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the major composition of these frying mediums is triacylglycerols . during frying triacylglycerols and fatty acids
these products include hydroperoxides , epoxides , epidioxides , hydroxides , and several other products with simple or complex structures [ 2 , 3 ] .
recent studies showed that several analytical methods can be applied to determine their absolute structures [ 4 , 5 ] .
however , due to the complex nature of food frying , it is still debatable to conclude on each specific method or oxidized compounds produced during frying .
it was found that the primary oxidation products are further oxidized to form secondary and tertiary oxidation products .
the aldehydes and other reactive substances are one of the main causes of rancidity in foods during preparation and storage .
one of the important oxidation products is known as malondialdehyde ( mda ) , which is considered as the main marker in lipid peroxidation .
thiobarbituric acid ( tba ) is reacted with mda , which is resulting in a colour compound , which can be determined spectrophotometrically , chromatographically , or through image processing techniques [ 810 ] . due to the reactivity of tba with several reactive substances in the biological sample , a more widely accepted terminology called thiobarbituric acid reactive substances ( tbars )
tbars is now considered as a standard marker for the lipid peroxidation induced oxidative stress .
meat or meat products upon frying also produced several oxidation products , which can be measured using tba - mda adduct with the help of hplc .
several hplc methods are available regarding the tbars assay [ 9 , 13 ] . however , due to the high cost and long operation timing of the hplc procedures , a more simple method is therefore required .
developed a spectrometric method for the tbars as marker for lipid oxidation in animal tissues , food , and feed substances .
however , the method involves 1,1,3,3-tetraethoxypropane ( tep ) as mda precursor , the hydrolysis of which would need a trained analytical chemist .
the method also involves hexane during extraction procedures , which was found to cause losses in the mda contents from the fried fast foods and was thus not applicable for fried fast foods .
recently papastergiadis et al .   developed a spectrophotometric method for analysis of mda in oxidized foods such as peanuts , almonds , walnuts , cookies , crisps , and trout samples .
their method was comparable with hplc method ; however , there is a lack of information about the mda analysis in complex fried food matrix such as traditional fried street foods .
this paper presents a simple and highly sensitive spectrometric method for the analysis of tba reactive substances in the traditional fried street foods .
thiobarbituric acid ( tba ) 99% pure was purchased from bdh ( bdh , england ) ; malondialdehyde tetrabutylammonium salt ( mda salt ) 96% pure and methanol 99.8% pure were from sigma - aldrich ( steinheim , germany ) . glacial acetic acid ( 99101% pure ) was purchased from daejung ( daejung , korea ) .
the standard solution of 4.0  mm of tba was prepared in glacial acetic acid . for this purpose , 57.66  mg of tba
standard stock solution of mda ( 1  mm ) was prepared in glacial acetic acid . mda ( 31.35  mg ) was accurately weighed and dissolved in 100  ml solvent . from the stock solution ,
different concentrations of 0.1 , 0.2 , 0.4 , 0.6 , and 0.8  mm were prepared .
the calibration curve was constructed in the concentration range of 0.1 to 1.0  mm .
one gram of each fried grinded sample ( samosa , shami kebab , fried bread , and potato chips ) was taken in 25  ml test tube and 5  ml of the solvent .
the solvent was either 100% glacial acetic acid ( aa ) or 50% glacial acetic acid in water ( aw ) .
the standard mda solution ( 1  ml ) was taken in a 10  ml test tube and mixed with tba ( 1  ml ) .
the test tubes were cooled at room temperature and absorbance was measured at 532  nm using uv - visible spectrophotometer model pharmaspec 1700 ( shimadzu , japan ) .
each standard for the calibration was repeated ( n = 3 ) according to the above procedure .
a blank sample was repeated ( n = 5 ) replacing standard or sample by acetic acid or water
. samples of the fast foods collected were samosa , shami kebab , fried bread , and potato chips .
two different kinds of samples extracts were prepared , that is , with 100% glacial acetic acid ( aa ) and 50% glacial acetic acid with water ( aw ) .
the extract of each sample ( 1  ml ) was mixed with 1  ml tba reagent and the above procedure was repeated five times ( n = 5 ) .
the tbars was calculated using the formula as m / g of the sample:(1)tbars  m / g = acv / w , where ac is the amount determined from the calibration curve and w is the weight of the sample taken while v is volume in ml or dilution factor of the total extract prepared .
the analytical method was validated according to the guidelines of the international conference on harmonization ( ich ) .
linearity was determined from the different concentration measured ( n = 3 ) in the range of 0.11.0  m .
limits of detection ( lod ) and limits of quantitation ( loq ) were determined from the standard calibration curve .
precision was determined in terms of intraday ( n = 3 ) and interday ( n = 5 ) at three concentration levels of 0.1 , 0.4 , and 0.8  m .
stability was measured ( n = 9 ) at concentration of 0.8  m . the accuracy of the method was measured using recovery studies in samosa samples ( n = 9 ) at two different extraction procedures .
the pure glacial acetic acid has extraction code of aa , while aw was code given to 50% glacial acetic acid in water .
glacial acetic acid was found to be the best solvent for extractions and preparation of reagent .
the present method is based on the unified solvent system with high dissolution and extraction power .
tep used in the previous studies [ 14 , 16 ] has been found to produce artifacts and therefore mda salt was used in this study .
this study thus does not require preparation of buffers needed for the hydrolysis of tep and also does not produce the false negative results from its partial hydrolysis .
the mda - tba mixture was reacted for 60  min ( figure 1 ) .
previous study and our observations showed that 60  min was optimum time for mda - tba reaction .
two extractions were performed using glacial acetic acid ( 100% ) and glacial acetic acid ( 50% ) in water .
the calibration curve of six points in triplicate ( n = 3 ) was established in the concentration range of 0.1 to 1.0  m . the present method was more sensitive than previously reported by papastergiadis et al . , who showed a linearity range of 0.610  m .
linear regression shows a correlation coefficient of 0.9929 with equation of y = 1.4167x + 0.0785 as shown in figure 2 .
the lod and loq were evaluated from the slope and residual standard deviations of the standard curve .
instrumental precision was determined by replicate ( n = 9 ) analysis of standard compounds .
the results showed higher precision ( 4.23 , % rsd ) for pure glacial acetic acid than 50% glacial acetic acid ( 6.37 , % rsd ) .
repeatability ( % rsd ) was determined using intraday and interday analyses of three standard concentrations ( 0.1 , 0.4 , and 0.8  m ) in replicates ( n = 3 ) .
the intraday precision for the concentration of 0.1  m was 11.3% , that for concentration of 0.4  m was 10.8% , and that for concentration of 0.8  m was 2.03% ( % rsd ) .
the interday precisions of 8.4 , 7.9 , and 13.1% were obtained for the concentration of 0.1 , 0.4 , and 0.8  m , respectively , as shown in table 1 .
the overall precision of this method was higher than reported methods [ 14 , 15 , 17 ] .
the specificity of the method was assessed using the absorption spectra of the standard mda - tba adduct formed after reaction and also in the sample .
it was found that absorption spectra were a good tool for determination of specificity of individual class of compound .
samples of samosa were spiked with 0.4  m of the standard addition method ( n = 9 ) .
the recovery studies were carried out in both pure glacial acetic acid and 50% glacial acetic acid .
results show a higher recovery for pure glacial acetic acid samples ( 114.3% ) than its corresponding 50% glacial acetic acid samples ( 96.01% ) .
the recovery of the method was higher than other spectrophotometric methods reported previously [ 15 , 18 ] .
the extraction using pure glacial acetic acid was found to have higher recovery than previously reported methods with hplc as shown in table 2 [ 10 , 17 ] .
four samples of the street foods were selected , because of the wide uses and popularity .
shami kebab is made of cooked grinded chickpeas , hot spices , onion , garlic extract , and grinded tomatoes and was fried in vegetable ghee .
the aa extract has lower tbars values ( 1.10  0.06  m / g , mean  sd ) than its corresponding aw extracts ( 1.505  0.09 , mean  sd ) .
revealed recently that deep fried shami kebab had higher amount of tbars than the grilled one and reported lower values of tbars than the present studies .
the difference may be due to the difference in the laboratory and fast food restaurant frying or frying medium .
these results indicate that the present simple method can give us a good look at the check and balance system in the fast food restaurants or streets foods frying .
the typical composition of samosa consists of wheat starch , onion , potatoes , and tomatoes .
only the outer crispy part of the samosa was analyzed , because this part is highly exposed to frying temperature and frying oil .
similar to shami kebab , the lower amount of tbars was obtained in aa samples ( 1.02  0.18  m / g , mean  sd ) than its corresponding aw samples ( 2.51  0.19  m / g , mean  sd ) .
a higher amount of tbars was obtained in aa samples ( 0.891  0.09  m / g , mean  sd ) than its corresponding aw samples ( 0.372  0.03  m / g , mean  sd ) .
the difference in the tbars may be due to its dissolution power and polarity of the solvent and solute .
the results of the tbars of the potato chips showed a higher amount in aw ( 2.911  0.13  m / g , mean  sd ) than aa ( 2.21  0.13  m / g , mean  sd ) .
the high amounts of lipids , high surface - to - volume ratio , and packaging in the presence of oxygen can result in oxidative deterioration of the chips .
thus , it is imperative to keep samples under standard conditions , while checking the proper healthy level using simple laboratory protocol .
in this study , a simple and sensitive spectrophotometric method was developed for the determination of tbars in fried street foods .
the method was precise , sensitive , and highly reproducible for quantitative determination of tbars .
the precision of extractions and analytical procedure was very high as compared to the reported methods .
the method was used to determine the tbars contents in the fried foods such as shami kebab , samosa , fried bread , and potato chips .
the method can be successfully used for the determination of tbars in other food matrices for quality control analysis in food industries or regular food inspection system .
a simple and highly sensitive spectrophotometric method was developed for the determination of thiobarbituric acid reactive substances ( tbars ) as a marker for lipid peroxidation in fried fast foods . 
 the method uses the reaction of malondialdehyde ( mda ) and tba in the glacial acetic acid medium . 
 the method was precise , sensitive , and highly reproducible for quantitative determination of tbars . 
 the precision of extractions and analytical procedure was very high as compared to the reported methods . 
 the method was used to determine the tbars contents in the fried fast foods such as shami kebab , samosa , fried bread , and potato chips . 
 shami kebab , samosa , and potato chips have higher amount of tbars in glacial acetic acid - water extraction system than their corresponding pure glacial acetic acid and vice versa in fried bread samples . 
 the method can successfully be used for the determination of tbars in other food matrices , especially in quality control of food industries .