Preparation Of Porous Titania By Water-regulated Morphology Of MIL-125(Ti) For CPPs Separation

As excellent chromatographic stationary phase,TiO₂ chromatographic stationary phase has been widely used in separation and analysis of biological and food samples.In order to improve its small specific surface area and pore diameter,some researchers have used MIL-125?Ti?,a kind of metal organic skeleton?MOFs?,to prepare porous TiO₂ chromatographic stationary phase with large specific surface.Appling TiO₂ chromatographic stationary phase to the separation and enrichment of phosphatide,the effect is good.However,in the process of adjusting the morphology of MIL-125?Ti?,impurities are easy to occur and block the pore structure of TiO₂chromatographic stationary phase,which will affect the separation and enrichment.Therefore,in this paper,TiO₂ chromatographic stationary phase was prepared by water-regulated MOFs and core-shell metal-organic framework?MOF@MOF?,respectively.TiO₂ chromatographic stationary phase was used for separation and analysis of theophylline.And it was also packed into solid phase extraction?SPE?column for separating and enriching casein phosphopeptide?CPPs?.This paper consists of the following four parts:?1?The morphology of precursor,MIL-125?Ti?was controlled by adding water,then calcined MIL-125?Ti?to prepare TiO₂ chromatographic stationary phase.To optimize the shape,particle size and specific surface of TiO₂ chromatographic stationary phase,we investigated the effect of volume ratios of DMF to MeOH,addition of water,molar ratios of Ti to H₂BDC,and conditions of calcination.And the MIL-125?Ti?and the TiO₂ chromatographic stationary phase were characterized by optical microscope,scanning electron microscope?SEM?,X-ray diffraction?XRD?,energy disperse spectroscopy?EDS?and other instruments.The synthesized TiO₂chromatographic stationary phase derived from that the volume ratio of DMF to MeOH is 9:1,the addition of water is 30?L,molar ratio of Ti to H₂BDC is 1:9 and the condition of calcination is from room temperture to 380°C at a speed of 5°C/min and kept at 380°C for 5 h in air directly.Under this condition,the specific surface area of the TiO₂ chromatographic stationary phase with porous structure and anatase crystal was 118.41 m²/g and particle size was about 3.5?m,which met the requirements for the application of HPLC stationary phase and SPE adsorbent.?2?MIL-125?Ti?@MIL-125?Ti?was prepared by epitaxial growth method.After calcining,the TiO₂ composite stationary phase with larger particle size and adjustable porous structure could be obtained.The effects of the addition of MIL-125?Ti?nuclear material,the dilution ratio of shell solution,the reaction time and the number of coating layers on the morphology and structure of MIL-125?Ti?@MIL-125?Ti?were investigated.And the MIL-125?Ti?@MIL-125?Ti?and the TiO₂ composite stationary phase under optimal conditions were characterized by SEM,specific surface area and aperture analyzer,Fourier infrared spectroscopy,XRD and EDS.According to the results,the optimal preparation condition of MIL-125?Ti?@MIL-125?Ti?is adding 30mg MIL-125?Ti?nuclear material,without diluting shell solution,cover with 1 layer,and react for 24 h.The specific surface area of the TiO₂ composite stationary phase was 76.00 m²/g.The thickness was about 2.5?m.And the particle size was about 7?m,which was larger than the TiO₂ chromatographic stationary phase prepared by the existing method.And the shape of it was more similar to the sphere,which was conducive to expanding the application of the TiO₂ in the separation and analysis of substances.?3?The TiO₂ chromatographic stationary phase based on MIL-125?Ti?was used as the stationary phase of HPLC,to prepare the TiO₂ normal phase chromatographic column.With column efficiency of the homemade column,retention time and half peak width of theophylline as a parameter,the effects of volume fraction of methanol,flow rate,temperature,pH value of acetate buffer and concentration of acetate buffer on theophylline retention were studied.After optimizing the conditions of HPLC,the theophylline content in the three tablets was determined.It is concluded the optimal chromatographic conditions of separating and analysing theophylline:the volume fraction of acetate buffer solution with pH value 6 and concentration 8 mmol/L was35%,volume fraction of MeOH was 65%,temperature was 45?,flow rate was 0.8mL/min,and detection wavelength was 273 nm.The contents of theophylline in three kinds of tablets were 463.40 mg/g,605.00 mg/g and 681.26 mg/g,respectively.?4?We focused on the enrichment of CPPs,using SPE column filled with homemade TiO₂ chromatographic stationary phase.With the number of titanium hydroxyl groups as the parameter,the activation conditions including the concentration of hydrochloric acid?HCl?,ultrasonic time,ultrasonic temperature and oscillation time were investigated;With the adsorption rate and desorption rate of CPPs on SPE column under dynamic conditions as parameters,the extraction conditions were optimized.The optimized conditions were used to isolate and concentrate the CPPs in the sample,and the sample solution,effluent and eluent were analyzed and identified by flight tandem mass spectrometry?Q-TOF LC/MS?.Then,HPLC and internal standard method were used to determine the CPPs content in the sample.Optimized activation conditions as follow.The concentration of HCl was 3mol/L;ultrasonic time was 30 min;ultrasonic temperature was 45?and oscillation time was 10 h.Optimized extraction conditions as follow.pH value of sample solution was 3;flow rate of SPE was 0.2 mL/min and the concentration of ammonia was 0.3 mol/L.Two phosphopeptides were identified using Q-TOF LC/MS in the sample,which were PS*PR and S*SRS*E,respectively.The total content of CPPs in the sample was 15.413 mg/g.According to all above results,it indicated that this method had better separation and concentration effect on CPPs.