The past decade has seen an increase in the number of research papers on ligand binding to proteins based on fluorescence spectroscopy. In most cases, determination of the binding affinity is made by analyzing the quenching of protein fluorescence induced by the ligand. However, many such articles, even those published in reputed journals, suffer from several mistakes with regard to analysis of fluorescence quenching data. Using the binding of phenylbutazone to human serum albumin as a model, we consider some of these mistakes and show how they affect the values of the association constant. In particular, the failure to correct for the inner filter effect and the use of unsuitable equations are discussed. Ligand binding data presented in these articles should be treated with caution, especially in the absence of data from complementary techniques.
The abuse of phenylbutazone among rheumatoid arthritis patients has recently become a subject of interest. Unscrupulous manufacturers take advantage of the miraculous analgesic property of phenylbutazone and deliberately add this toxic drug in their preparations without declaring its presence on the label. In a recent survey, many such illicit preparations were seized from Chinese medical halls in Johor and sent to the Department of Chemistry, Johor Bahru for analysis. Here a Gas Chromatograph Mass Selective Detector (GC-MSD) method was developed for the determination of phenylbutazone in illicit traditional preparations.
Drug eruptions from indeginous medicine is often difficult to diagnosis and confirm. It is known that a number of these now supplied by bomohs and Chinese sinsehs contain known drugs and are dispensed as tablets and capsules. We report 3 cases of adverse drug eruption to "Jamu", a Malay herb. A particular brand, "Jamu Indonesia, Toko Air Pancur", from Johor Bahru, Malaysia, is especially recommended for "sakit pinggang" or backache. The cases occurred between January and February 1985, and all had taken brown kidney shaped tablets. The adverse reactions were moderately severe. Two had erythroderma with hepatitis, and one, Steven Johnson Syndrome. Analysis of this jamu for analgesics led to the discovery of adulteration with phenylbutazone and diazepam. Records from local cases from 1974-1984 showed that 8 other patients, all Chinese had adverse cutaneous eruptions from phenylbutazone, oxybutazone and propyphenazone. The skin manifestations were erythroderma (2 cases), vasculitis (2 cases) and toxic epidermal necrolysis (4 cases). Those with toxic epidermal necrolysis had 100% mortality.
Pazopanib (PZP) is a multi-targeting tyrosine kinase inhibitor and is currently approved by FDA for the treatment of soft tissue sarcoma and renal cancer. Molecular interaction mechanism of PZP with human serum albumin (HSA) was explored under simulated physiological conditions (pH = 7.4), using fluorescence and UV absorption spectroscopy along with computational methods. Based on the inverse correlation between the Stern-Volmer constant (Ksv) and temperature, it was concluded that PZP quenched the protein fluorescence through static quenching mechanism. This was also confirmed from the UV-vis absorption spectral results. Moderate binding affinity between PZP and HSA was evident from the Ka values (5.51 - 1.05 × 105 M-1) while PZP-HSA complex formation was driven by hydrophobic and van der Waals interactions as well as hydrogen bonds, as revealed by positive entropy change (ΔS = +98.37 J mol-1 K-1) and negative enthalpy change (ΔH = -60.31 kJ mol-1). Three-dimensional fluorescence spectral results disclosed microenvironmental perturbations around Trp and Tyr residues of the protein upon PZP binding. Interestingly, the addition of PZP to HSA significantly protected the protein against thermal stress. Competitive drug displacement results obtained with warfarin, phenylbutazone and diazepam elucidated Sudlow's Site I, positioned in subdomain IIA of HSA, as the preferred binding site of PZP which was well supported by molecular docking analysis, while molecular dynamics simulation results suggested the stability of the PZP-HSA complex.Communicated by Vsevolod Makeev.