Abstract
A new simple, rapid, specific, accurate, precise and novel Reverse Phase High Performance Liquid Chromatography (RP-HPLC) method has been developed for the estimation of Sitagliptin Phosphate in the pharmaceutical dosage form. The chromatographic separation for Sitagliptin was achieved with mobile phase containing methanol, Thermoscientific C18 column, (250x4.6 particle size of 5µ) at room temperature and UV detection at 267 nm. The compounds were eluted in the isocratic mode at a flow rate of 1ml/min. The retention time of Sitagliptin was 2.37min. The above method was validated in terms of linearity, accuracy, precision, LOD, LOQ , interday, intraday , selectivity , range , stability and robustness in accordance with ICH guidelines.
Keywords
Sitagliptin phosphate, Spectrophotometry, HPLC,UV etc.
Introduction
SITAGLIPTIN PHOSPHATE
Diabetes mellitus is a common disease characterized by hyperglycemia, excessive thirst, frequent urination, excessive hunger, general body malaise, and weakness due to perturbed metabolism of fat, carbohydrates, and protein. Cancer and heart disease are killer illnesses that currently afflict the population more rapidly because of their higher prevalence and link to a greater likelihood of morbidity and mortality. The major classes of oral anti-diabetic drugs include sulfonylureas, glinides, biguanides, thiazolidinediones, ?-glucosidase inhibitors, and dipeptidyl peptidase-4 (DPP-4) blockers.[1] The DPP-4 enzyme degrades the incretins glucagon-like peptide 1 (GLP-1), a gastrointestinal hormone secreted in response to eating. To prevent GLP-1 breakdown, sitagliptin phosphate increases insulin discharge by suppressing glucagon release from the ? cells in the pancreas, thereby regulating blood sugar levels.[2]
Drug Profile:
HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY[4]
Another name for high performance liquid chromatography is high pressure liquid chromatography. It is a well-liked analytical method for figuring out, identifying, and quantifying each component of a combination. HPLC is a sophisticated method of liquid chromatography in columns. The early 20th century saw the discovery of liquid chromatography as an analytical approach that was first used to the separation of colored substances. This is the origin of the terms "chromatography," which means "color," and "graphy," which means "writing." A crude method of chromatographic separation was employed by Russian botanist Mikhail S. T-Swett to separate plant pigment mixtures into their individual components. The foundation of every chromatographic separation is the pigments' interaction with a stationary phase, which is how he separated the pigments. In his separation, the solvent served as the mobile phase, and he employed powdered chalk and aluminum as the stationary phase. Because HPLC operates at considerably higher pressures (50 bar to 350 bar), it may be distinguished from classical ("low weight") liquid chromatography. On the other hand, the portable stage in conventional liquid chromatography usually relies on gravity to move it through the segment. Column section measurements range between 2.1 and 4.6 mm in diameter and between 30 and 250 mm in length due to the minimal sample quantity that is isolated in scientific HPLC. Furthermore, smaller sorbent particles (2 ?m to 50 ?m in normal molecule size) are used in the production of HPLC segments. Because of its ability to identify components while separating mixtures, HPLC has a high determining or resolving power, making it a popular chromatographic technique.
HPLC provides advantages such as simultaneous analysis and high resolution.
- Repeatability is good.
- A small sample size.
- Analysis conditions are moderate.
- Easy to fractionate and purify the sample.
Fig. 2: High-performance liquid chromatography[4]
TYPES OF HPLC
HPLC types are typically determined by the phase system employed in the procedure. The following kinds of HPLC are commonly employed in the analysis
- Normal Phase HPLC is a process that separates analytes depending on their polarity. NP-HPLC has a polar stationary phase and a non-polar mobile phase. As a result, the stationary phase is normally silica, while the mobile phases are often hexane, methylene chloride, chloroform, diethyl ether, and combinations of these. As a result, polar samples remain on the polar surface of the column packing for longer than less polar materials.
- Reverse-phase chromatography:
An aqueous, moderately polar mobile phase and a non-polar stationary phase make up reverse phase HPLC, also known as RPC or RP-HPLC. Hydrophobic interactions, which arise from repulsive forces between a polar eluent, the comparatively non-polar analyte, and the non-polar stationary phase, are the basis for RPC's operation. When the analyte molecule associates with the ligand in the aqueous eluent, its binding to the stationary phase is proportional to the contact surface area around the non-polar region of the molecule.
- Ion exchange chromatography:
In this method, solute ions are attracted to charged sites that are attached to the stationary phase, which results in retention. The same-charged ions are not included. Water purification, ligand-exchange chromatography, protein ion-exchange chromatography, high-pH anion-exchange chromatography of carbohydrates and oligosaccharides, and other applications are common uses for this type of chromatography.
- Size exclusion chromatography (SEC):
also referred to as gel filtration or gel permeation chromatography, is a technique used to primarily separate particles according to size. Determining the quaternary and tertiary structures of proteins and amino acids is another beneficial application of it. This method is frequently employed to determine the molecular weight of polysaccharides.
- Bio-affinity chromatography:
A method of separation based on the particular, reversible interactions that ligands and proteins have. In a bio-affinity matrix, ligands are covalently bonded to solid support, retaining proteins that interact with the column-bound ligands. Proteins attached to a bio-affinity column can be extracted in two ways:
- Bio-specific elution involves using a free ligand in the elution solution to compete with column-bound ligands.
- A particular elution is characterized by changes in pH, salt, and other factors that impair the protein's interaction with column-bound substrates.
Because of the selectivity of the contact, affinity chromatography may achieve very high purification rates in a single step (10 - 1000-fold).
Instrumentation of HPLC[5]
- Solvent reservoir and pump
- under high pressure (1000–5000 psi
- An injector
Reduced pressure:
cease the flow
Value at high pressure
d. Desk
- Normal Phase: hydrocarbon-free, organic mobile phase
- Non-aqueous silica gel
- Adsorption
- Aqueous mobile phase in reverse phase (C8, C18)
Dividends
- Water-based ion-exchange mobile phase
- Aqueous mobile phase-molecular sieve
- Dimensions
e. The detector
? Particular
Absorbance
- Fluorescence
- Electrochemical
- Indefinite
- Index of Refraction
Radioactivity
Conductivity
LITERATURE REVIEW:
METHODS OF SITAGLIPTIN PHOSPHATE (ONE-TIME ASPECT):
CONCLUSION:
The documented spectrophotometric and chromatographic techniques that were created and verified for sitagliptin estimation are shown in this review. This review led to the conclusion that, in order to increase resolution, various spectroscopic and chromatographic techniques are available for both single and combination forms of sitagliptin. Additionally, it was discovered that most chromatographic methods used a mobile phase that included phosphate buffer, methanol, and acetonitrile. It was shown that the most typical pairing of sitagliptin and metformin (JANUMET) was prevalent. To obtain a satisfactory retention time for the chromatographic technique, a flow rate of 0.8–1.5 ml/min is observed. Methanol is a popular solvent used in the majority of spectroscopic procedures. As a result, all of the techniques were discovered to be straightforward, precise, affordable, accurate, and repeatable. However, it became evident from this assessment that the Design of Expert (DOE) process might be used to improve the existing methodologies and produce more exact and accurate results.
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10.5281/zenodo.13619846
