Muhammad Khalid Saeed

Chromatographic fingerprint analysis of Cephalotaxus sinensis from various sources by high-performance liquid chromatography-diodearray detection-electrospray ionization-tandem mass spectrometry

Selective and efficient analytical methods are required not only for quality assurance but also for authentication of Chinese herbal medicine. A simple, rapid and valid fingerprint method has been first carried out for the quality control of Cephalotaxus sinensis by using high-performance liquid chromatography (HPLC) coupled with photodiode array detection (DAD) and electrospray ionisation-tandem mass spectrometry (ESI-MS). The characteristic analytical fingerprints of this plant extract showed 18 common peaks, and out of these, 10 compounds involving 2 new compounds were identified by comparing the retention time, UV and ESI-MS/MS spectrum of each standard with those of each peak separated by on-line HPLC-DAD-MS/MS. Moreover, the effects of collecting locations, harvesting time, storage time, drying methods, and medicinal portions on herbal chromatographic fingerprints were examined by similarity analysis and principal component analysis (PCA) along with markers. Using the reference fingerprint along with markers, the best harvesting time, cultivation location and medicinal part were determined. The results obtained suggest that the chromatographic fingerprint combining similarity evaluation and PCA along with markers or pharmacologically active constituents can efficiently identify raw herb of Cephalotaxus sinensis from different sources, which provide helpful clues to the study of plants secondary metabolites and benefit quality control.

Thermal Characterization of Purified Glucose Oxidase from A Newly Isolated Aspergillus Niger UAF-1

An intracellular glucose oxidase was isolated from the mycelium extract of a locally isolated strain of Aspergillus niger UAF-1. The enzyme was purified to a yield of 28.43% and specific activity of 135 U mg−1 through ammonium sulfate precipitation, anion exchange and gel filtration chromatography. The enzyme showed high affinity for D-glucose with a Km value of 2.56 mM. The enzyme exhibited optimum catalytic activity at pH 5.5. Temperature optimum for glucose oxidase, catalyzed D-glucose oxidation was 40°C. The enzyme showed a high thermostability having a half-life 30 min, enthalpy of denaturation 99.66 kJ mol−1 and free energy of denaturation 103.63 kJ mol−1. These characteristics suggest the use of glucose oxidase from Aspergillus niger UAF-1 as an analytical reagent and in the design of biosensors for clinical, biochemical and diagnostic assays.

Radical scavenging activity of various fractions of Cephalotaxus sinensis (Rehd et Wile) Li

Cephalotaxus sinensis (Rehd et Wile) Li, belongs to family Cephalotaxaceae, common in China, is an indigenous medicinal plant due to its antiascariasis, antiinflammation, anticough and antidyspepsia activity. The overindulgence of free radicals has been concerned in the development of chronic diseases and play an imperative role in the development of neurodegenerative disorders, such as Alzheimers and Parkinsons diseases. Various fractions of this plant were evaluated for their radical scavenging activity by the DPPHldr and reducing power activity assays. Also, the Folin-Ciocalteu method was used to determine the total phenolic content. The results of both methods revealed that Cephalotaxus. sinensis possesses potent antioxidant activity. The polyphenolic content and antioxidant activities of the different fractions tested decreased in the order of EaF > BtF > CfF > WtF > PtF. The radical scavenging activity of the ethyl acetate fraction higher than BHT and slightly lower than Vit-C. A positive linear correlation between antioxidant activity and contents of polyphenol was established. This information shows that ethyl acetate fraction of C. sinensis can be used as antioxidant in medicinal preparations.

Optimization of Condition for Maximum Recovery of Antioxidants from Cephalotaxus sinensis and their FRAP Assay

In this study, Cephalotaxus sinensis (Rehd et Wile) Li (Cephalotaxaeeae), plant was examined for phenolic constituents and antioxidant capacity, to determine their potential as a source of naturat antioxidants. Total polyphenol content (TPC) were determined spectrophotometrically, after extraction of ptant material with assorted solvents at 80degC for 3 hour. Antioxidant activity of aqueous extracts and fractions were evaluated by FRAP assay. The highest antioxidant activity was observed in the extract of 80% aq. ethanol followed by fractionation with 40% (v/v) aq. ethanoi using AB-8 resin, which had soaring leveis of potyphenot. Resutts strongty suggested that potyphenot content shoufd be considered as an imperative feature of Cephalotaxus sinensis ptant and demonstrate a impending for exptoit as natural antioxidants.

LC-MS Analysis of Markers for Advanced Glycation Endprocducts -Protein/Peptides Adducts

Advanced glycation end products (AGEs) are a heterogeneous group of proteins that have been modified with glucose or carbohydrates adducts, and are thought to be responsible for many complications of diabetes and aging. Free amino groups of proteins react slowly with reducing sugars such as glucose by the glycation or Maillard reaction, initially forming Schiff bases, which undergo rearrangement to form the relatively stable Amadori products. The Amadori products subsequently degrade into a-dicarbonyl compounds that react with amino groups of proteins to form cross-links, stable end products called advanced glycation end products (AGEs). AGEs have been shown to react (cross-link) with the amino group of the N-terminal amino acid residue and the side-chains of arginine and lysine residues. Therefore, glucose has been shown to be responsible, at least in part, for protein cross-linkage, oxidative stress and cytotoxicity. In this in vitro study model, the cross-linking of glucose (0.05, 0.5, 1.0 and 2.5 M) with bovine serum albumin incubated for different intervals of time (0, 7,14, 21, 28 and 35 days) was studied using LC-MS and Mascot database to search for the possible markers for glucose-peptides adducts. A total of twenty-seven peptides were matched in the database search query for control BSA out of which seven (m/z 922.69, 464.36, 582.41, 681.95, 834.49, 847.59 and 812.87) were found to be the possible markers for AGEs-protein/peptides adducts.