In order to improve the stationary phase retention of polar solvent systems and aqueous two-phase systems (ATPSs), we designed a multiple spiral disk assembly for type-J high-speed counter-current chromatography (HSCCC). The stationary phase retention was studied under different elution modes by using two solvent systems that contained 1-butanol-acetic acid-water (4:1:5, V/V/V) and polyethylene glycol (PEG) 1000-K2HPO4-water (12.5:12.5:75, W/W/W). The best retention was obtained in L-I-T, U-O-H, L-I-H three modes by pumping lower mobile phase from inner terminal (I) to outer terminal (O), and upper mobile phase from outer terminal (O) to inner terminal (I) at a relatively high flow rate. Meanwhile, the relationship between retention percentage of the stationary phase (Sf) and various parameters such as flow-rate (F), rotation speed (w) and column temperature (T) was also studied. Sf increased with the increase of w and decreased with the increase of F. Regression analysis showed a linear relationship between Sf and F1/2/w. The influence of T on Sf was not obvious between 20 degrees C and 40 degrees C, lower temperature than 20 degrees C was not suitable for viscous ATPSs. Acceptable resolutions were achieved when it was applied for the separation of dipeptides including Leu-Tyr and Val-Tyr by using 1-butanol-acetic acid-water (4:1:5, V/V/V) solvent system. The proteins including cytochrome C and myoglobin, lysozyme and myoglobin, and fresh chicken egg-white proteins were well separated by 12.5% PEG1000-12.5% K2HPO4-75% water (pH 9.0) and 16% PEG 1000-12.5% K2HPO4-71.5% water (pH 8.0) system.
Countercurrent Distribution ; instrumentation ; methods ; Peptides ; isolation & purification ; Proteins ; isolation & purification

Enramycin is a polypeptide antibiotic and new, safe animal feed additive. A new purification process was developed, based on pre-purification by macroporous resin and refining by reversed phase chromatography. AB-8 macroporous resin was used for the pre-purification process of enramycin, with an elution buffer of 0.012 mol/L aqueous HCl solution-methanol (50: 50, V/V). Then, enramycin a and enramycin b were separated effectively by C18 reversed phase chromatography, with a elution buffer of 0.05 mol/L aqueous KH2PO4 solution-acetonitrile (70: 30, V/V, pH 4.5). The purities of enramycin a and enramycin b were up to 98.5% and 98.0%, respectively. The yield reached 29.2%. This study would provide a useful reference for the preparation of enramycin a and enramycin b with a high purity.
Adsorption ; Anti-Bacterial Agents ; isolation & purification ; Chromatography, Reverse-Phase ; methods ; Peptides ; isolation & purification

OBJECTIVETo screen human stem cell factor (hSCF) mimetic peptides in vitro with a phage-display random peptide library.

METHODSPhage clones with high hSCF receptor (rc-kit/Ig 1-3)-binding activity was screened from phage-displayed random hepta/dodecapeptide library by phage enzyme-linked immunosorbent assay (ELISA). Phage single DNA was extracted and sequenced. Four kinds of peptide with higher c-Kit/Ig 1-3 binding activity were chosen for synthesis and characterized by using cell proliferation assay with 3-(4,5-dimethylthiazol-2-yl)2,5-diphenyltetrazolium bromide (MTT) method in UT-7 cells.

RESULTSEleven Ph.D.-C7C clones and eight Ph.D-12 phage clones with high hSCF receptor-binding activity were selected from phage-displayed random hepta/dodecapeptide library, respectively. Sequence analysis showed there were no homologous sequence between hSCF and these screened mimetic peptides except one homologous sequence DPSPHTH found in heptapeptide library. All these four synthesized peptides (CE3, CE16, LE4, and LE20), particularly CE16 and LE20, stimulated UT-7 cell proliferation.

CONCLUSIONFour hSCF mimetic peptides were successfully isolated from phage-displayed random peptide library..

Humans ; Peptide Library ; Peptides ; genetics ; isolation & purification ; Stem Cell Factor ; genetics ; isolation & purification

OBJECTIVETo separate and identify cyclopeptides of tubers of Rubia schumanniana.

METHODThe 70% methanol extracts from tubers of Rubia schumanniana were separated and purified by silica gel, RP-18, Sephedax LH-20 and HPLC. Their structures were identified by spectral analysis.

RESULTNine cyclopeptides were separated and identified as RA- II (1), RA-V (2), RA-VIII (3), rubiyunnanin C (4), RA-X (5), RY-II (6), RA- I (7), RA-XIII (8) and RA-XIII-OMe (9), respectively.

CONCLUSIONAll of nine cyclopeptides were separated from R. schumanniana for the first time.

Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Molecular Structure ; Peptides, Cyclic ; analysis ; isolation & purification ; Rubia ; chemistry

OBJECTIVETo identify the tyrosinase inhibitory constituent quickly from Potentilla bifurca.

METHODThe active constituent was found through fraction collecting and tyrosinase inhibitory activity by bioassay-linked HPLC method.

RESULTThe methanol extracts and BuOH fraction of Potentilla bifurca showed strong tyrosinase inhibitory activities. From BuOH fraction of Potentilla bifurca, the tyrosinase inhibitory constituent was isolated and identified as flavonoid, quercetin-4'-O-beta-D-glucoside. It express stronger tyrosinase inhibition than the known tyrosinase inhibitor, kojic acid (IC50 = 0.28 mmol x L(-1)) with IC50 value of 0.001 9 mmol x L(-1).

CONCLUSIONBioassay-linked HPLC fractionation method was provided for determination the active constituents quickly from herbal medicines.

Enzyme Inhibitors ; chemistry ; isolation & purification ; Kinetics ; Peptides ; chemistry ; isolation & purification ; Plant Extracts ; chemistry ; isolation & purification ; Potentilla ; chemistry

OBJECTIVETo study the chemical constituents from the mycelia of Flammulina velutipes.

METHODThe compounds were isolated with silica gel column chromatography and their structures were elucidated on the basis of spectral analysis (IR, EL-MS, FAB-MS, 1H-NMR, 13NMR).

RESULTSeven compounds were identified as cyclo-(R-pro-R-leu) (1), cyclo-(R-isoleu-R-leu) (2), phenylalanine (3), alanine (4), leucine (5), guanosine (6), adenosine (7),

CONCLUSIONThe compounds 1-6 were isolated from the mycelia of Flammulina velutipes for the first time.

Agaricales ; chemistry ; Alanine ; chemistry ; isolation & purification ; Dipeptides ; chemistry ; isolation & purification ; Fermentation ; Leucine ; chemistry ; isolation & purification ; Mycelium ; chemistry ; Peptides, Cyclic ; chemistry ; isolation & purification ; Phenylalanine ; chemistry ; isolation & purification

As the technologies of separation, purification and determination develop rapidly, more and more peptide compounds, which have special bioactive and medical value, have been separated from natural plants, such as oligopeptides and cyclopeptides. The chemical structures and function of these plant peptides have been researched profoundly. This paper mainly reviews the composition, structure, bioactive function and medicine value of representative plant peptides in recent years, and can give some references about research and application of plant bioactive peptides.
Animals ; Antihypertensive Agents ; chemistry ; isolation & purification ; pharmacology ; Antineoplastic Agents, Phytogenic ; chemistry ; isolation & purification ; pharmacology ; Antioxidants ; chemistry ; isolation & purification ; pharmacology ; Glutathione ; chemistry ; isolation & purification ; pharmacology ; Gramicidin ; chemistry ; isolation & purification ; pharmacology ; Humans ; Hypoglycemic Agents ; chemistry ; isolation & purification ; pharmacology ; Oligopeptides ; chemistry ; isolation & purification ; pharmacology ; Peptides ; chemistry ; isolation & purification ; pharmacology ; Peptides, Cyclic ; chemistry ; isolation & purification ; pharmacology ; Plants ; chemistry ; Plants, Medicinal ; chemistry

A fraction named GFC-1 with high antioxidant activities in vitro was isolated from the enzymatic hydrolysates of roasted pills of Asini Corii Colla, and the peptides in this fraction were identified. The enzymatic hydrolysates were isolated and purified with anion exchange chromatography and Sephadex G-25 filtration chromatography successively. GFC-1, a fraction isolated from the hydrolysates, exhibited the highest DPPH and ABTS scavenging capacity (DPPH 47. 95% at 2.0 g x L(-1) and ABTS 97.20% at 0.40 g x L(-1). Nine peptides from GFC-1 were identified by LC-ESI-MS/MS coupled with TurboSEQUEST search software and Swiss-Prot data base, and a high repetition core sequence GPAGPP*GPP* was also found.
Animals ; Antioxidants ; chemistry ; isolation & purification ; Equidae ; Hydrolysis ; Mass Spectrometry ; Peptides ; chemistry ; isolation & purification ; Protein Hydrolysates ; chemistry ; Skin ; chemistry

OBJECTIVE[corrected] To study the cyclic peptides from Psammosilene tunicoides.

METHODThe constituents were separated and purified with chromatographic methods, identified by NMR, MS, UV and IR.

RESULTThree cyclic dipeptides: cyclo(-Pro-Val-) (1), cyclo(-Pro-Ala-) (2) and cyclo(-Pro-Pro-) (3), were isolated and identified.

CONCLUSIONCompound 1 and 2 are new natural products, compound 3 was isolated for the first time from Psammosilene tunicoides.

Caryophyllaceae ; chemistry ; Dipeptides ; chemistry ; isolation & purification ; Molecular Structure ; Peptides, Cyclic ; chemistry ; isolation & purification ; Plant Roots ; chemistry ; Plants, Medicinal ; chemistry

To develop a timesaving and easy operating Reverse Phase (RP) chromatography method, we adopted Thermo Pierce RP C18 Tip to separate small amount hippocampus peptide mixtures and to compare with high performance liquid chromatography (HPLC). According to the separation performance of 4 ACN gradient optimization methods, we determined the best ACN concentration gradient. The results showed that, the experiment took only 10 min by separating with eight ACN concentration gradient, which accounted 1/4 for HPLC. But as for the identified proteins, RP C18 Tip accounted 85.5% for HPLC. ACN gradient of 5%, 15%, 20% and 90% had best repeatability (P = 0.429) and result for separating 30 μg peptides. This method is easy to operate, timesaving and has low cost. It could be used into pretreatment of small amount complex proteomic samples.
Chromatography, High Pressure Liquid ; Chromatography, Reverse-Phase ; methods ; Peptides ; Proteins ; isolation & purification ; Proteomics ; methods