Materials and methods Procedure method of counting

solid as shown in Table II-6 . For semi-solid medium , the composition is given byTable II -7.Table II -6. Composition in grams per 1 liter of semisolid agar RCMRCM broth Hirsch and Grinsted ............................................ .................................. 32 gRCM agar Hirsch and Grinsted ............................................ .................................... 8 gwater .................................................................................................................................. 1000 mlMaterials and methods40- Suspend in 1 liter of distilled or deionized water ;- Bring to a boil slowly , stirring until completely dissolved ;- Dispense tubes screw diameter 16 mm x 160 at 10 ml per tube ;- Sterilize at 121 ° C for 15 minutes.Prepared media should be routinely deaerated before use even inIf they are prepared the day of use, deaerated environments are brought to 46 ± 1 ° Cbefore use.• Agar ( cap)Agar is used as a cap obtained by complete dissolution of the agarbacteriological type E in 1 liter of distilled or deionized water, divided into vials screwmaximum rate of 100 ml per vial and then sterilized at 121 ° C for 15 minutes.6.3.3 . Procedure / method of counting• Thermal testPerform a heat test at 80 ° C for 10 minutes effective .• Inoculation and incubation mediaThree tubes per dilution and serially diluted 3- Transfer 1 ml of the primary dilution in 3 tubes aerated environment, do the same with the 2following dilutions ;- Thoroughly mix the inoculum and medium without introducing air ;- Cool rapidly to semi- solidification ;- Add 3 ml of a plug of agar solution maintained at 46 ± 1 ° C ( approximately 1.5 cm ) ;- Cool again until solidification of the agar plug;- Incubate the tubes at 37 ± 1 ° C for 5 days (120 h ± 4 h) .• Readings tubes and expression of results- Record as positive tubes with gas production (lifting the cap,separation of the tube wall , the agar cracks ... ) ;- Calculate the number of positive tubes per dilution and use this number to read out the coefficientMPN table ;- Only the coefficients belonging to categories 1 and 2 are acceptable;- Express the result as the number of spores per gram of product, taking into account the factorthe first dilution.Materials and methods417 Sensory Analysis .Sensory analysis was conducted in three stages. We conducted a testtriangular , a placement test and a taste test .7.1 . subjectsThe panel consisted of 09 male subjects ; Student graduation and positionsgraduation from the Institute of Nutrition , Food and Agri-Food Technologies( INATAA ) of Mentouri Constantine University and recruited according to their motivation andavailability to participate in the test. It is shown how their ballots will be met,by using bulletins enlarged projected on a screen . We avoided discussingthe food to be tested , explaining the method and analysis protocolsused to reduce confusion and make it easier for tasters. It is importantthey understand the procedures and how to complete scorecards toto participate in the tests on the same basis.Should be advised to avoid the use of tasters products smellpronounced as soaps, lotions and perfumes before participating in a panel and avoideating, drinking or smoking at least 30 minutes prior to testing.7.2 . Sequence analysisSensory analyzes were conducted in three tests ; triangular test, testclassification and hedonic test. Prior to analysis, cheese specialties are analyzingpreviously cut into cube of 10 g samples and equilibrated at room temperature( 21 ° C ) .7.2.1 . triangular testThe triangle test is a form of difference test which is used commonly fordetermining if there are noticeable differences between two samples (ISO 4120 , 1983) . this testis also used to determine the ability of tasters to distinguish between differencesappearance , odor , flavor or texture of foods ( WATTS et al . 1991).Task description tasters : We present three tasterscoded samples , two identical and different, they are asked to identify the sampledifferent and fill out the form given in Figure II-3 . They must select a sample ,although they can not distinguish the difference between samples ( that is , theymust choose at random in case of doubt ) .Materials and methods42Figure II-3 . Bulletin of the triangular testPresentation of samples: The two different samples (A and B ) are showntasters in groups of three , that is to say whether they receive two samples A andB or both samples B and A. The three samples are presented in containersidentical coded with random 3-digit numbers . The three numbers codessamples provided to each taster must be different , even if two samples areidentical .There are six options for order of presentation with the triangular test and are indicatedTable II -8. We must use every order of an equal number of times tobalance the order of presentation. This is only possible if there are six or a multiple tasterssix . Samples are presented together each taster in the order chosenso that they can evaluate the samples from the left to the right. They can tasteseveral times .TEST RECORD TRIANGULARNAME : .............................NAME: .......................DATE: ..............................Question: goutez samples listed below and note the sample among the threeyou perceive different?Order codeMaterials and methods43Table II -7. Six possible orders to serve in a triangular test samplesnumbertasterOrder of presentation of samplesFirst Second Third1256 ( A) 831 ( A) 349 ( B )2256 ( A) 349 ( B ) 831 ( A)3670 ( B ) 256 ( A) 831 ( A)4349 ( B ) 670 ( B ) 256 ( A)5349 ( B ) 256 ( A) 670 ( B )6831 ( A) 349 ( B ) 670 ( B )Data analysis: the significance of the results were analyzed using a binomial testa tail . This test is suitable since it is known that there is a sample that is different and that there istherefore a "correct" answer . The number of panelists who identified the different samplewas totaled and the meaning of the total was checked using the table in Annex 10.7.2.2 . Test rank orderDetermine the extent of this test goal in which the consumer acceptsproduct . The acceptance of a food product usually indicates the actual consumption of theproduct ( purchase and consumption).Task description tasters : Tasters were asked to classifyamong samples coded according acceptance from least to most acceptableacceptable (Figure II-4) . Generally , it does not allow the tie .Sample presentation : We present five samples in containersidentical, coded with random 3-digit numbers . Each sample numberdistinct. All samples are presented simultaneously to each taster in orderplanned in advance or at random, and they are entitled to enjoy repeatedly samples .Figure II-4 , gives the ballot to fill the rank order of acceptance.Data analysis: For the purposes of data analysis, we total rankingsassigned to each sample . This is followed by the determination of the meaning ofdifferences by comparing the totals of rankings for all possible pairs ofsamples using the Friedman test . The differences between all possible pairstabulated rankings are compared to the critical value in the table given in Annex 11 ,to a level of significance of 5 % .Materials and methods44Figure II-4 . Bulletin test rank order7.3.3 . hedonic testIn this study , we followed a process of realization of sensory profiles , andwe are interested in the actual analysis of samples on the basis of the list oftexture descriptors such as tranchabilité on slices of bread , which is a propertyof a food product allowing it to be easily sliced ​​mechanically,Liquid aspect, spreadability which is a property of a food product allowing itto be spread over a surface and appearance of the brittle texture during mastication with the teeth ,taste (bitter , pungent, sweet, salty and sour ) , color and odor (Figure II -5).After evaluating our product, the results were presented as atable and subjected to analysis of variance (ANOVA ) .SHEET PLACEMENT TESTNAME : .............................NAME: .......................DATE: ..............................- Please rank the five samples in order of preference .Order codeMaterials and methods45Figure II-5 . Bulletin for the hedonic test with a rating scale ranging from January to SeptemberData analysis: The results were processed by two independent analyzesXLSTAT using a statistical software (2008 ) . The first analysis to determine theway ANOVA the significance of differences ( significance level was set at 0.05)a cheese product to another , and a sensory characteristic to another withinof the same product . The second analysis method allows component analysismain (ACP ) to reduce a complex system of correlations to a smaller number ofdimensions whose goal is to visualize the correlation between physicochemical parameters andsensory parameters.TEST RECORD hedonicNAME : .............................NAME: .......................DATE: ..............................Please examine and taste each cheese sample , and provide a score from January to Septemberdepending on the intensity of the character .A B C D Esliceable textureliquid texturespreadable texturebrittle texturebitter tastepungencysweetnesssalty tastesour tastecolorodorNOTE:If the taste is mentioned in the record is not detected in the product, you put 0 , notemust be assigned depending on the intensity of taste .Chapter III: Results and Discussion46Chapter III: Results and DiscussionThis study was conducted in order to understand the evolution of parameters andphysicochemical phenomena that occur during the manufacturing process of thespecialty cheese . But first , a partial substitution of milk protein materialby modified starch (E 1422 ) was conducted and the impact of this substitution on the behaviorrheological and texture of the finished product was studied.1. EVOLUTION OF PARAMETERS IN THE PHYSICOCHEMICALMANUFACTURING1.1 . pHObserving the experimental values ​​shows that the pH increases during meltingcheeses in both Metija (Figure III-1 ) process, these results are consistent withobtained by Lee et al. (2004 ) and DIMITRELI et al . (2005 ) . This may be the result ofthe influence of water on the ionic environment of the cheese product by inducingionization of calcium phosphate complexes and different functional groupsamino acids.Moreover , these values ​​show a significant decrease ( p < 0.05 ) pHto 5.67 and 5.97 for Metija Ladhidh Camembert in the finished product , this fall isprobably due to the buffering capacity of the melting salts which could adjust the pH to the right value( Gupta et al , 1984 . BEDROOM et al, 1997 . ) salts of iron chelate calcium boundproteins and thus transform paracaseinate insoluble calcium in paracaseinate sodiumsoluble , which results in the unfolding and dissociation of protein chains ( peptization )( Sood et al , 1979 . LEE et al , 1986 . MARCHESSEAU et al, 1997 . ) .Results and Discussion47Figure III-1 . Evolution of pH during the process of manufacturing MetijaFigure III -2. Evolution of pH during the manufacturing process of Camembert Ladhidh1.2 . solidsThe functional properties of the cheese are controlled by the chemical composition ,including humidity (McMahon et al . 1999). From there, the mutilation of the formulacheese preparations involves effects on the functional behavior in applicationspractices.The decrease in dry matter was not significant (p > 0.05 ) during themanufacturing process of cheese preparation Metija ( Figure III -3 ) . Values45.85 % , 43.84 % , 42.66 % , 41.99 % respectively are recorded in the mixer , after theprecooking after UHT treatment and the conditioner ( finished product Metija ) .5.755.855.715.675.555.605.655.705.755.805.855.90Mixer Precooking UHT ConditionerpH6.126.065.975.855.95.9566.056.16.156.2After grinding Conditioner After cookingpHResults and Discussion48For Ladhidh Camembert , the rate of solids decreased significantly (p <0.05 ) ( Figure III -4 ) . This decrease is very clear after cooking and remains light until theconditioner when the finished product was 39.84 % dry extract.These results are very similar to those made ​​by Hennelly et al. (2005) andNORONHA et al. ( 2008b). The injection of the cold water ( 25 % of the total amount of water injected )in the mixer can be the main cause of the increase of moisture content in themixture and therefore the decrease of the dry extract . After heat treatment; theprecooking ( 68 ° C to 110 ° C ) and the UHT treatment ( 132 ° C to 145 ° C ) , there may be lossesCondensate which are compensated by the injection of hot water at 70 ° C ( 75 % of the total wateradded ) .Figure III-3 . Evolution of the solids during the process of manufacturing MetijaFigure III-4 . Evolution of the solids during the process of manufacturing the LadhidhCamembert45.8543.8442.6641.99394041424344454647Mixer Precooking UHT ConditionerSolids (%)64.2839.94 39.84010203040506070After grinding Conditioner After cookingSolids (%)Results and Discussion491.3 . shorteningAs shown in Figure III -5 , the fat content was 26.00 % , the latternot vary significantly ( p> 0.05 ) after the raw material mixture , but afairly sharp decrease in fat may be noted after precooking itreached 21.50% after the UHT treatment , the latter is kept constant by the followingto finished product. Ladhidh on Camembert , according to Figure III -6 , the mixture offormula contains a rate 30.50% fat . The latter was mainlypies and used as vegetable fat added . After cooking andpackaging , the rate drops to fat 17.50% .The decline in fat can be glossed by the effect of treatmentsthermal, melting salts which have a role emulsifier and homogenising ; that generatea reduction in the size of fat globules up to 1 m in diameter ( RAYAN et al , 1980. ;Heertje et al , 1981. ; KIMURA et al , 1986. ; Lelievre et al, 1990 . ; Tamime et al . 1990)but also a destruction of the membrane that the membrane is replaced by micellescasein and submicelles ( KEENAN et al , 1988; . . Michalski et al , 2002) to formsaid pseudo- proteins which interact with the casein micelles become particles and byFollowing an integral part of the dough and therefore are beyond the quantization ( TUNICKet al , 1997. ; Michalski et al . 2002).Figure III-5 . Evolution of the fat in the process of manufacturing Metija26.00 25.0021.50 21.500.005.0010.0015.0020.0025.0030.00Mixer Precooking UHT ConditionerFat (%)Results and Discussion50Figure III -6. Evolution of the fat in the process of manufacturing the LadhidhCamembert1.4 . Fat / dryThe ratio of the fatty substance relative to the dry matter , during the various stagesmanufacturing Metija is 56.70 % , 57.02 % , 50.39 % and 51.20 % respectively inmixer after precooking after UHT treatment and in the finished product ( Figure III -7 ) . thesevalues ​​are similar to those observed by DIMITRELI et al . (2007) and GLIGUEM et al.( 2009a). The decrease in MG / ES report can be noted after cooking the mixture during themanufacturing Ladhidh Camembert ( Figure III -8 ) .The decrease in the ratio (MG / ES) was not significant (p> 0.05), but indicatesreduction of the fat during the manufacturing process due to the increasethe water content in the dough during the melting under the effect of agitation ( Emmons et al.1980; METZGER and MISTRY , 1994; DRAKE et al. , 1995). According McMAHON et al. ( 1999) ,fat globules may be trapped in the pockets of the serum to be more abundantand larger because of the increased moisture content .30.5018.00 17.500.005.0010.0015.0020.0025.0030.0035.00After grinding Conditioner After cookingFat (%)Results and Discussion51Figure III -7. Evolution of MG / O ratio in the manufacturing process of MetijaFigure III -8 . Evolution of MG / O ratio in the manufacturing process of the LadhidhCamembert1.5. Protein levelsThe protein content ( total nitrogen x 6,38) in the finished product , expressed as g/100 g cheesewas 15.50% and 17.30% for Metija and Ladhidh Camembert respectively . Ratestwo proteins cheese specialties are slightly lower than those reported inwork MOUNSEY and O'Riordan (2008a , 2008b) where the specialty cheese with 3 %had a protein content of 16.90 % and 20.50% in the absence of starch in the formulation .In stages of the melting process , under the action of heating , and the comminuted cheese dispersedwater loses its compact structure. Emulsifying salts calcium chelate protein boundcheese , transforming paracaseinate insoluble calcium in paracaseinate sodiumsoluble . During this process of destabilization of natural cheese , the progress of56.70 57.0250.39 51.200.0010.0020.0030.0040.0050.0060.0070.00Mixer Precooking UHT ConditionerMG / ES (%)47.00 45.0044.000.0010.0020.0030.0040.0050.0060.00After grinding Conditioner After cookingMG / ES (%)Results and Discussion52protein chains and increasing the number of negative charges resulting bothdisappearance of the calcium bridges , enhances the ability of interaction groupspolar side proteins with water molecules ; this is the phase of swelling andhydration ( PACKAGE , 1988). Along this runs hydration process , theparacaseinate sodium emulsifies the fat released in the form of large fat globulesin the mixture during melting . The homogenization caused by the simultaneous effects ofemulsifying salts , heating and agitation reduces the size of the fat globules and improvesand the stability of the emulsion ( RAYAN et al . 1980). Gelation occurs duringcooling the molten mixture is formed in which a protein structureholding dimensional highly water of hydration and the fat emulsified( PACKAGE , 1988). Despite the deep structural changes that occur in theprotein system , the rate of protein remains unchanged during the process of melting .1.6. Rate NaClThe results obtained show that the rate of salt between the two cheese specialtiesare slightly different, they are 0.6% and 0.75% for Metija Ladhidh and Camembert(Table III -1). This difference is probably due to the salt content of the natural cheesesused as raw material , knowing that location is also for Metija cheddar and CamembertLadhidh for Camembert . Nevertheless, the effect of NaCl on the strength of the gels depends on itsconcentration in the product. The NaCl improve the balance between attractive forces andrepulsive forces . At high concentrations , the repulsion between protein moleculesbecome excessive to allow the protein-protein interactions institution tooptimal level.The addition of NaCl in the formula is intended to result in an increase of forcegels and their ability to retain water . Minerals according to their natural properties andtheir concentration in the environment, can influence the attractions and repulsionsElectrostatic intra or inter protein molecules by reducing the electrostatic charge ofgroups COO- and NH3 + proteins ( BAU et al. , 1985).1.7. Ash contentThe determination of ash content in the finished product showed rates ash4.36 ± 0.075% in the specialty cheese Metija , and 3.80 ± 0.09% in the LadhidhCamembert (Table III -1); this result is too close to that reported in the work ofResults and Discussion53Hennelly et al. (2005 ) in which the moisture content in the cheese product was determined46 % and the ash content was 4.56 ± 0.07 % , and 3.93 ± 0.17 % for other specialtycheese with a moisture content of 54 % .In the finished product , the ash is composed mainly of minerals fromdairy raw materials and powders, such as salts of iron; we find thecalcium , phosphorus, potassium , chlorine, sodium , magnesium, and table salt .VARUNSATIAN et al. (1983 ) reported that they have a significant role in thegelation of casein which depends on the ionic strength and the nature of the ions ; ionic strengthequal , the Na + ions would prevent aggregation heated above 70 ° C proteins , whileCa + + ions or Mg + + the favor . This effect of calcium would still be dependent on itsconcentration , an amount of calcium required for being crosslinks interprotéines( KOHNHORST and MANGINO 1985; GUINEA and O'KENNEDY , 2009). From theseAuthors , calcium in high concentrations , inhibits gel formation by promotingprotein-protein interactions leading to aggregation , which will negatively influence theviscosity of the cheese specialty . In the absence of NaCl , the calcium is moreconcentration as sodium and magnesium can adversely affect the consistency of thedough forming salt bridges between molecules of caseins and partially déplisséesnegatively charged ( STEPHEN , 1992).Table III-1 . Summary of physicochemical characteristics of finished products ( and MetijaLadhidh camembert )Metija Ladhidh CamembertpH 5.67 ± 0.30 5.97 ± 0.02Solids (%) 41.99 ± 0.93 39.84 ± 1.04Humidity ( %) 58.01 ± 0.93 60.16 ± 1.04Fat (%) 21.50 ± 0.00 17.5 ± 0.95MG / ES (%) 51.20 ± 0.002 44.00 ± 2.00Protein ( %) 15.50 ± 0.00 17.30 ± 0.00Rate of NaCl ( % ) 0.6 0.75Ash content (%) 4.365 ± 0.075 3.808 ± 0.099Flow (mm) 126.67 ± 5.13 138.33 ± 3.061.8. Morphology of starch granules1.8.1 . Morphology of starch granules1422 )Microscopic observationhydroxypropylated E 1422 ) , under an optical microscope ( 100 x G ) leaves appearstarch are white lenticular shapes, smooth( Figure III -9 ) .Figure III -9. Morphological Observation crosslinked starch granuleshydroxypropyl1.8.1 . Hydrothermal behavior of starch grainsDuring the steps of manufacturing the cheese product , the starch content in theformula suffered the effect of hydrothermal treatmentmechanical , resulting in morphological and textural changes of grainsstarch ( Figure III -10 ) .In the presence of moistureduring precooking (temperaturenative origin ) swelling of the granulesdue to the absorption of watercrystalline structure : issolubilization of granular contentsoptimum cooking with a smudgy (Fin phosphate Distarchcross-linked starch ( distarch phosphateand free of pores or(phosphate DistarchE 1422 ) under an optical microscope ( GX100 )in the finished productand shear forcese water , accompanied by heat treatmentabove the gelatinization temperature of starchcrosslinked starch occurs and volume increaseswater . This swelling of the starch granules is accompanied by a lossthe irreversible gelatinizationgranular . It seems that the starch grains have reached the stage ofFigure III- 10a).Starch grainsResults and Discussion54hydroxypropyl ( Ethe grainscracks, 90 ° Ce but it has the.Grains con crosslinked starchgood water retention . Furthermore ,few granules with breakscrosslinked starch granules couldthermal ( UHT 138 ° C for 3 secondsacidity which can lead to the breakdown of starch grainsoverfire giving a gelatinous texture .Figure III -10. Morphological observation of starch grainsbroke in the specialty cheese state1.9. Variation of the viscosity of the cheese body during the creamingWe can see from Figureis a gradual increasebehavior of the fat , starch and crosslinked para2008b) ; reducing the sizeprotein , favoring the swelling of the starch granulespara- casein during the process of manufacturing the cheese specialty .apparent Metija reaches its threshold75 % . This increase is probably due to the decrease of free watersince the crosslinked starch granules and parato absorb a large portion of the waterpart, resulting in increased viscositycaseins . According Savello et alviscosity by their buffering capacitycontinue to maintain the state of swelling. Microscopic observation revealed the existence of(Figure III- 10b). According Nayouf et al. (2003 )starch be justified by the constraints imposed by theseconds) , but also by forcesstarch , they have reached the stagecrosslinked bservation (a