{"id":632,"date":"2015-02-15T07:35:02","date_gmt":"2015-02-15T07:35:02","guid":{"rendered":"http:\/\/biomedpharmajournal.org\/?p=632"},"modified":"2020-04-25T07:22:15","modified_gmt":"2020-04-25T07:22:15","slug":"determination-of-azathioprine-and-rocuronium-in-biological-fluid-samples-by-voltammetry","status":"publish","type":"post","link":"https:\/\/biomedpharmajournal.org\/staging\/vol2no1\/determination-of-azathioprine-and-rocuronium-in-biological-fluid-samples-by-voltammetry\/","title":{"rendered":"Determination of Azathioprine and Rocuronium in Biological Fluid Samples by Voltammetry"},"content":{"rendered":"

Introduction<\/strong><\/p>\n

Azathioprine [6-[(1-methyl-4-nitro-1H<\/em>-imidazol-5-yl)sulfanyl]-7H<\/em>-purine] (AZP) \u00a0is a 6-mercaptopurine derivative used as an immunosuppressive agent for prevention of transplant rejection and for the treatment of rheumatoid arthritis and various autoimmune diseases.<\/p>\n

Rocuronium [[3-hydroxy-10,13-dimethyl-2-morpholin-4-yl-16- (1-prop-2-enyl-2,3,4,5-tetrahydropyrrol-1-yl)-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro- 1H<\/em>-cyclopenta[a] phenanthren-17-yl] acetate] (RCR) is a non-depolarizing agent neuromuscular blocking agent used as an adjunct in general anaesthesia and as skeletal muscle relaxant during surgery.<\/p>\n

AZP was determined by 1H NMR spectroscopy (1), spectrophotometric (2) and chromatographic (3- 6) methods. RAN was determined by HPLC (20), GC-MS (22) and LC (23) methods which are expensive and time consuming. In the present work, a cheaper and selective voltammetric determination of AZP and RCR has been reported.<\/p>\n

Experimental<\/strong><\/p>\n

Voltammograms were recorded with Metrohm 757 VA computrace (Herisau, Switzerland).AZP and RCR were purchased from Sigma. Graphite powder (l-2 mm particle size), paraffin oil and Clay from Aldrich India Ltd., Bangalore. All chemicals used for the preparation of buffers and supporting electrolytes are of reagent grade.<\/p>\n

Recommended Procedure<\/strong><\/p>\n

A suitable amount of analyte is transferred into the electrolytic cell containing Britton-Robinson buffer solution. To remove oxygen, the solution is purged with nitrogen gas for 10 min. The voltammograms are recorded after each aliquot of the standard solution is added.<\/p>\n

Results and Discussion<\/strong><\/p>\n

Cyclic Voltammetry<\/strong><\/p>\n

Figs. 1 and 2 illustrate the cyclic voltammograms recorded for AZPand RCR at carbon paste (CPE) and clay modified carbon paste electrode (CMCPE). On scanning towards a negative potential on a bare carbon paste electrode, only a much smaller cathodic peak is observed. When CMCPE is used a large increase in the peak currents is observed. No peaks are observed in the anodic sweep indicating that the reduction of AZP and RCR \u00a0is of irreversible process.<\/p>\n\n\n\n
\u00a0\"Figure<\/td>\nFigure 1: Typical CV of 1.2 x 10-7<\/sup> M AZP\u00a0RCR \u00a0 at (a) bare CPE (b) CMCPE.<\/strong><\/p>\n

 <\/p>\n

Click here to View figure<\/a><\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

 <\/p>\n\n\n\n
\u00a0\"Figure<\/td>\nFigure 2: Typical CV of 3.5xl0-6<\/sup> M\u00a0at (a) bare CPE; (b) CMCPE.<\/strong><\/p>\n

 <\/p>\n

Click here to View figure<\/a><\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

Differential Pulse Voltammetry<\/strong><\/p>\n

Figs. 3 and 4 illustrate differential pulse voltammograms obtained at bare carbon paste electrode and clay modified carbon paste electrodes of AZP and RCR in BR buffer. From the obtained results the peak current obtained at clay modified carbon paste electrode are almost twice than those at carbon paste electrode of AZP and RCR.<\/p>\n\n\n\n
\u00a0\"Figure<\/a><\/td>\nFigure 3:Typical DPAdSV of 1.4 x 10-9<\/sup> M AZP\u00a0 at (a) bare CPE (b) CMCPE.<\/strong><\/p>\n

 <\/p>\n

Click here to View figure<\/a><\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

 <\/p>\n\n\n\n
\u00a0\"Figure<\/td>\nFigure 4: Typical DPAdSV of 1.6×10-9<\/sup> M\u00a0 RCR (a) bare CPE; (b) CMCPE.<\/strong><\/p>\n

 <\/p>\n

Click here to View figure<\/a><\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

 <\/p>\n

This peak in the voltammogram of AZP is attributed to the four electron reduction of nitro group to the corresponding hydroxylamine group according to the currently accepted mechanism for electroreduction of nitro compounds (82-84).\u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0\u00a0<\/strong><\/p>\n\n\n\n
\u00a0\"Scheme<\/td>\nScheme 1: Reduction mechanism of Azathioprine.<\/strong><\/p>\n

 <\/p>\n

Click here to View Scheme<\/a><\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

 <\/p>\n

The peak in the voltammogram\u00a0 of RCR is attributed to the reduction of carbon, carbon double bond according to the currently accepted mechanism for the electroreduction of carbon, carbon double bond containing compounds.(25, 26).<\/p>\n\n\n\n
\u00a0\"Scheme<\/td>\nScheme 2: Reduction mechanism of Rocuronium.<\/strong><\/p>\n

 <\/p>\n

Click here to View Scheme<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

\u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0 \u00a0\u00a0<\/strong><\/p>\n

Table 1: \u00a0Chosen Experimental Conditions.<\/strong><\/p>\n\n\n\n\n\n\n\n\n\n\n\n\n
Variables<\/strong><\/td>\n\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0 Chosen Value<\/strong><\/td>\n<\/tr>\n
\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0 AZP<\/strong><\/td>\n\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0 RCR<\/strong><\/td>\n<\/tr>\n
pH<\/td>\n8.5<\/td>\n3.0<\/td>\n<\/tr>\n
Buffer volume (ml)<\/td>\n10<\/td>\n10<\/td>\n<\/tr>\n
Accumulation potential (V)<\/td>\n– 0.16<\/td>\n-0.4<\/td>\n<\/tr>\n
Accumulation time (s)<\/td>\n160<\/td>\n150<\/td>\n<\/tr>\n
Rest time (s)<\/td>\n15<\/td>\n25<\/td>\n<\/tr>\n
Stirring rate (rpm)<\/td>\n2000<\/td>\n\u00a0\u00a0 2000<\/td>\n<\/tr>\n
Scan rate (mVs-1<\/sup>)<\/td>\n10<\/td>\n10<\/td>\n<\/tr>\n
Pulse amplitude (mV)<\/td>\n50<\/td>\n50<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n


\nTable 2: Experimental data of AZP and RCR<\/strong><\/p>\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n
Parameters<\/strong><\/td>\nAZP<\/strong><\/td>\n\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0 RCR<\/strong><\/td>\n<\/tr>\n
CPE<\/strong><\/td>\nCMCPE<\/strong><\/td>\nCPE<\/strong><\/td>\nCMCPE<\/strong><\/td>\n<\/tr>\n
Linearity range (M)<\/td>\n2.0 \u00b4 10-8<\/sup> to<\/p>\n

3.0 \u00b4 10-7<\/sup><\/td>\n

2.2 \u00b4 10-9<\/sup> to<\/p>\n

1.2 \u00b4 10-5<\/sup><\/td>\n

1.2 \u00b4 10-8<\/sup> to<\/p>\n

1.0 \u00b4 10-5<\/sup><\/td>\n

0.2 \u00b4 10-9<\/sup> to<\/p>\n

1.2 \u00b4 10-5<\/sup><\/td>\n<\/tr>\n

Calibration curve equation<\/td>\nY(mA)=0.30214X +0.03535<\/td>\nY(mA)=0.2985X+ 0.0442<\/td>\nY(mA)=0.9815X +0.1139<\/td>\nY(mA)= 0.9865 X +0.1063<\/td>\n<\/tr>\n
Correlation coefficient<\/td>\n0.9971<\/td>\n0.9995<\/td>\n0.9865<\/td>\n0.9815<\/td>\n<\/tr>\n
L.O.D (M)<\/td>\n1.5 \u00b4 10-8<\/sup><\/td>\n2.1 \u00b4 10-9<\/sup><\/td>\n1.4 \u00b4 10-8<\/sup><\/td>\n0.4 \u00b4 10-9<\/sup><\/td>\n<\/tr>\n
L.O.Q (M)<\/td>\n0.5 \u00b4 10-7<\/sup><\/td>\n0.667 \u00b4 10-8<\/sup><\/td>\n0.466 \u00b4 10-7<\/sup><\/td>\n0.133 \u00b4 10-8<\/sup><\/td>\n<\/tr>\n
Repeatability of<\/p>\n

peak currents<\/p>\n

%RSD)<\/td>\n

4.28<\/td>\n4.86<\/td>\n4.29<\/td>\n4.32<\/td>\n<\/tr>\n
Repeatability of<\/p>\n

Peak potentials<\/p>\n

%RSD)<\/td>\n

0.32<\/td>\n0.38<\/td>\n0.54<\/td>\n0.59<\/td>\n<\/tr>\n
Reproducibility of peak currents<\/p>\n

%RSD)<\/td>\n

3.92<\/td>\n4.02<\/td>\n4.91<\/td>\n4.98<\/td>\n<\/tr>\n
Reproducibility of potentials<\/p>\n

%RSD)<\/td>\n

0.51<\/td>\n0.53<\/td>\n0.32<\/td>\n0.38<\/td>\n<\/tr>\n
Numbers of assays<\/td>\n12<\/td>\n12<\/td>\n12<\/td>\n12<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n


\nTable 3: Determination of AZP and RCR in\u00a0 spiked human serum samples<\/strong><\/p>\n\n\n\n\n\n\n\n\n\n
Name of the drug<\/strong><\/td>\nAmount labelld (m.g\/L)<\/strong><\/td>\n*Average amount found<\/strong><\/p>\n

(m.g\/L)<\/strong><\/td>\n

Recovery percentage<\/strong><\/p>\n

(%)<\/strong><\/td>\n

+<\/u> S.D<\/strong><\/td>\nRSD<\/strong><\/td>\n<\/tr>\n
AZP<\/td>\n2<\/td>\n1.961<\/td>\n98.0<\/td>\n0.0200<\/td>\n1.02<\/td>\n<\/tr>\n
<\/td>\n4<\/td>\n3.93<\/td>\n98.25<\/td>\n0.0461<\/td>\n1.173<\/td>\n<\/tr>\n
<\/td>\n6<\/td>\n5.956<\/td>\n99.26<\/td>\n0.0208<\/td>\n0.349<\/td>\n<\/tr>\n
RCR<\/td>\n4<\/td>\n3.923<\/td>\n98.07<\/td>\n0.0513<\/td>\n1.307<\/td>\n<\/tr>\n
<\/td>\n8<\/td>\n7.93<\/td>\n99.125<\/td>\n0.0655<\/td>\n0.825<\/td>\n<\/tr>\n
<\/td>\n12<\/td>\n11.9167<\/td>\n99.30<\/td>\n0.0611<\/td>\n0.5127<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

* Each value is an average of three determinations.<\/p>\n

 <\/p>\n

Table 4: Determination of \u00a0AZP and RCR in spiked human urine samples<\/strong><\/p>\n\n\n\n\n\n\n\n\n\n
Name of the drug<\/strong><\/td>\nAmount Spiked (m.g\/L)<\/strong><\/td>\n*Average amount found<\/strong><\/p>\n

(m.g\/L)<\/strong><\/td>\n

Recovery percentage<\/strong><\/p>\n

(%)<\/strong><\/td>\n

+<\/u> S.D<\/strong><\/td>\nRSD<\/strong><\/td>\n<\/tr>\n
AZP<\/td>\n2<\/td>\n1.97<\/td>\n98.50<\/td>\n0.004<\/td>\n0.2030<\/td>\n<\/tr>\n
<\/td>\n4<\/td>\n3.94<\/td>\n98.5<\/td>\n0.0624<\/td>\n1.583<\/td>\n<\/tr>\n
<\/td>\n6<\/td>\n5.71<\/td>\n95.31<\/td>\n0.07<\/td>\n1.17<\/td>\n<\/tr>\n
RCR<\/td>\n2<\/td>\n1.98<\/td>\n99.0<\/td>\n0.0206<\/td>\n1.040<\/td>\n<\/tr>\n
<\/td>\n4<\/td>\n3.903<\/td>\n97.57<\/td>\n0.032<\/td>\n0.81<\/td>\n<\/tr>\n
<\/td>\n6<\/td>\n5.72<\/td>\n95.33<\/td>\n0.0670<\/td>\n1.17<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

* Each value is an average of three determinations.<\/p>\n

Conclusion<\/strong><\/p>\n

Clays are complex micro porous media with an appreciable surface area helped in increasing the sensitivity of the proposed method. Due to CMCPE, stability, accuracy and low expensive, it offers a good possibility as a substitute for the previous approaches used in routine analysis, such as colorimetric, spectrophotometric and chromatographic methods. Results obtained in the developed method shows that drugs can be determined accurately and reliably at a lower expens<\/p>\n

References<\/strong><\/p>\n

    \n
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    Introduction Azathioprine [6-[(1-methyl-4-nitro-1H-imidazol-5-yl)sulfanyl]-7H-purine] (AZP) \u00a0is a 6-mercaptopurine derivative used as […]<\/p>\n","protected":false},"author":2,"featured_media":0,"comment_status":"closed","ping_status":"closed","sticky":false,"template":"","format":"standard","meta":{"footnotes":""},"categories":[6],"tags":[],"class_list":["post-632","post","type-post","status-publish","format-standard","hentry","category-vol2no1"],"_links":{"self":[{"href":"https:\/\/biomedpharmajournal.org\/staging\/wp-json\/wp\/v2\/posts\/632","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/biomedpharmajournal.org\/staging\/wp-json\/wp\/v2\/posts"}],"about":[{"href":"https:\/\/biomedpharmajournal.org\/staging\/wp-json\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"https:\/\/biomedpharmajournal.org\/staging\/wp-json\/wp\/v2\/users\/2"}],"replies":[{"embeddable":true,"href":"https:\/\/biomedpharmajournal.org\/staging\/wp-json\/wp\/v2\/comments?post=632"}],"version-history":[{"count":5,"href":"https:\/\/biomedpharmajournal.org\/staging\/wp-json\/wp\/v2\/posts\/632\/revisions"}],"predecessor-version":[{"id":32976,"href":"https:\/\/biomedpharmajournal.org\/staging\/wp-json\/wp\/v2\/posts\/632\/revisions\/32976"}],"wp:attachment":[{"href":"https:\/\/biomedpharmajournal.org\/staging\/wp-json\/wp\/v2\/media?parent=632"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"https:\/\/biomedpharmajournal.org\/staging\/wp-json\/wp\/v2\/categories?post=632"},{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/biomedpharmajournal.org\/staging\/wp-json\/wp\/v2\/tags?post=632"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}