OBJECTIVETo study the anticoagulant constituents in dried leech (Whitmania pigra Whitman).

METHODThe plasma recalcification time (PRT) as index, the constituents with anticoagulant activity were isolated and purified by anion-exchange chromatography on Sephadex DEAE A-50, gel permeation chromatography on Sephadex G-25 and Sephadex LH-20 columns, and then reversed phase high-performance liquid chromatography successively.

RESULTThree anticoagulant polypeptides were isolated and purified. Compounds 1 and 2 can be translated each other in natural conditions, and their molecular weights are 7100 and 5531, respectively. Compound 3 was identified as a pure polypeptide by HPLC and SDS-PAGE, and its molecular weight was determined as 8 608 by MALDI-TOF-MS. The amino acid composition of compound 3 was also determined.

CONCLUSIONCompound 3 was inferred to be a novel anticoagulant, and named whitmanin.

Animals ; Anticoagulants ; analysis ; chemistry ; isolation & purification ; Leeches ; chemistry ; Molecular Weight

OBJECTIVEApplying for the activity of enzyme in vitro,the research optimized the best preparation procedure for the anticoagulated blood region from Lumbricus.

METHODAll through our experiment, the content of protein and theactivity of enzyme were examined. The extraction process, the refining technology, concentration processes of Lumbricus were optimized with single factor checking and orthogonal design method.

RESULTAt 37 degrees C, the coarse powder of Lumbricus soaking with 15 fold of 0.9% sodium chloride and ultrasonic extracting 40 minites for three times was the best ultrasonic extraction. Utrafiltration membrane with molecular weights of 30 x 10(3) for refining and 10 x 10(3) for concentrating were selected.

CONCLUSIONUltrasonic extraction and membrane separation technology, to well improve the effect of purification for the anticoagulant site of Lumbricus, is conducive to further study.

Animals ; Anticoagulants ; chemistry ; isolation & purification ; Drug Compounding ; methods ; Oligochaeta ; chemistry ; enzymology ; Temperature ; Ultracentrifugation ; Ultrasonics

Anti-Bacterial Agents ; therapeutic use ; Anticoagulants ; therapeutic use ; Arcanobacterium ; isolation & purification ; Humans ; Lemierre Syndrome ; drug therapy ; microbiology ; Male ; Middle Aged

Mosquitoes secrete saliva that contains biological substances, including anticoagulants that counteract a host's hemostatic response and prevent blood clotting during blood feeding. This study aimed to detect heparin, an anticoagulant in Aedes togoi using an immunohistochemical detection method, in the salivary canal, salivary gland, and midgut of male and female mosquitoes. Comparisons showed that female mosquitoes contained higher concentrations of heparin than male mosquitoes. On average, the level of heparin was higher in blood-fed female mosquitoes than in non-blood-fed female mosquitoes. Heparin concentrations were higher in the midgut than in the salivary gland. This indicates presence of heparin in tissues of A. togoi.
Aedes/*metabolism ; Animals ; Anticoagulants/*isolation & purification ; Blood Coagulation/physiology ; Female ; Gastrointestinal Tract/*metabolism ; Heparin/*isolation & purification ; Male ; Salivary Ducts/metabolism ; Salivary Glands/*metabolism

BACKGROUNDSnake venom contains a number of components with different pharmacological and biological activities, especially in cancer therapy, and has increasingly become a research focus. This study was designed to isolate and purify a novel anti-clotting protein component from the venom of Agkistrodon acutus, and to explore its physico-chemical properties and biological activity.

METHODSThe venom of Agkistrodon was isolated and purified by ion-exchange chromatography on diethylaminoethyl (DEAE)-Sepharose Fast Flow, molecular sieve filtration through Sephadex G75, SP-Sepharose Fast Flow and molecular sieve filtration through Sephadex G50. We detected the activated partial thromboplastin time (APTT) of the eluant to select the anti-clotting protein component of interest. The molecular weight was determined by sodium dodecyl sulfate-polyacrylamid gel electrphoresis (SDS-PAGE) and liquid chromatography. Its protein content was detected by bicinchoninic acid (BCA).

RESULTSSDS-PAGE vertical gel electrophoresis showed that the anticoagulant factor is a tripolymer composed of three proteins whose molecular weights are 25 KDa, 30 KDa and 50 KDa. The factor contains about 65% percent protein.

CONCLUSIONSA novel anti-clotting protein component was purified by ion-exchange chromatography and molecular sieve filtration from the venom of Agkistrodon acutus and was found to be composed of three kinds of proteins.

Agkistrodon ; metabolism ; Animals ; Anticoagulants ; chemistry ; isolation & purification ; Chromatography, High Pressure Liquid ; Chromatography, Ion Exchange ; Crotalid Venoms ; chemistry ; Electrophoresis, Polyacrylamide Gel ; Proteins ; chemistry ; isolation & purification

Family Schisandraceae is composed of two genera Schisandra and Kadsura, which are quite different in regard to the used part and medical function. Traditionally, the fruits of Schisandra plants were called 'Wuweizi' used as tonic for neurasthenia, antitussive, and sedative agents. The stems and roots of Kadsura plants, with names related to 'XueTeng', possess activating blood circulation and eliminating stasis and are used for the treatment of rheumatism, fractures and irregular menstruation. The dried fruit of Schisandra chinensis was recorded in Chinese Pharmacopoeia (2010 Edition) as 'Wuweizi', while the dried fruit of S. sphenanthera was recorded as an official origin of 'Nanwuweizi'. Historically, there was no evidence that the fruits of Kadsura plants were substituted as 'Wuweizi' either in ancient literatures or contemporary marketing. However, genus Kadsura is still popularly called 'Nanwuweizi' and plant K. longipedunculata is regarded as the origin of 'Nanwuweizi', thus this will cause confusion as well as misunderstanding of genus Kadsura. The authors recommended, therefore, the Chinese name of genus Kadsura should be 'Lengfantengshu' and K. longipedunculata with the name of 'Lengfanteng' in order to guarantee the drug authenticity.
Anticoagulants ; isolation & purification ; Antitussive Agents ; isolation & purification ; Drug Contamination ; Drugs, Chinese Herbal ; isolation & purification ; Fruit ; chemistry ; Kadsura ; chemistry ; Plant Roots ; chemistry ; Plant Stems ; chemistry ; Plants, Medicinal ; classification ; Schisandra ; classification ; Terminology as Topic

OBJECTIVETo separate anticoagulant components from the pepsin enzymolysis of centipede by gel filtration and reverse-phase C18 chromatography, and to detect the distribution range of their molecular mass.

METHODCingula and 280 nm ultraviolet spectrometry were used to detect and collect the chromatographic solutions. The components' anticoagulant activity in vitro was detected with activated partial thromboplastin time (APTT) as the index, and the molecular mass range of the active composition was detected by MALDI-TOF-MS.

RESULTAnticoagulant active compounds were produced by gel filtration and reverse-phase C18 chromatography. The distribution range of relative molecular mass was determined to be from 597 to 1 146.

CONCLUSIONGel filtration and reverse-phase C18 chromatography are feasible for separating and purifying the pepsin enzymolysis of Centipede. The anticoagulant active compounds are oligopeptides.

Animals ; Anticoagulants ; isolation & purification ; metabolism ; Arthropods ; chemistry ; enzymology ; Chromatography, High Pressure Liquid ; Male ; Partial Thromboplastin Time ; Pepsin A ; chemistry ; metabolism ; Peptides ; isolation & purification ; metabolism ; Rabbits ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

SHG was sulfated by chlorosulfonic acid-pyridine method, and six samples which we got were prepared in different reaction conditions. There is a characteristic absorption peak near 260 nm in UV spectra and there are two characteristic absorption peaks near 1240 cm(-1) and 810 cm(-1) in the FT-IR. Degree of sulfation (DS) was calculated by elemental analysis and turbidimetry. Under the same conditions the absorption peaks become strong with the DS increase. The anticoagulant activity of SHG and sulfated modification samples was evaluated by the classic coagulant assays of prothrombin time (PT), activated partial thrombin time (APTT) live enzymes, and plasma thrombin time (TT). Results show that sulfated SHG has a good anticoagulant activity in vitro, and DS increased activity within a certain range.
Animals ; Anticoagulants ; chemistry ; isolation & purification ; pharmacology ; Blood Coagulation Tests ; Fabaceae ; chemistry ; Glucans ; chemistry ; isolation & purification ; pharmacology ; Partial Thromboplastin Time ; Plants, Medicinal ; chemistry ; Prothrombin Time ; Rabbits ; Spectrophotometry, Ultraviolet ; Spectroscopy, Fourier Transform Infrared ; Sulfonic Acids ; chemistry ; Thrombin Time

OBJECTIVETo investigate the effects of polysaccharide 2-1 from Gastrodia elata (PGE2-1) on blood coagulation and thrombosis.

METHODClotting time (CT) and bleeding time (BT) of mice were measured by glass method and tail-cutting method. Bleeding capacity (A540) was measured by cutting tail in 5 min. Plama recalcificatic time (RT) were measured in mice. Platelet aggregation was caused by adenosine diphosphate (ADP). Activated partial thromboplastin time (APTT), prothrombin time (PT), and thrombin time (TT) were measured by reagent boxes. During thrombosis in vitro, their lengths, wet and dry weights were measured by instrument; wet weights of arteriovenous experimental thrombosis were measured and the impressive rates were analyzed.

RESULTCT and BT of groups PGE2-1 (60, 120 mg x kg(-1)) were remarkably prolonged, and bleeding capacity (A540) were significantly increased (P < 0.05 or P < 0.01). RT of groups PGE2-1 (30, 60, 120 mg x kg(-1)) were remarkably prolonged, and platelet aggregation (PAG) were inhibited (P < 0.05 or P < 0.01). Human serous TT and APTT of groups PGE2-1 (10, 20, 40 mg x mL(-1)) were remarkably prolonged (P < 0.05 or P < 0.01), but the difference of effect on PT had no statistic significance. PGE2-1 (30, 60, 120 mg x kg(-1)) could make the mice obviously eliminate thrombus symptom and reduce the time of restoring independent activity (P < 0.05 or P < 0.01); thrombosis in vitro: Lengths, wet and dry weights of groups PGE2-1 (30, 60, 120 mg x kg(-1)) were significantly decreased (P < 0.05 or P < 0.01); wet weights of arteriovenous experimental thrombosis were dramatically decreased (P < 0.01), and impressive rates were respectively 32.5%, 49.0% and 61.5%.

CONCLUSIONPGE2-1 has remarkable effects of anticoagulation and antithrombosis, so it may be the main component of the isolation from G. elata in the field of antithrombosis.

Animals ; Anticoagulants ; isolation & purification ; pharmacology ; Blood Coagulation Tests ; Dose-Response Relationship, Drug ; Female ; Fibrinolytic Agents ; isolation & purification ; pharmacology ; Gastrodia ; chemistry ; Glycoproteins ; isolation & purification ; pharmacology ; Humans ; Male ; Mice ; Partial Thromboplastin Time ; Phytotherapy ; Plants, Medicinal ; chemistry ; Platelet Aggregation ; drug effects ; Prothrombin Time ; Random Allocation ; Rats ; Rats, Wistar ; Thrombin Time ; Thrombosis ; pathology ; prevention & control

Thrombophlebitis of the portal venous system (PVS) with superimposed bacterial infection (septic pylephlebitis) is an extremely rare complication of Crohn's disease (CD), and therefore diagnosis of septic pylephlebitis is difficult without high clinical suspicion. A 16-year old male patient who was diagnosed with CD 3 months earlier was admitted with recurrent fever and abdominal pain. CD activity had been well controlled with conventional medical treatment during a follow-up period. Abdominal contrast-enhanced computed tomography showed massive thrombosis in the PVS without evidence of intra-abdominal infection, and blood cultures were positive for Streptococcus viridians. There was no evidence of deep vein thrombosis or pulmonary thromboembolism, and all laboratory tests for thrombophilia were normal. Based on these findings, the patient was diagnosed with septic pylephlebitis complicated with CD, and was successfully treated with intravenous antibiotic therapy combined with anticoagulation. This case suggests that early comprehensive evaluation is crucial for immediate diagnosis and proper treatment of septic pylephlebitis in patients with CD who present with fever and abdominal pain of unknown origin, even with stable disease activity and absence of other intra-abdominal infections.
Adolescent ; Anti-Bacterial Agents/therapeutic use ; Anticoagulants/therapeutic use ; Colonoscopy ; Crohn Disease/complications/*diagnosis ; Humans ; Male ; Phlebitis/complications/*diagnosis ; Portal Vein/radiography ; Sepsis/*diagnosis/microbiology ; Streptococcal Infections/diagnosis/drug therapy ; Thrombosis/drug therapy/radiography ; Tomography, X-Ray Computed ; Viridans Streptococci/isolation & purification