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Annals of Chromatography and Separation Techniques

RP-HPLC Method Development and Validation for the Simultaneous Estimation of Diphenhydramine and Bromhexine in Tablet Dosage Forms

[ ISSN : 2473-0696 ]

Abstract Citation Introduction Materials and Methods [7-10] Results and Discussion [11-13] Conclusion References
Details

Received: 14-Jan-2018

Accepted: 02-Jul-2018

Published: 06-Jul-2018

Research Article | Volume 4 - Issue 1 | Article DOI :

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Sivagami B¹, Nagaraju B¹, Pavan Kumar V¹, Sireesha R¹, and Chandrasekar R²*

¹ Department of Pharmaceutical Analysis, Seven Hills College of Pharmacy, India
² Department of Pharmacognosy, Seven Hills College of Pharmacy, India

Corresponding Author:

Sivagami B, Department of Pharmaceutical Analysis, Seven Hills College of Pharmacy, Venkataramapuram, Tirupati, Chitoor Dist, 517561, A.P, India, Tel: 0877 329 9990; Email: sivagamib_27@rediffmail.com

Keywords

Bromhexine; Diphenhydramine; RP-HPLC; Simultaneous estimation; Validation

Abstract

Background: A simple, Accurate, precise method was developed for the simultaneous estimation of the Diphenhydramine and Bromhexine in tablet dosage form by RP-HPLC method.

Methods: Chromatogram was run through standard discovery 150 x 4.6 mm, 5m. Mobile phase containing Buffer 0.01N Potassium Dihydrogen Phosphate: Acetonitrile taken in the ratio 50:50 was pumped through column at a flow rate of 1 ml/min. Buffer used in this method was 0.01N Potassium Dihydrogen Phosphate and pH adjusted to 3.0 with dilute Orthophosphoric acid solution. Temperature was maintained at 30°C. Optimized wavelength selected was 225 nm. Retention time of Diphenhydramine and Bromhexine were found to be 2.458 min and 2.972.

Results: % Relative Standard Deviation of the Diphenhydramine and Bromhexine were found to be 0.5 and 0.3 respectively. % Recovery was obtained as 99.20% and 99.40% for Diphenhydramine and Bromhexine respectively. Limit of Detection, Limit of Quantitation values obtained from regression equations of Diphenhydramine and Bromhexine were 0.07, 0.20 and 0.11, 0.33 respectively. Regression equation of Diphenhydramine is y = 9539.x + 42940, and y = 9765x + 8034 of Bromhexine.

Conclusion: Since the retention time decreased the run time also decreased. So the method developed was simple and economical that can be applied successfully for simultaneous estimation of both Diphenhydramine and Bromhexine in bulk and combined tablet formulation.

Citation

Sivagami B, Nagaraju B, kumar PV, Sireesha R and Chandrasekar R. RP-HPLC Method Development and Validation for the Simultaneous Estimation of Diphenhydramine and Bromhexine in Tablet Dosage Forms. Ann Chromatogr Sep Tech. 2018; 4(1): 1034.

Introduction

Diphenhydramine is a histamine H1 antagonist used as an antiemetic,antitussive,for dermatoses and pruritus,for hypersensitivity reactions, as a hypnotic,an antiparkinson,and as an ingredient in common cold preparations. It has some undesired antimuscarinic and sedative effects.Chemically diphenhydramine is [2-(diphenylmethoxy) ethyl] dimethylamine. Diphenhydramine competes with free histamine for binding at HA-receptor sites. This antagonizes the effects of histamine on HA-receptors, leading to a reduction of the negative symptoms brought on by histamine HA-receptor binding [1-3] (Figure 1).

Figure 1: Structure of Diphenhydramine.

Bromhexine is an expectorant/mucolytic agent.Bromhexine is an oral mucolytic agent with a low level of associated toxicity. Bromhexine acts on the mucus at the formative stages in the glands, within the mucus-secreting cells.Bromhexine disrupts the structure of acid mucopolysaccharide fibres in mucoid sputum and produces less viscous mucus, which is easier to expectorate.Chemically Bromhexine is 2,4dibromo6 {[cyclohexyl(methyl)amino]methyl} aniline [4-6] (Figure 2).

Figure 2: Structure of Bromhexine.

The literature review revealed that several analytical methods have been reported for Diphenhydramine and Bromhexine in UV-Spectrophotometry, RP-HPLC, individually and in combination. This research work implicates the simultaneous estimation of Diphenhydramine and Bromhexine by RP-HPLC in tablet dosage forms.

Materials and Methods [7-10]

Materials

Combination Diphenhydramine and Bromhexine tablets (Histachlor Oyster Labs Limited) received from spectrum lab, Distilled water, Acetonitrile, Phosphate buffer, Methanol, Potassium dihydrogen ortho phosphate buffer,Ortho-phosphoric acid. All the above chemicals and reagents used were analytical grade and procured from Rankem Laboratories Pvt Ltd.

Instruments

Electronics Balance-Denver, pH meter-BVK enterprises,India,Ultrasonicator-BVK enterprises,WATERS HPLC 2695 SYSTEM equipped with quaternary pumps, Photo Diode Array detector and Auto sampler integrated with Empower 2 Software. UV-VIS spectrophotometer PG Instruments T60 with special bandwidth of 2 mm and 10mm and matched quartz cells integrated with UV win 6 Software was used for measuring absorbance’s of Diphenhydramine and Bromhexine solutions.

Methods

Diluents: Based up on the solubility of the drugs, diluents was selected, Acetonitrile and Water taken in the ratio of 50:50.

Preparation of standard stock solutions: Accurately weighed 25mg of Diphenhydramine, 8mg of Bromhexine and transferred to 10ml and 10ml individual volumetric flasks and 3/4th of diluents was added to these flask and sonicated for 10 minutes. Flask were made up with diluents and labeled as Standard stock solution. (2500µg/ml of Diphenhydramine and 800µg/ml Bromhexine).

Preparation of standard working solutions (100% solution): 1ml from each stock solution was pipetted out and taken into a 10ml volumetric flask and made up with diluent (250µg/ml of Diphenhydramine and 80µg/ml of Bromhexine).

Preparation of sample stock solutions: 5 tablets were weighed and the average weight of each tablet was calculated, then the weight equivalent to 1 tablet was transferred into a 10 ml volumetric flask, 10ml of diluents was added and sonicated for 25 min, further the volume was made up with diluents and filtered by HPLC filters (2500µg/ml of Diphenhydramine and 800µg/ml of Bromhexine).

Preparation of sample working solutions (100% solution): 1ml of f iltered sample stock solution was transferred to 10ml volumetric f lask and made up with diluents (250µg/ml of Diphenhydramine and 80µg/ml of Bromhexine).

Preparation of buffer: 0.1% OPA Buffer: 1ml of orthophosphoric acid was diluted to 1000ml with HPLC grade water. Buffer: 0.01N Potassium dihyrogen ortho phosphate.

Accurately weighed 1.36gm of Potassium dihyrogen orthophosphate in a 1000ml of Volumetric flask add about 900ml of milli-Q water added and degas to sonicate and finally make up the volume with water then added 1ml of Triethylamine then PH adjusted to 3.0 with dil.Orthophosphoric acid solution (Tables 1 & 2).

Table 1: Optimization of chromatographic conditions.

Trials Mobile phase Flow rate Column Detector wave Column Injection Run Diluent
length temp volume time
Trial 1 Acetonitrile and 0.1%OPA 1 ml/min BDS C18 (4.6 x 225nm 30°C 10µL 10 min Water and Acetonitrile
taken in the ratio 50:50 150mm, 5µm) in the ratio 50:50
Trial 2 0.01N Kh2po4: 1 ml/min BDS C18 (4.6 x 225nm 30°C 10µL 10 min Water and Acetonitrile
Acetonitrile (50:50) 150mm, 5µm) in the ratio (50:50)
Trial 3 50% Water: 50% 1 ml/min BDS C18 (4.6 x 225nm 30°C 10µL 10 min Water and Acetonitrile
Methanol 150mm, 5µm) in the ratio 50:50
Trial 4 60% OPA (0.1%): 40% 1 ml/min Hiber BDS C18 (4.6 x 225nm 30°C 10µL 10 min Water and Acetonitrile
Acetonitrile 150mm, 5µm) in the ratio 50:50
Optimized 50% 0.01N kh2po4 : 50% 1 ml/min Discovery BDS C18 225nm 30°C 10µL 6 min Water and Acetonitrile
method Acetonitrile (4.6 x 150mm, 5µm) in the ratio 50:50

Table 2: Results of chromatographic conditions.

Trials

Results

Trial 1

Bromhexine and Diphenhydramine were eluted but peak shapes and Bromhexine peak having less USP plate count so further trial was

carried out

Trial 2

Bromhexine and Diphenhydramine eluted but retention time was more so further trial was carried out

Trial 3

Diphenhydramine eluted but Bromhexine peak was not eluted and peak shape was not good so, further trail was carried out.

Trial 4

Diphenhydramine and Bromhexine both peak are eluted but retention times were more and peak shape also no good so, further trail was

carried.

Optimized method

Both peaks have good resolution, tailing Factor, theoretical plate count and resolution.

Results and Discussion [11-13]

Optimized method (Figure 3)

Observation: Diphenhydramine and Bromhexine were eluted at 2.458 min and 2.972 min respectively with good resolution. Plate count and tailing factor was very satisfactory, so this method was optimized and to be validated.

Figure 3: Optimized Chromatogram.

System suitability: All the system suitability parameters were within the range and satisfactory as per ICH guidelines [14] (Table 3) (Figure 4).

Figure 4: System suitability Chromatogram.

Table 3: System suitability parameters for Diphenhydramine and Bromhexine.

S.No Diphenhydramine Bromhexine
Inj RT(min) USP Plate Tailing RT(min) USP Plate Tailing
Count Count
1 2.456 5716 1.16 2.969 6061 1.11
2 2.456 5716 1.16 2.969 6061 1.11
3 2.458 5769 1.18 2.972 6370 1.1
4 2.458 5757 1.17 2.972 6386 1.09
5 2.465 5673 1.13 2.978 6229 1.1
6 2.465 5679 1.13 2.978 6229 1.1

According to ICH guidelines plate count should be more than 2000, tailing factor should be less than 2 and resolution must be more than 2. All the system suitable parameters were within the limits.

Specificity: Retention times of Diphenhydramine and Bromhexine were 2.458 min and 2.972 min respectively. We did not found and interfering peaks in blank and placebo at retention times of these drugs in this method. So this method was said to be specific (Figure 5).

Figure 5: Chromatogram of blank.

Linearity: Six linear concentrations of Diphenhydramine (62.5-375/ml) and Bromhexine (20-120µg/ml) were injected in a duplicate manner. Average areas were mentioned above and linearity equations obtained for Diphenhydramine was y = 9539.x+42940 and of Bromhexine was y = 9765x+8034 Correlation coefficient obtained was 0.999 for the two drugs (Table 4) (Figures 6 & 7).

Figure 6: Calibration curve of Diphenhydramine.

Figure 7: Calibration curve of Bromhexine.

Table 4: Linearity table for Diphenhydramine and Bromhexine.

Diphenhydramine Bromhexine
Conc (μg/mL) Peak area Conc (μg/mL) Peak area
0 0 0 0
62.5 653277 20 207024
125 1283232 40 414399
187.5 1849097 60 594388
250 2396559 80 767086
312.5 3029852 100 994188
375 3609261 120 1180466

Precision System Precision: From a single volumetric flask of working standard solution six injections were given and the obtained areas were mentioned above. Average area, standard deviation and % RSD were calculated for two drugs. % RSD obtained as 0.3% 0.2% respectively for Diphenhydramine and Bromhexine. As the limit of Precision was less than “2” the system precision parameters were within the limits (Table 5) (Figure 8).

Table 5: System precision table of Diphenhydramine and Bromhexine.

S. No Area of Diphenhydramine Area of Bromhexine
1 2389976 760114
2 2382256 761254
3 2370867 763684
4 2385746 762691
5 2388631 763872
6 2380954 764192
Mean 2383072 762635
S.D 6925.8 1632.2
%RSD 0.3 0.2

Figure 8: System precision chromatogram.

Repeatability: Multiple sampling from a sample stock solution was done and six working sample solutions of same concentrations were prepared, each injection from each working sample solution was given and obtained areas were mentioned in the above table. Average area, standard deviation and % RSD were calculated for two drugs and obtained as 0.5% and 0.3% respectively for Diphenhydramine and Bromhexine. As the limit of Precision was less than “2” the system precision parameters were within the limits (Table 6) (Figure 9).

Figure 9: Repeatability chromatogram.

Table 6: Repeatability table of Diphenhydramine and Bromhexine.

S. No Area of Diphenhydramine Area of Bromhexine
1 2375405 760462
2 2361582 760702
3 2352717 762728
4 2388878 762293
5 2375021 761496
6 2365100 766017
Mean 2369784 762283
S.D 12681.5 2028.6
%RSD 0.5 0.3

Intermediate precision (Day_Day Precision): Multiple sampling from a sample stock solution was done and six working sample solutions of same concentrations were prepared, each injection from each working sample solution was given on the next day of the sample preparation and obtained areas were mentioned in the above table. Average area, standard deviation and % RSD were calculated for two drugs and obtained as 1.2% and 0.3% respectively for Diphenhydramine and Bromhexine. As the limit of Precision was less than “2” the system precision parameters were within the limits (Table 7) (Figure 10).

Figure 10: Inter Day precision Chromatogram.

Table 7: Intermediate precision table of Diphenhydramine and Bromhexine.

S. No Area of Diphenhydramine Area of Bromhexine
1 2295635 765308
2 2251596 760614
3 2287512 761673
4 2304762 760091
5 2301136 761269
6 2336636 760082
Mean 2296213 761506
S.D 27561.1 1968
%RSD 1.2 0.3

Accuracy: Three levels of Accuracy samples were prepared by standard addition method.Triplicate injections were given for each level of accuracy and mean %Recovery was obtained as 99.20% and 99.40% for Diphenhydramine and Bromhexine respectively (Tables 8 & 9) (Figures 11-13).

Figure 11: Accuracy 50% Chromatogram of Diphenhydramine and Bromhexine.

Figure 12: Accuracy 100% Chromatogram of Diphenhydramine and Bromhexine.

Figure 13: Accuracy 150% Chromatogram of Diphenhydramine and Bromhexine.

Table 8: Accuracy table of Diphenhydramine.

% Level Amount Spiked Amount recovered % Recovery Mean
(μg/mL) (μg/mL) %Recovery
  125 123.46 98.77  
  125 124.53 99.63  
50% 125 124.96 99.97  
  250 247 98.8  
  250 247.7 99.08  
100% 250 247.07 98.83  
  375 371.87 99.17  
  375 372.19 99.25  
150% 375 372.48 99.33

99.20%

Table 9: Accuracy table of Bromhexine.

% Level Amount Spiked Amount recovered % Recovery Mean
(μg/mL) (μg/mL) %Recovery
  40 39.91 99.76  
  40 40.2 100.5  
50% 40 39.27 98.19  
  80 78.85 98.56  
  80 79.45 99.31  
100% 80 79.41 99.26  
  120 118.91 99.09  
  120 119.82 99.85  
150% 120 120.04 100.04 99.40%

Sensitivity (Table 10)

Robustness: Robustness conditions like Flow minus (0.9ml/min), Flow plus (1.1ml/min),mobile phase minus (55B:45A),mobile phase plus (45B:55A),temperature minus (25°C) and temperature plus (35°C) was maintained and samples were injected in duplicate manner. System suitability parameters were not much affected and all the parameters were within the limits.% RSD was within the limit (Table 11) (Figures 14 &15).

Figure 14: Flow minus Chromatogram of Diphenhydramine and Bromhexine.

Figure 15: Flow plus Chromatogram of Diphenhydramine and Bromhexine.

Table 10: Sensitivity table of Diphenhydramine and Bromhexine.

Molecule LOD LOQ
Diphenhydramine 0.07 0.2
Bromhexine 0.11 0.33

Table 11: Robustness data for Diphenhydramine and Bromhexine.

S.No Condition %RSD of %RSD of
Diphenhydramine Bromhexine
1 Flow rate (-) 1.1ml/min 0.2 0.5
2 Flow rate (+) 1.3ml/min 0.5 0.5
3 Mobile phase (-) 0.4 0.8
55B:45A
4 Mobile phase (+) 0.7 1.1
45B:55A
5 Temperature (-) 25°C 0.3 0.6
6 Temperature (+) 35°C 1.2 1

Assay: Oyster Labs Limited, bearing the label claims Diphenhydramine 25mg, Bromhexine 8mg (Histachlor). Assay was performed with the above formulation. Average % Assay for Diphenhydramine and Bromhexine obtained was 99.24and 99.75% respectively (Tables 12 & 13) (Figures 16 & 17).

Figure 16: Chromatogram of working standard solution.

Figure 17: Chromatogram of working sample solution.

Table 12: Assay Data of Diphenhydramine.

S.no Standard Area Sample area % Assay
1 2389976 2375405 99.48
2 2382256 2361582 98.9
3 2370867 2352717 98.53
4 2385746 2388878 100.04
5 2388631 2375021 99.46
6 2380954 2365100 99.05
Avg 2383072 2369784 99.24
Stdev 6925.8 12681.5 0.53
%RSD 0.3 0.5 0.54

Table 13: Assay Data of Bromhexine.

S.No Standard Area Sample Area % Assay
1 760114 760462 99.52
2 761254 760702 99.55
3 763684 762728 99.81
4 762691 762293 99.76
5 763872 761496 99.65
6 764192 766017 100.24
Avg 762635 762283 99.75
Stdev 1632.2 2028.6 0.3
%RSD 0.2 0.3 0.3

Degradation

Degradation Studies: Degradation studies were performed with the formulation and the degraded samples were injected. Assay of the injected samples was calculated and all the samples passed the limits of degradation (Tables 14 &15) (Figure 18-21).

Figure 18: Acid chromatogram of Diphenhydramine and Bromhexine.

Figure 19: Base chromatogram of Diphenhydramine and Bromhexine.

Figure 20: Peroxide chromatogram of Diphenhydramine and Bromhexine.

Figure 21: Thermal chromatogram of Diphenhydramine and Bromhexine.

Table 14: Degradation Data of Diphenhydramine.

S.NO Degradation % Drug Purity Purity
Condition Degraded Angle Threshold
1 Acid 4.77 0.159 0.361
2 Alkali 2.73 0.131 0.335
3 Oxidation 1.89 0.306 0.327
4 Thermal 0.97 0.159 0.358
5 UV 0.58 0.128 0.327
6 Water 0.64 0.306 0.325

Table 15: Degradation Data of Bromhexine.

S.NO Degradation % Drug Purity Purity
Condition Degraded Angle Threshold
1 Acid 4.86 1.01 1.266
2 Alkali 2.9 0.781 0.971
3 Oxidation 1.88 0.73 0.903
4 Thermal 1 0.936 1.202
5 UV 0.81 0.764 0.956
6 Water 0.73 0.717 0.889

Conclusion

A simple, Accurate, precise method was developed for the simultaneous estimation of the Diphenhydramine and Bromhexine in Tablet dosage form. The RP-HPLC method developed and validated allows a simple and rapid quantitative determination of Diphenhydramine and Bromhexine in tablet dosage forms. All the validation parameters were found to be within the limits according to ICH guidelines. The proposed method was found to be simple, accurate and specific for the drugs of interest irrespective of the excipients present and the short retention times allows the analyst to analyze number of samples in a short period. The method developed was found to be simple, accurate, precise, rugged, robust and stable under forced degradation conditions. So the established method can be successfully applied for the routine analysis for marketed formulations.

References

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2. Diana S Church, Martin K Church. Pharmacology of Antihistamines. World Allergy Organ J. 2011; 4: S22-S27.

3. George Wong HC. Clinical Professor Long-term use of diphenhydramine. CMAJ. 2015; 187: 1078.

4. John R Horton, Ken Sawada, Masahiro Nishibori, Xiaodong Cheng. Structural Basis for Inhibition of Histamine N-Methyltransferase by Diverse Drugs. J Mol Biol. 2005; 353: 334-344.

5. Peter G Pavlidakey, Erin E Brodell, Stephen E Helms. Diphenhydramine as an Alternative Local Anesthetic Agent. J Clin Aesthet Dermatol. 2009; 2: 37-40.

6. Alessandro Zanasi, Massimiliano Mazzolini, Ahmad Kantar. A reappraisal of the mucoactive activity and clinical efficacy of bromhexine. Multidiscip Respir Med. 2017; 12: 7.

7. Porel A, Sanjukta Haty, Kundu. A Stability-indicating HPLC Method for Simultaneous Determination of Terbutaline Sulphate, Bromhexine Hydrochloride and Guaifenesin. Indian Journal of Pharmaceutical Sciences. 2011; 73: 46-56.

8. Amit Kumar, Sanju Nanda. A validated high performance liquid chromatographic method for estimation of bromhexine and terbutaline in bulk and tablet dosage forms. Pharm Methods. 2011; 2: 218-222.

9. Hirak Joshi V, Shah Ujash A, Patel JK, Patel SM. Development and Validation of Analytical Method for Simultaneous Estimation of Bromhexine HCl and Enrofloxacine in Combined Pharmaceutical Dosage Form. Eurasian J Anal Chem. 2017; 12:1631-1638.

10. Njaria PM, Abuga KO, Kamau FK, Chepkwony HK. A versatile HPLC method for the simultaneous determination of bromhexine, guaifenesin, ambroxol, salbutamol/terbutaline, pseudoephedrine, triprolidine, and chlorpheniramine maleate in cough-cold syrups. Chromatographia. 2016; 79: 1507-1514.

11. Shabana Sulthana, Barun kumar Mehta, Anuradha V, Mandava Basaveswara Rao V. Simultaneous estimation of bromhexine and diphenhydramine in pharmaceutical formulations by reversed phase-high performance liquid chromatography. Journal of Pharmacy Research. 2018; 12: 255-260.

12. Vanita Rohit D, Jinal Tandel, Payal Chauhan, Samir Shah. A novel stability indicating RP-HPLC method development and validation for estimation of Phenylephrine hydrochloride and Bromhexine hydrochloride in their tablet dosage form. Journal of Current Pharma Research. 2016; 6: 1839-1851.

13. Jayalakshmi B, Ramesh J, Kalpana TN, Vijayamirtharaj R. Analytical method development and validation of simultaneous determination of Diphenhydramine HCL, Guaiphenesin and Bromhexine HCL in liquid dosage form by RP-HPLC technique. Journal of Pharmacy Research. 2010; 3: 2868-2870.

14. ICH Harmonised Tripartite Guideline, validation of analytical procedures: Text methodology, Q2 (R1) (2005). International Conference on Harmonization, Geneva, 1-13.

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Structural Characterization of Bamboo Lignin Isolated With Formic Acid and Alkaline Peroxide by Gel Permeation Chromatography and Pyrolysis Gas Chromatography Mass Spectrometry

Fractionation is an effective technology to maximize the utilization of lignocelluloses for the production of chemicals and materials. In this case, bamboo was subjected to a two-step fractionation process based on the concept of biorefinery: (a) formic acid treatment at boiling point under atmospheric pressure for 2 h, and (b) post treatment with alkaline hydrogen peroxide solution containing 1% NaOH and 1% H2 O2 at 80 ºC. The combination of formic acid delignification and alkaline hydrogen peroxide degradation achieved an effective removal of both lignin (delignification rate 94.9%) and hemicelluloses (removal rate 87.4%) from bamboo, producing cellulose rich pulp, formic acid lignin and sugars. To investigate the structural modification of lignin during the fractionation process, the residual lignin in the treated samples was isolated and characterized with multiply techniques including gel permeation chromatography, pyrolysis gas chromatography mass spectrometry, Fourier-transform infrared spectroscopy, etc. The relative ratio of S/G was 1.63 for bamboo milled wood lignin (L1), whereas the lignin isolated from the formic acid treated cellulose-rich fraction (L2) presented a chromatograph similar to that of L1 but had a lower S/G ratio of 1.28. This indicated that a preferential removal of S units during the formic acid fractionation process. In addition, alkaline hydrogen peroxide treatment resulted in more removal of S units, as indicated by a lower S/G ratio of 0.71.

Xun Li¹, Chang-Zhou Chen¹, and Ming-Fei Li¹*

 

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Volatile Compound Profiles by HS GC MS for the Evaluation of Postharvest Conditions of a Peach Cultivar

Volatile Organic Compounds (VOCs) profile of foods obtained by Gas Chromatography/Mass Spectrometry (GC/MS) can be considered a potent tool of food products quality changes occurring as a result of different processing, such as ripening and deterioration. The aim of the present study was the evaluation of volatiles profiles of peaches (cv Springcrest) during their storage in conditions similar to those of long distance transport that normally these products undergo before being placed on market. We investigated control sample (no stored fruit) and peaches stored in cardboard boxes wrapped in heat-sealed HD polythene bags, both in normal and modified atmosphere (0% and 23% CO2 ) after 1 and 8 days of storage at 4°C. GC/MS analysis of these samples allowed the identification of a total of 115 VOCs.

The comparison of the VOCs profile of the three peach samples (control, normal atmosphere and 23% CO2 ) shows that fruits packaged in normal atmosphere released a greater amount of esters of medium chain fatty acids, such as ethyl nonanoate and ethyl dodecanoate. On the other hand, fruits stored in normal atmosphere and modified atmosphere after 8 days of storage (increased concentration of CO2 in packs) released a greater amount of esters of long chain fatty acids, such as ethyl hexadecanoate.

Livia Malorni¹, Antonella Martignetti¹, and Rosaria Cozzolino¹*

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Current Trends in Lignocellulosic Analysis with Chromatography

The conversion of lignocellulosic biomass into biofuel and biomaterial is promising for the substitution of fossil resources in energy and material applications. Given the complexity of plant cell wall, the main challenge is to obtain lignocelluloses with high yield and purity. For a better understanding of lignocellulosic biomass, chromatography stands out as a powerful separation method that can support the lab directed research and pilot scale production of biomaterial and biochemical. This paper provides a review on the characterization of cellulose, hemicellulose and lignin along with their derivatives and decomposed sugar monomers, in particular their isolation and purificationmethods using various specific types of chromatography. Methods with various specific types of chromatography. This review also summarizes different chromatographic methods for obtaining the molecular weights of cellulose, hemicellulose and lignin that have been used in recent years, and highlights future opportunities for the application of those biopolymers.

Fengbo Sun¹ and Qining Sun²*

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Current Status of Two-Dimensional Gel Electrophoresis and Multi-Dimensional Liquid Chromatography as Proteomic Separation Techniques

Proteomics is very important component in the era of post-genomics because it can address functions of genes and some important non-gene-determined biological issues such as Post Translational Modifications (PTMs), splicing, translocation, and spatial structure. Proteome is very complex, including multiple parameters such as kind of proteins, copy number of each protein, PTMs, isoforms, spatial structure of each protein, protein-protein interaction, and protein-other molecule interaction, etc. Moreover, proteome is dynamic, and alters with different conditions such as different physiological processes, different pathological processes, and different disease status.

Xianquan Zhan¹,²,³,⁴*

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Validation of Assay Indicating Method Development of Imatinib in Bulk and Its Capsule Dosage Form by Liquid Chromatography

A novel, simple and economic reverse phase High Performance Liquid Chromatography (RP-HPLC) method has been developed for the quantification of Imatinib in bulk and capsule dosage form with greater precision and accuracy. Separation was achieved on Analytical technologies, C-18, (250mm*4.6mm) column in isocratic mode with mobile phase consisting of acetonitrile: potassium dihydrogen phosphate buffer (pH 2.5) (30:70v/v) with a f low rate of 0.8 mL/min. The detection was carried out at 268 nm. The retention time of Imatinib was found to be 2.67 min. The method was validated as per ICH guidelines. Linearity was established for Imatinib in the range 5-35 μg / ml with r2 value 0.996. The percentage recovery of Imatinib was found to be in the range 99.49-99.67 %. The high recovery and low relative standard deviation confirm the suitability of the proposed method for the estimation of the drug in bulk and capsule dosage forms. Validation studies demonstrated that the proposed RP-HPLC method is simple, specific, rapid, reliable and reproducible for the determination of Imatinib for quality control level.

Nalini Kanta Sahoo¹, Madhusmita Sahu¹, V Alagarsamy¹, B Srividya², and Chinmaya Keshari Sahoo³*

 

 
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