Biochemical Analysis Determination Of Sugar Content Biology Essay
Determination of sugars ( entire sugar, cut downing sugar and non-reducing sugar ) were carried out though Lane and Eynon Method as described by James ( 1995 ) .Entire sugar and cut downing sugar: Took 5 g of sample into a beaker and added 100 milliliter of warm H2O. The solution was stired until all the soluble affairs were dissolved and filtered through wattman paper into a 250 volumetric flask. Pipetted 100 milliliter of the solution prepared into a conelike flask, added 10 milliliter diluted HCl and boiled for 5 min. On chilling, neutralize the solution to phenolphthalein with 10 % NaOH and do up to volume in a 250 volumetric flask.
This solution was used for titration against Fehling ‘s solution and reading was calculated as follow.% Entire sugar = Factor ( 4.95 ) ten dilution ( 250 ) x 2.
5Titre x wt of sample x 10% Entire sugar = Factor ( 4.95 ) ten dilution ( 250 ) x 2.5Titre x wt of sample x 10% Reducing sugar = Factor ( 49.
5 ) ten dilution ( 250 )Titre x wt of sample x 10Non-reducing sugar was estimated as the difference between the entire sugar content and cut downing sugar content.
Determination of dietetic fibre
Entire dietetic fiber content is measured harmonizing to the AOAC enzymatic-gravimetric method. The footing of this method is the isolation of dietetic fiber by enzymatic digestionof the remainder of the components of the stuff. The residue is measured gravimetrically.Starch is digested by uniting amylase ( pH=6.0, Ta=100 & A ; deg ; C, t=30min ) and amyloglucosidase ( pH=7.5, Ta=60 & A ; deg ; C, t=30min ) ; protein is digested with peptidase( pH = 4.5, T a = 60 & A ; deg ; C, t = 30 min ) .
Analysiss are performed utilizing a entire dietetic fiber assay kit ( Sigma merchandise N & A ; deg ; TDF 100 kit ) . Ethanol is added to precipitate the soluble fiber. Filtration is carried out in melting pots with 0.5 g of moisture and homogeneously distributed celite. The residue is so filtered off and washed with 78 % and 95 % ethyl alcohol and propanone. Crucibles with the residue are dried to mensurate the weight of residue.
Protein, ash and amylum were measured in each residue in order to rectify the values for dietetic fiber. Weight of dietetic fibre gave was calculated by the undermentioned expression.Crud Fiber % = ( c-b ) – ( d-b ) x 100( a )[ 6 ] .
3. Determination of wet content
Moisture content is one of the most of import factors act uponing seed quality and storability, Therefore, its appraisal during seed quality finding is of import. Seed wet content can be expressed either on moisture weight footing or on dry weight footing, in seed testing, it is ever expressed on a wet weight footing, and the method for its computation is given subsequently. Seed wet content can be determined either by air oven or moisture metre. However, if prescribed criterion for wet content is less than 8 % , air oven method shall be used.
( 1 ) Air oven method: In this method, seed wet is removed by drying at a specified temperature for a specified continuance. The wet content is expressed as a per centum of the original weight ( wet weight footing ) . It is the most common and standard method for seed wet finding.( 2 ) Moisture metres: A assortment of wet metres are available in the market.These metres estimate seed wet rapidly but the appraisal is non every bit precise as by the air-oven method. The metres should be calibrated and standardized against the air-oven method.The wet content of the sample is calculated utilizing the undermentioned equation:W % = A-B ten 100BacillusWhere:% W = Percentage of wet in the sample,A = Weight of wet sample ( gms ) , andB = Weight of dry sample ( gms )
Determination of ash content
For finding of ash content, method of AOAC ( 2000 ) was followed. Harmonizing to the method, 10 g of each sample was weighed in a silicon oxide melting pot. The melting pot was heated in a muffle furnace for about 3-5 H at 600 & A ; deg ; C. It was cooled in desiccators and weighed to completion of ashing. To guarantee completion of ashing, it was heated once more in the furnace for half an hr more, cooled and weighed. This was repeated accordingly till the weight became changeless ( ash became white or grey white ) . Weight of ash gave the ash content was calculated by the undermentioned expression.
Ash % = Weight of ashed sample x 100Weight of sample taken
5. Determination of protein
Protein was determined utilizing micro Kjeldahl method as describe in AOAC ( 2000 ) .2 g of sample stuff was taken in a Kjeldahl flask and 30 milliliter concentrated sulphuric acid ( H2SO4 ) was added followed by the add-on of 10 g K sulfate and 1 g Cu sulfate. The mixture was heated foremost gently and so strongly once the frothing had ceased. When the solution became colourless or clear, it was heated for another hr, allowed to chill, diluted with distilled H2O ( rinsing the digestion flask ) and transferred to 800 milliliters Kjeldahl flask. Three or four pieces of granulated Zn and 100 milliliter of 40 % acerb sodium carbonate were added and the flask was connected with the splash caputs of the distillment setup. Following 25 milliliter of 0.1 N sulfuric acid was taken in the receiving flask and distilled.
When two-thirds of the liquid had been distilled, it was tested for completion of reaction. The flask was removed and titrated against 0.1 N acerb sodium carbonate solution utilizing methyl ruddy index for finding of Kjeldahl N, which in bend gave the protein content. The nitrogen per centum was calculated by the undermentioned expression.N % = 1.4 ( V2-V1 ) x Normality of Hcl x 250 ( dilution )Weight of SampleWhereas, protein content was estimated by transition of nitrogen per centum to protein ( James, 1995 ) .Protein % = N % x Conversion factor ( 6.
25 )Where transition factor = 100/N ( N % in fruit merchandises )
6. Determination of fat
Fat was determined by Mojonnier method ( James,1995 ) . The fat content was determined gravimetrically after extraction with diethyl quintessence ( ether ) and crude oil quintessence from an ammonium hydroxide alcoholic solution of the sample. About 10 g of sample was taken into a Mojonnier tubing. Added 1 milliliter of 0.880 with 10 milliliters ethanol assorted good and cooled.
Added 25 milliliter diethyl quintessence, stopper the tubing, shacked smartly and so added 25 milliliters crude oil quintessence and left the tubing to be stand for 1 hour. The extraction was replaced thrice utilizing a mixture of 5 milliliters crude oil quintessence and adding the extraction to the distillment flask. Distilled off the dissolvers, dried the flask for 1 hour at 100 C and reweighed.
The per centum fatContent of the sample was calculated by the following expression which gave that the difference in the weight or the original flask and the flask plus extracted fat represent the weight of fat nowadays in the original sample.% Fat content of sample = W2 W1 x 100W3Where:W1 = Weight of empty flask ( g )W2 = Weight of flask + fat ( g ) andW3 = Weight of sample taken ( g ) .