Prelab week 1 Calculations Preparation of 1.5μmol/L mixed low-level standard dilution 150μmol/L × V1=1.5μmol/L × 10ml V1=(1.5μmol/L×10ml)/(150μmol/L)=0.1ml Conversion of milliliters to microliters (0.1ml×1000)μL= 100μL Preparation of 3μmol/L mixed low-level standard dilution 150μmol/L × V1=3μmol/L × 10ml V1=(3μmol/L×10ml)/(150μmol/L)=0.2ml Conversion of milliliters to microliters (0.2ml×1000)μL= 200μL Preparation of 3μmol/L mixed low-level standard dilution 150μmol/L × V1=7.5μmol/L × 10ml V1=(7.5μmol/L×10ml)/(150μmol/L)=0.5ml Conversion of milliliters to microliters (0.5ml×1000)μL= 500μL Preparation of the blank samples The volumetric flask will be filled to the mark with 150μmole/L of stock solution to act as blank (reference). Additional two blanks will …show more content…
2010). RP-HPLC is recognized by a condition in which the mobile phase is made to be more polar as compared to the stationary phase. In reversed HPLC, the stationary phase is hydrophobic. It is also chemically bonded to the surface of silica support elements. The reversed HPLC mobile phase for neutral analytes is composed of organic modifier and water. Organic modifiers are used to change the retention time of different analytes. Organic modifiers lower mobile phase polarity. By increasing the amount of water lead to the repulsion of hydrophobic analytes out of the mobile phase. The hydrophobic analytes are pushed onto the stationary phase where they reside for duration up to the partitioning into the mobile phase. When ionic analytes exist in the sample, the addition of ion and buffer to the mobile phase are necessary. This addition aids in controlling the reproducibility and retention. Separation of the mixture via RP-HPLC can be done using continuous gradient or stepwise to move out the sample components. For every separation, the ideal gradient and volume must be