Original article formulation and characterization of rifampicinloaded p3hbco4hb nanoparticles amirah m. Experiment eight assay test for rifampicin capsules. Materials and methods instruments lab india double beam uv visible spectrophotometer uv 3092 with 1 cm matched quartz cells were used for all absorbance measurements with uv win software. The inhibitory effect of rifampicin on the amyloid formation of hen egg white lysozyme was assessed with both thioflavin t tht fluorescence assay and fourier transform infrared ftir difference spectroscopy. Full text biodegradable rifampicinreleasing coating of. The results showed that the addition of pva and pvp significantly improved drugloading, encapsulation efficiency and release characteristics. A new approach to the use of uv spectrophotometry was used for pyrazinamide pza, rifampicin rif and isoniazid inh assay in fixeddose. Rifampicin and isoniazid were kindly donated by a pharmaceutical industry and used without further purification. Rifampicin in urine was detected by a simple plate diffusion assay using as the test organism a strain of staphylococcus aureus made resistant to other antituberculosis drugs. Rp hplc method development and validation for estimation of isoniazid and rifampicin in tablet dosage form dissertation submitted to the tamilnadu dr. Uv spectrophotometric methods which are stabilityindicating were developed and validated for the determination of rifampicin in bulk drug, capsules and spiked human urine. Liquid chromatography mass spectroscopy, plasma, rifampicin rifampicin 34methyl piperazinyliminomethyl rifamycin is a firstline semisynthetic agent along with isoniazid and pyrazinamide in the treatment of pulmonary and extrapulmonary tuberculosis 1, 2. The mechanism of rifampicin inhibition of escherichia coli rna polymerase was studied with a newly developed steady state assay for rna chain initiation and by analysis of the products formed with several 5terminal nucleotides.
Mycobacterium tuberculosis ribosomal protein s1 rpsa and. The absorption or reflectance in the visible range directly affects the perceived color of the. In addition, the liver tissue of male mice treated with rifampicin showed a decrease. Simultaneous determination of isoniazid and rifampicin by.
Stabilityindicating uvspectrophotometric assay of rifampicin. The assay results of samples 16 list of figures figure 1. Rphplc,7,20 hptlc8 and uv spectrophotometry9 and for the determination of piperine and rifampicin in individually submission date. The degree of resistance varies depending on the site of mutation in the rna polymerase. Rifampicinloaded mesoporous silica nanoparticles for the. Obviously, the presence of rifampicin interfereswitht assay. Simultaneous estimation of rifampicin and isoniazid in. Apr 28, 2017 on the other hand, simple spectrophotometric methods are available to assay rifampicin alone tella et al. In the first method fcr method, inh is reacted with folinciocalteu reagent in na 2 co 3 medium and the resulting blue colored chromogen measured at 760 nm.
The assay is based on the ultraviolet uv absorbance maxima at about 202 wavelength of gentamycin using water as solvent. Rifampicin is active against grampositive bacteria but less active against gramnegative bacteria. Samples were withdrawn at predetermined time points and centrifuged at 38,000 rcf for 10 min. Biodegradable rifampicinreleasing coating of surgical meshes. This means it uses light in the visible and adjacent ranges. Evaluation of the meltpro tbstr assay for rapid detection. Highly luminescent and biocompatible p and n codoped. Uvspectrophotometric methods which are stabilityindicating were developed and validated for the determination of rifampicin in bulk drug, capsules and spiked human urine. Development and validation of high performance liquid. Two methods for the assay of rifampicin in serum are described. Ironii, formed as a result of reaction between inh and ironiii, is made to react with ferricyanide, and the. The assay results for the capsule by proposed method were.
College of pharmacy, madurai 625107, in partial fulfilment of the university rules and regulations for award of master of. It is an aggressive disease with one third of the worlds population reportedly being infected with mtb. Simultaneous spectrophotometric determination of rifampicin. Analysis procedure identification of rifampicin standard, isoniazid standard and pyrazinamide standard by fourier transform. Interpretation of the ir spectra of ethanol, acetone and. Common calculation for determination of the assay result chromatography. Increases in rifampicin concentration increased the fluorescence quenching of the imbedded bsaau ncs in the waxprinted paper sensor.
If the light is in the visible region, meaning the wavelength of 400700 nm or more broadly 360900 nm, the color of the assay can be visibly captured by naked eyes. The stabilityindicating assay is a method that is employed for the analysis of stability of samples in pharmaceutical industry. Assessment of the inhibitory effect of rifampicin on amyloid. There is a paucity of data on application of this test directly to clinical specimens, although based on a. Constant monitoring of rifampicin is important for controlling the side effects and preventing overdose caused by chronic medication. Jun 26, 2015 the intensity of the colorimetric assay performed on the paperbased platforms can be easily captured using a digital camera and subsequently analyzed. Three antituberculosis medications are investigated in this work consist of rifampicin, isoniazid and pyrazinamide. The antimycobacterial activity of the isolated piperine was tested against m. Rifampicin is used to treat tuberculosis and tuberculosisrelated mycobacterial infections. The chromatographic conditions were the same described on the usp xxii under rifampin assay conditions. The purity of the compound was ascertained by uv, ftir studies and by differential scanning calorimetry dsc. The major effect of rifampicin was found to be a total block of the translocation step that would ordinarily follow. It is more common in developing countries, with more than half of the reported cases occurring in asia. Ion mass spectrometry was monitored in positive ion that has a mass 823.
We can divide spectroscopy into two broad classes of techniques. The cell deathbased assay can be used to determine the involvement of a gene in pti. Most tests use the uv visible uv vis spectroscopy as the detection method, which usually falls into the wavelength range of 1001100 nm. Determination of proteinunbound, active rifampicin in. An integral part of the strategy to fight the disease is use of quality antitb drugs. Assay of formulations and % recovery data sample labelled amount mg amount found mg in method % recovery proposed method proposed reported rifampicin capsule tablet 150 150 148.
Each colorimetric assay was performed using a total volume of 30 l prepared bsaau ncs, and the evaluation was performed with a total of 30 l spiked urine containing different concentrations of rifampicin. The first is a conventional plate diffusion method, measuring concentrations down to 002. Assay of rifampicin in bulk and its dosage forms by visible. The decreased fluorescence intensity of bsaau ncs in the presence of rifampicin allows for the sensitive detection of rifampicin in a range from 0. Approximately 50% of multidrugresistant tuberculosis and over 90% of. The polymeric core was synthesized in the following order. Since simple uv spectrophometric method for the assay of rifampicin in combined dosage form is. In absorption spectroscopy a photon is absorbed by an atom or mol. Preparation of microparticles containing rifampicin as dry. Isolation, identification and antimycobacterial evaluation. Original article formulation and characterization of rifampicin loaded p3hbco4hb nanoparticles amirah m. The method, which requires a simple protein precipitation procedure as sample preparation, was performed to compare venous and arterial plasma kinetics after a single therapeutic dose of rifampicin 8. For instance, lossoffunction mutants for a gene of interest can be tested using this assay.
Encapsulation of zincrifampicin complex into transferrinconjugated silver quantumdots improves its antimycobacterial activity and stability and facilitates drug delivery into macrophages. A commercial line probe assay for the rapid detection of. Two spectrophotometric methods are described for the determination of isoniazid inh in pharmaceuticals. Two uv spectrophotometric methods which are stabilityindicating were developed and validated for the determination of rifampicin in bulk drug, capsules and spiked human urine.
Chemometric assisted spectrophotometric methods for the. Rifampicin or rifampin r is a common drug used to treat inactive meningitis, cholestatic pruritus and tuberculosis tb, and it is generally prescribed for longterm administration under regulated dosages. We designed a polymeric micelle to improve the delivery of antitb drugs rifampicin rf and isoniazid inh. Redoxreaction based spectrophotometric assay of isoniazid in. Multiwavelength spectrophotometric determination of. To standardize a highperformance liquid chromatography hplc method for the deter. Research article assessment of the inhibitory effect of. The hplc chromatogram of uv stressed rif, inh and pza. Analysis procedure identification of rifampicin standard, isoniazid. The homogeneity of rifampicin microparticleinhalac 230. Mar 26, 2018 the fluorescence intensity of probe is linearly proportional to rifampicin over the concentration rage from 0 to 20. Besides, this probe can be also used to assay rifampicin in serum samples with satisfactory recoveries.
Development and validation of liquid chromatographymass. Uvspectrophotometry9 for the assay of rif in formulations in. Either test a alone or any two of tests b, c and d may be applied. Validation of a simple hplcuv method for rifampicin.
Simultaneous content analysis of rifampicin, isoniazid and. Ungraded products supplied by tci america are generally suitable for common industrial uses or for research purposes but typically are not. Detection of residual rifampicin in urine via fluorescence. Preparation of calibration curve of metformin 67 12. Uvvis spectroscopy is usually applied to molecules and inorganic ions or complexes in solution. Assay for pathogenassociated molecular pattern pamp. Unbound rifampicin was analyzed using ultra performance liquid chromatography with a beh c18 column, isocratic elution and ultraviolet uv detection. A digimed dm21v6 ph meter was used for the determination of the end point in the titration of inh. Spectra were automatically obtained by uv probe system software.
Each inh and rif can be assayed by ultraviolet spectrophotometry. Uvspectrophotometry with its variants constitutes an evident alternative to the already existing methods. The potential of polymeric micelles constructed by coalescing natural and synthetic polymers for tuberculosis tb treatment was evaluated in this work. A typical example of a straightforward tablet assay is the analysis of furosemide frusemide tablets. Assay of rifampicin in bulk and its dosage forms by visible spectrophotometry using chloranilic acid elisha divakar tella 1. Certificate this is to certify that the dissertation entitled rphplc method development and validation for estimation of isoniazid and rifampicin in tablet dosage form submitted by mr. A validated highperformance liquid chromatography method for the determination of rifampicin and desacetyl rifampicin in plasma and urine a. In this study, we present an easy to use, effective. Assay results of inh and rif determination in pharmaceutical formulations n 6.
Most tests use the uvvisible uvvis spectroscopy as the detection method, which usually falls into the wavelength range of 1001100 nm. An hplcuv method for determining rifampicin concentration in plasma using rifapentine as an internal standard was validated. Encapsulation of zincrifampicin complex into transferrin. Two uvspectrophotometric methods which are stabilityindicating were developed and validated for the determination of rifampicin in bulk drug, capsules and spiked human urine. Interaction of rifampicin and isoniazid with large. A validated highperformance liquid chromatography method for. Assay of rifampicin in serum journal of clinical pathology. Simultaneous determination of rifampicin, isoniazid, pyrazinamide and ethambutol in 4fdc tablet by raman spectroscopy associated to chemometric approach article mar 2017. A shimadzu model 1700 japan double beam uv visible spectrophotometer with spectral width of 2 nm, wavelength accuracy of 0. Tuberculosis tb is one of the most important human bacterial diseases caused by mycobacterium tuberculosis mtb commonly affecting the lungs. Rifampicin is a bactericidal antibiotic drug used to treat inactive meningitis and tuberculosis. Rifampicin was extracted from plasma utilizing liquidliquid extraction process using a 70. It is the measurement of the wavelength and intensity of absorption of near ultraviolet and visible light by a sample.
Simultaneous determination of isoniazid and rifampicin by uv spectrophotometry article pdf available in farmacia 652. The blend of microparticles with inhalac 230 was prepared in ratio of 1. Assay of rifampicin in bulk and its dosage forms by. On the mechanism of rifampicin inhibition of rna synthesis. An hplc uv method for determining rifampicin concentration in plasma using rifapentine as an internal standard was validated. The surface topology of the developed beads were analyzed by scanning electron microscopy and the crosslinking process was investigated by. Line probe assay has been shown to be highly sensitive and specific in the detection of rifampicinresistant tb when used on culture isolates.
Pdf simultaneous determination of isoniazid and rifampicin. The supernatant was diluted in methanol and quantified for rifampicin by uvvis spectroscopy as described above. Medical university, chennai in partial fulfilment for the award of the degree of master of pharmacy in pharmaceutical analysis department of pharmaceutical analysis k. The deficiency in delivery of proper dose of rifampicin has serious implications as it is known that doses of rifampicin. Chemical structure of rifampicin which absorbance of an aqueous solution. Determination binary mixture of isoniazide and rifampicin. Wahab 1school of pharmaceutical sciences, universiti sains malaysia, 11800 penang, malaysia. Biodegradable rifampicinreleasing coating of surgical. Negative control reagent was used as notemplate control. In one class of techniques there is a transfer of energy between the photon and the sample. Pdf one of the most effective antituberculosis treatments is the. Ultraviolet and visible light are energetic enough to promote outer electrons to higher energy levels. Common pharmacopeial calculations in usp monographs.
The mechanism of rifampicin inhibition of esche richia coli rna polymerase was studied with a newly developed steady state assay for rna chain initiation and by analysis of the products formed with several 5 terminal nucleotides. Ultraviolet visible spectroscopy or ultraviolet visible spectrophotometry uv vis or uv vis refers to absorption spectroscopy or reflectance spectroscopy in part of the ultraviolet and the full, adjacent visible spectral regions. Zincalginate beads for the controlled release of rifampicin. The fluorescence intensity of probe is linearly proportional to rifampicin over the concentration rage from 0 to 20. The correction for the absorption of rifampicin at the emission wavelength trivial process or reabsorption was carried out by. Versatile phresponsive chitosangpolycaprolactonemaleic. We reveal that tht fluorescence assay gives a false positive result due to rifampicin interference, while ftir difference spectroscopy provides a reliable assessment. Sigmaaldrich offers a number of rifampicin products. A sample of drug was dissolved in water to produce a solution containing. On the other hand, simple spectrophotometric methods are available to assay rifampicin alone tella et al. Packaging 1, 5, 25 g in glass bottle 250 mg in glass insert. Biochemical characterization of the mycobacterium smegmatis.
Out of the 4 combinations of inducer and challenge inoculations given in table 1, it is possible that only one combination may result in a phenotype in your plant. Spectral and absorbance measurements were made on shimadzu double beam spectrophotometer uv140 with matched 1cm quartz cells and ph measurements were carried out using on systronics ph meter 335. Five milligrams of rifampicinloaded msnp were suspended in 10 ml of pbs sink conditions and agitated at 37 c. Evaluation of the meltpro tbstr assay for rapid detection of. Sep 26, 2017 rifampicin assay spectrophotometry fc reagent ferricyanide abstract of article is published in zhurnal prikladnoi spektroskopii, vol. Pdf simultaneous determination of isoniazid and rifampicin by uv. Uv spectra of rifampicin 200 gml and isoniazide 100gml in methanol. Preparation of calibration curve of ciprofloxacin 64 10. Assessment of the inhibitory effect of rifampicin on. It interferes with the synthesis of nucleic acids by inhibiting dna dependent rna polymerase. Two novel ultraviolet spectrophotometric method for determination. Priyanshu and madhav, 20 but in case of combine dosage form the reported assay methods are quite complex khamar and patel, 2012b.
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