; therapeutic use ; Cockroaches ; chemistry ; Culture Media ; chemistry ; Female ; Fermentation ; Liver Neoplasms ; drug therapy ; Male ; Materia Medica ; pharmacology ; Mice ; Polysaccharides ; biosynthesis ; pharmacology ; Reishi ; growth & development ; metabolism ; Triterpenes ; metabolism ; pharmacology