Methods of identification of brix and fruit in natural juice Charalampos Proestos, Lecturer in Food Chemistry Department of Chemistry, National and Kapodistrian University of Athens AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Legislation 1. EU: The new Fruit Juice Directive (2012/12) Relating to fruit juices and certain similar products intended for human consumption Until 28 April 2015 products placed on the market or labeled before 28 October 2013 can be sold. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Legislation The composition of fruit juices, concentrated fruit juices, dehydrated fruit juices and fruit nectars, their reserved names, their manufacture and labelling characteristics are subject to specific Community rules under Directive 2001/112/EC amended in August 2009 by the Commission Directive2009/106/EC AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Legislation 2.CODEX GENERAL STANDARD FOR FRUIT JUICES AND NECTARS. (CODEX STAN247-2005).   3. Association of the Industry of Juices and Nectars from fruits and vegetables (AIJN) – EUROPEAN FRUIT JUICE ASSOCIATION Reference guidelines (RF) + Code of Practice for evaluation of quality and authenticity of juices. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

Analytical criteria for authentic juices commercial standards have existed for a number of years in countries such as France, Germany, and the Netherlands German Fruit Juice Industry guide values, known as the RSK values AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

Adultaration-Current issues (1) overdilution of juices with water (2) use of cheaper solid ingredients (particularly sugars) (3) blending of cheaper with more expensive juices. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

Overdilution of juices with water Application of a minimum solids content (measured in degrees Brix). They normally apply to juices prepared by adding water to concentrate and not to ‘(NFC) not from concentrate’ products. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

Use of cheaper solid ingredients (particularly sugars) by far the most common E.g. apple juice will normally contain around 11% by weight of solids. 90% of these solids are carbohydrates – sucrose, dextrose and fructose predominate. Addition of cheaper sources of carbohydrates can be used to ‘stretch’ apple juice AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Continue.. One obvious means of concealing added sugar is to increase acidity with added organic acids. commercial malic acid was used that contained both D- and L- forms of the acid pure fruit juices only contain the L-form. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

Blending of cheaper with more expensive juices. A cheaper juice can be used to adulterate a more expensive one E.g. elderberry juice can be used to extend strawberry or raspberry juice Juice authenticity AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

Definitions (Directive 12/2012) (a) Fruit juice The fermentable but unfermented product obtained from the edible part of fruit which is sound and ripe, fresh or preserved by chilling or freezing of one or more kinds mixed together having the characteristic colour, flavour and taste typical of the juice of the fruit from which it comes. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

(b) Fruit juice from concentrate The product obtained by reconstituting concentrated fruit juice with potable water that meets the criteria set out in Council Directive 98/83/EC of 3 November 1998 on the quality of water intended for human consumption AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 SS and Brix The soluble solids content of the finished product shall meet the minimum Brix level for reconstituted juice specified in Annex V. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 ANNEX V AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Annex 5 continue.. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

FJFC - Definitions continue.. If a juice from concentrate is manufactured from a fruit not mentioned in Annex V, the minimum Brix level of the reconstituted juice shall be the Brix level of the juice as extracted from the fruit used to make the concentrate. Concentrated fruit juice The product obtained from fruit juice of one or more fruit species by the physical removal of a specific proportion of the water content. Where the product is intended for direct consumption, the removal shall be at least 50 % of the water content. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

3. Water extracted fruit juice The product obtained by diffusion with water of: — pulpy whole fruit whose juice cannot be extracted by any physical means, or — dehydrated whole fruit. 4. Dehydrated/powdered fruit juice The product obtained from fruit juice of one or more fruit species by the physical removal of virtually all the water content. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 5. Fruit nectar The fermentable but unfermented product which: — is obtained by adding water with or without the addition of sugars and/or honey to the products defined in points 1 to 4 to fruit purée and/or to concentrated fruit purée and/or to a mixture of those products, and — meets the requirements of Annex IV. Flavour, pulp and cells obtained by suitable physical means from the same species of fruit may be restored to the fruit nectar. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Annex IV AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Continue.. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 More Annex IV AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

Definitions of Raw Materials (1) Fruit All fruits. For the purposes of this Directive, tomatoes are also considered as fruit. (2) Fruit purée The fermentable but unfermented product obtained by suitable physical processes such as sieving, grinding, milling the edible part of whole or peeled fruit without removing the juice. (3) Concentrated fruit purée The product obtained from fruit purée by the physical removal of a specific proportion of its water content. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Continue.. (4) Flavour (5) Sugars — sugars as defined by Council Directive 2001/111/EC of 20 December 2001 relating to certain sugars intended for human consumption, — fructose syrup, — sugars derived from fruits. (6) Honey The product defined by Council Directive 2001/110/EC of 20 December 2001 relating to honey (7) Pulp or cells The products obtained from the edible parts of fruit of the same species without removing the juice. Furthermore, for citrus fruit, pulp or cells are the juice sacs obtained from the endocarp. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

Estimation of juice or fruit content 1. assessing the ash content, 2. levels of potassium ions, 3. phosphorous content, 4. total level of free nitrogen by measuring the formol value, 5.levels of specific amino acids and a range of other AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

Brix measurement in juices AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Refractometers AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Testing the Amount Of Juice, The Acid And The Sugar Levels In Citrus Fruit Testing Fruit Percent Juice Content: Juice content is an important measure of internal quality. Under or over-ripe fruit tend to be less juicy, which directly affects eating quality. The juice content is determined by weighing components of the whole fruit and the juice. - Brix (total soluble solids): In the citrus industry this is a measure of the total soluble solids in the juice. These soluble solids are primarily sugars; sucrose, fructose, and glucose. Citric acid and minerals in the juice also contribute to the soluble solids. Brix is reported as "degrees Brix" and is equivalent to percentage. For example, a juice which is 12 degrees Brix has 12% total soluble solids. The National standards state the minimum brix sugar/ acid ratio. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Continue Acid (titratable acidity): This is a measure of the total acid present in a juice. The predominant acid naturally occurring in orange juice is citric acid. There are also small amounts of malic acid and tartaric acid present. The amount of acid present in the juice is reported as percent citric acid. It needs to be noted that the total acid cannot be measured by pH because the acids concerned are “weak acids” and not completely ionized. The acid content must be measured using a titration with sodium hydroxide. Sodium benzoate determination!!! AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

Determining soluble solids content (% sugar or °Brix) AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Continue.. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Determination of total acidity The acidity of natural fruit juices is the result mainly of their content of various organic acids. For example, most fruits contain the tricarboxylic acid, citric acid, whereas grapes are rich in tartaric acid & peaches, apricots & plums in malic acids. Both tartaric & malic acids are dicaroxylic acids. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Tartaric acid MW=150.09 malic acid MW=134.09 citric acid MW=192.43 AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 The acidity of fruit juice may be determined by simple direct titration with 0.1M sodium hydroxide, using phenolphthalein (ph.ph) as an indicator. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Method : 1. Weight 10g of sample in conical flask & add 50ml distilled water 2. Titrate using 0.1M NaOH & phenolphthalein (ph.ph) as indicator 3- Calculate the total acidity of your fruit juice AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 citric acid= 0.1 x vol.of NaOH(ml) x 10-³ x 192.43/3 Total acidity= wt of acid/wt of sample *100 0.39-1.1 Normal range for citric acid = AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Example AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

Determining sugar/acid ratio AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Industry Standards AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

Determination of sodium benzoate Sodium benzoate (C6H5COO Na ,MW=144 ) is a commonly used preservative which is added to fruit juices to prevent the growth of microorganisms. When added in high conc., it affects the taste of juice. Sodium benzoate is usually permitted at a conc. Of up to 1.3g/l of juice. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 The benzoate anion is not soluble in non-polar solvents because of its negative charge. However, in acid solution, benzoic acid is formed. This is neutral & quite non-polar. Moreover, it is soluble in non-polar solvents, into which it may be extracted at acidic PH, i.e. well below the pka of the carboxyl group , which is 4.20 . AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 In an experiment , benzoic acid is extracted into chloroform, which is then removed by evaporation. After dissolving the residue in 50% (v/v) neutralized ethanol, the benzoic acid is titrated with 0.05M sodium hydroxide, using phenolphthalein as an indicator. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Method: Weight 10g of sample into a graduated flask & add 1ml of 10% NaOH solution & 12 g NaCl. Add sufficient water to bring the vol. up to about 50ml & let it stand for 30 min. with frequent shaking . 2- Add drops of ph.ph (the color will change) , add drops of HCL until the color change (or disappear), then add excess 3 ml HCL 3. Transfer into a separatory funnel 4. Add 25ml of chloroform AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 4. let it sand for 30min with frequent shaking. 5. Transfer 12.5ml of the chloroform layer (low layer) into a conical flask & evaporate off the chloroform on a steam bath 6. Add 50ml of 50%ethanol solution 7. Titrate with 0.05M NaOH using phenolphthalein(ph.ph) as indicator 8. Calculate the amount of sodium benzoate in the sample. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 1ml of 0.05M NaOH =0.0072 sodium benzoate. Ttitre ml of NaOH = x Wt of sodium benzoate(x) = ml * 0.0072 %of sodium benzoate= wt sodium benzoate/ wt of sample *100 Normal range not exceed 0.13% AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

Determination of total solids Remnants of pulp & other solids are easily determined gravimetrically, after evaporation of water from a known weight of sample. The solid present in juice include sugar , organic acid & pectins AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Method: 1-Weight a dry dish 2-Add 15ml of juice & weight 3-Place the dish & its content on a boiling water bath & evaporate to dryness 4-Place the dish in an oven for 2hrs. 5-Put the dish in descicator to cool % weight. AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 %total solid=( wt of beaker after heating_ wt of empty beaker *100)/ Wt of sample(g) AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

Code of Practice for evaluation of fruit juices AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014

AGR 57476 , Cairo, Egypt, 10 - 11 November 2014 Any Questions? harpro@chem.uoa.gr AGR 57476 , Cairo, Egypt, 10 - 11 November 2014