Development On The Monascus Pigment Reference Material

The Monascus pigment is a natural microbial fermentation pigment. It is widely used in food coloring and medicine because for its advantages no toxic side effects, thermal stability, prominent medicinal value and so on So far, there is neither purification technology researches of the monascus, nor certified reference materiel available in the mark.This is found on the study of the technique of extraction, separation and purification, analysis, stability.1. Extraction:Used Soxhlet extraction to study pigment extraction rates of different solvents including n-hexane, ethyl acetate, methanol and methylene chloride. The best extraction solvent was n-hexanes-EtoAc(1:1).2. Purification: reversed-phase were used to isolate and purify then preparative liquid chromatography, and optimized Monascus pigment standards process flow. 220mm (L)×25mm (id) glass column with packed Silicycle silica gel 60, hexane dissolve the sample,9:1 hexane:ethyl acetate was used to wash the impurity,5:1 n-hexane:ethyl acetate was eluted with orange components, and then separated and purified in the OBDTM-Prep C18 column (150mm×19mm× 5μm) by preparative liquid chromatography. The isolated components were recrystallized to obtain monascorubrin which molecular weight is 382 and rubropunctatin which molecular weight is 354.The yield were 0.037%,0.067%. The yield of preparation was stability and good reproducibility.3. Analysis:Using HPLC-MS to detect the mass spectrogram of three pigments and analyze the data of purity; By the analysis of 1HNMR detection data and comparing with literature data to determine the material structure; Two orange pigments in 300-500 nm have absorption peak by spectrophotometry detection, the maximum wavelength is 382 nm; Ultra high performance liquid chromatography (UPLC) with X Bridge TM C18 column (4.6 mm×150 mm×5 microns) and 70% methanol water in the maximum absorption wavelength of 382 nm wavelength to detect. Used the peak area normalization method to calculate the purity of monascus standard product were 100%. While setting up 370 nm,354 nm,320 nm,410nm,465nm wavelength to detect at the same time, found no other impurity peaks. Experiments finally studied out a set of standard by HPLC-MS, spectrometry, nuclear magnetic resonance (NMR),. 4. Stability:After solutions were kept for 30 days in the dark, measures the content of monascorubrin and rubropunctatin were stable,while them was unstable under natural light. Experimental study on the monascorubrin absorbance changes in brown glass bottles protected from light, aluminum foil and dark, single-wavelength ultraviolet light, natural light exposure. It showed that the visible light was faster than the single-wavelength UV decomposition, while brown glass the dark was better than dark effect of the aluminum foil, we initially identified storage conditions. Farther stability remains to be further studied.This is study established the monascus pigment standard product process, provided it standard analysis method, and the stability test. Which for development of the monascus standard provided reference data.