Novel method facilitates quantification of active pharmaceutical ingredients

Researchers deploy mass spectrometry imaging to identify and quantify compounds present in drugs, cosmetics and foods quickly, cheaply and without toxic waste.

By Karina Toledo  |  Agência FAPESP – A quick and inexpensive method of identifying and quantifying active substances in pharmaceuticals, cosmetics and foods is described by researchers at the University of Campinas (UNICAMP) in São Paulo State, Brazil, in an article recently published in Scientific Reports, an online journal published by Springer Nature.

Developed with FAPESP’s support, the technique is based on mass spectrometry imaging (MSI).

According to the authors, one of its key advantages is its versatility. The same platform can be deployed without adaptation to analyze a diverse array of substances, including biological fluids such as blood. Moreover, because it does not use large amounts of solvent or other chemicals, the method does not produce potentially toxic waste.

“There are many possible applications, both in the pharmaceutical, cosmetics and food industries and in the field of clinical and toxicological analysis,” said Rodrigo Ramos Catharino, a professor at UNICAMP’s School of Pharmaceutical Sciences (FCF) and head of UNICAMP’s Innovare Biomarker Laboratory.

The method was developed as part of Maico de Menezes’s Master’s research and Diogo Noin de Oliveira’s PhD research, conducted with a scholarship from FAPESP. Both were supervised by Catharino under the aegis of the Thematic Project “Mitochondrial energy metabolism, redox state and functionality in cell death and cardiometabolic and neurodegenerative disorders”, for which the principal investigator is Aníbal Eugênio Vercesi, also a professor at UNICAMP.

“One of the Thematic Project’s aims is to study the effects of drugs used to treat cardiometabolic disorders, and to do this we needed a practical deformulation method to find out what substances are present in these drugs and in what proportions,” Catharino said. “In the article, we describe how we validated this novel method by analyzing rosuvastatin, the drug most widely used to control cholesterol.”

Steps

The rosuvastatin used in the study had exactly the same composition as the drug sold in tablet form by pharmacies. It was dissolved in a mixture of water and methanol and applied to filter paper previously prepared with a solution of α-cyano-4-hydroxycinnamic acid (CHCA), which acted as a revealing substance.

“When the filter paper is exposed to a laser, this revealing substance absorbs the light and converts it to heat energy. The CHCA solution is heated rapidly until it sublimates and transfers a proton to the analyte – the compound to be deformulated, in this case rosuvastatin,” Catharino explained. “The resulting ions can be analyzed by the mass spectrometer.”

Each laser shot also generates a pixel, so that eventually a 1 cm² image is formed. The concentration of each substance can be determined based on the intensity of the color produced by the pixels.

Using this MSI technique, known as MALDI, the researchers were able to identify and determine the proportions of ions related to the active ingredient and the other substances present in the sample.

To validate the new technique, they compared the results of their MALDI-MSI analysis with a separate analysis of the same sample using high-performance liquid chromatography (HPLC), currently considered the gold standard for analyzing rosuvastatin. The main problem with HPLC is that it entails the use of large amounts of solvents.

“We achieved the same result in about a tenth of the time and at half the cost or less, depending on the compound involved,” Catharino said.

Applications

As noted by the authors in the article, the techniques used to deformulate drugs have many applications in the pharmaceutical and cosmetics industries, ranging from quality control (to reduce impurities) and product development to compliance with regulatory requirements.

To produce a generic or similar drug, for example, a pharmaceutical company must determine the concentrations of both the active pharmaceutical ingredient (API) and the excipients present in the reference drug.

According to Catharino, pharmaceutical companies currently combine different methods to achieve this goal, such as liquid chromatography, magnetic resonance, infrared analysis, and thermal gravimetric analysis, in which changes in physical and chemical properties are measured as a function of increasing temperature.

“This new method achieves the same result much more quickly and cheaply, since the main input is paper,” Catharino said. “The method can be used to deformulate anything. You could use it to measure the blood level of a specific amino acid or toxin, for example. It could be used in anti-doping tests and in a plethora of other fields.”

The article “Capillary-induced homogenization of matrix in paper: a powerful approach for the quantification of active pharmaceutical ingredients using mass spectrometry imaging” (doi: 10.1038/srep29970) can be read at ncbi.nlm.nih.gov/pmc/articles/PMC4954946.