+++++

( 1 ) OJ NO L 54 , 5 . 3 . 1979 , P . 1 .

( 2 ) OJ NO L 359 , 15 . 12 . 1981 , P . 1 .

( 3 ) OJ NO L 54 , 5 . 3 . 1979 , P . 48 .

( 4 ) OJ NO L 359 , 15 . 12 . 1981 , P . 6 .

( 5 ) OJ NO L 360 , 22 . 12 . 1978 , P . 1 .

COMMISSION REGULATION ( EEC ) NO 1108/82 OF 21 APRIL 1982 DETERMINING COMMUNITY METHODS FOR THE ANALYSIS OF WINES AND REPEALING REGULATION ( EEC ) NO 2984/78

THE COMMISSION OF THE EUROPEAN COMMUNITIES ,

HAVING REGARD TO THE TREATY ESTABLISHING THE EUROPEAN ECONOMIC COMMUNITY ,

HAVING REGARD TO COUNCIL REGULATION ( EEC ) NO 337/79 OF 5 FEBRUARY 1979 ON THE COMMON ORGANIZATION OF THE MARKET IN WINE ( 1 ) , AS LAST AMENDED BY REGULATION ( EEC ) NO 3577/81 ( 2 ) , AND IN PARTICULAR ARTICLE 63 THEREOF ,

HAVING REGARD TO COUNCIL REGULATION ( EEC ) NO 338/79 OF 5 FEBRUARY 1979 LAYING DOWN SPECIAL PROVISIONS RELATING TO QUALITY WINES PRODUCED IN SPECIFIED REGIONS ( 3 ) AS LAST AMENDED BY REGULATION ( EEC ) NO 3578/81 , AND IN PARTICULAR ARTICLE 14 ( 3 ) THEREOF ,

WHEREAS ARTICLE 63 ( 2 ) OF REGULATION ( EEC ) NO 337/79 PRESCRIBES THE ADOPTION OF METHODS OF ANALYSIS FOR ESTABLISHING THE COMPOSITION OF THE PRODUCTS INDICATED IN ARTICLE 1 OF THAT REGULATION AND OF RULES FOR CHECKING WHETHER THESE PRODUCTS HAVE BEEN SUBJECTED TO TREATMENTS IN VIOLATION OF AUTHORIZED OENOLOGICAL PRACTICE ;

WHEREAS ARTICLE 14 ( 1 ) OF REGULATION ( EEC ) NO 338/79 PROVIDES FOR AN ANALYTICAL TEST , INCLUDING AT THE LEAST AN ASSESSMENT OF THE CHARACTERISTICS , AS LISTED IN THE ANNEX TO THAT REGULATION , OF THE QUALITY WINE PSR IN QUESTION ;

WHEREAS THE VERIFICATION OF THE PARTICULARS ON DOCUMENTS CONCERNING THE PRODUCTS IN QUESTION CALLS FOR THE INTRODUCTION OF UNIFORM METHODS OF ANALYSIS TO ENSURE THAT ACCURATE AND COMPARABLE INFORMATION IS OBTAINED ; WHEREAS , CONSEQUENTLY , THESE METHODS SHOULD BE COMPULSORY FOR ALL COMMERCIAL TRANSACTIONS AND ALL VERIFICATION PROCEDURES ; WHEREAS , HOWEVER , IN VIEW OF CONTROL REQUIREMENTS AND THE TRADE'S LIMITED FACILITIES , A RESTRICTED NUMBER OF USUAL PROCEDURES SHOULD BE ADMITTED ENABLING THE REQUISITE FACTORS TO BE DETERMINED RAPIDLY AND WITH REASONABLE ACCURACY ;

WHEREAS , TO THE EXTENT POSSIBLE , GENERALLY RECOGNIZED METHODS SUCH AS THOSE DEVELOPED UNDER THE 1954 INTERNATIONAL CONVENTION FOR THE UNIFICATION OF METHODS OF ANALYSIS AND APPRAISAL OF WINES , WHICH ARE PUBLISHED IN THE RECUEIL DES METHODES INTERNATIONALES D'ANALYSE DES VINS ( COMPENDIUM OF INTERNATIONAL METHODS FOR THE ANALYSIS OF WINES ) BY THE INTERNATIONAL VINE AND WINE OFFICE , MAY USEFULLY BE RETAINED ; WHEREAS , HOWEVER , THE TEXTS IN QUESTION HAVE BEEN PUBLISHED IN ONE LANGUAGE ONLY ; WHEREAS ALL THE METHODS OF ANALYSIS RECOGNIZED BY COMMUNITY LAW SHOULD BE PUBLISHED IN ALL THE OFFICIAL LANGUAGES OF THE COMMUNITY IN ORDER TO HELP THOSE CONCERNED GAIN A PROPER UNDERSTANDING THEREOF ;

WHEREAS THE COMMUNITY METHODS OF ANALYSIS APPLICABLE TO WINE ARE LAID DOWN IN REGULATION ( EEC ) NO 2984/78 ( 5 ) ;

WHEREAS SCIENTIFIC PROGRESS HAS MADE IT POSSIBLE TO REPLACE A METHOD FOR ANALYZING CARBON DIOXIDE WHICH IS SELDOM USED BY A PROVEN METHOD ; WHEREAS CERTAIN ERRORS IN THE METHODS OF ANALYSIS PUBLISHED SHOULD BE CORRECTED AND THE PRESENTATION OF THE METHODS SHOULD IN SOME CASES BE IMPROVED ; WHEREAS THE TEXT HAS BEEN FOUND TO CONTAIN SOME CLERICAL ERRORS ; WHEREAS , ACCORDINGLY , ALL THE COMMUNITY METHODS OF ANALYSIS SHOULD BE PUBLISHED AGAIN AND REGULATION ( EEC ) NO 2984/78 SHOULD BE REPEALED :

WHEREAS THE MEASURES PROVIDED FOR IN THIS REGULATION ARE IN ACCORDANCE WITH THE OPINION OF THE MANAGEMENT COMMITTEE FOR WINE ,

HAS ADOPTED THIS REGULATION :

ARTICLE 1

1 . THE ANNEX TO THIS REGULATION SETS OUT COMMUNITY METHODS WHICH MAKE IT POSSIBLE , IN THE CONTEXT OF COMMERCIAL TRANSACTIONS AND ALL CONTROL OPERATIONS , TO :

- ESTABLISH THE COMPOSITION OF THE PRODUCTS LISTED IN ARTICLE 1 OF REGULATION ( EEC ) NO 337/79 ,

- CHECK WHETHER THESE PRODUCTS HAVE BEEN SUBJECTED TO TREATMENTS IN VIOLATION OF AUTHORIZED OENOLOGICAL PRACTICE .

2 . FOR PRODUCTS FOR WHICH REFERENCE METHODS AND CUSTOMARY METHODS ARE LAID DOWN , THE RESULTS OBTAINED BY THE APPLICATION OF THE REFERENCE METHODS SHALL PREVAIL .

3 . THE METHODS OF ANALYSIS REFERRED TO IN ARTICLE 63 ( 3 ) OF REGULATION ( EEC ) NO 337/79 MAY BE USED FOR THE ANALYSIS OF SUBSTANCES FOR WHICH NO METHOD OF ANALYSIS IS SPECIFIED IN THE ANNEX TO THIS REGULATION OR FOR SUPPLEMENTING OR CONFIRMING THE RESULTS GIVEN BY THE REFERENCE METHOD .

4 . UNTIL COMMUNITY PROVISIONS ARE ADOPTED ON THE LEVELS OF THE CONSTITUENTS WHICH INDICATE THAT CERTAIN OENOLOGICAL PRACTICES HAVE BEEN EMPLOYED AND ON TABLES ENABLING A COMPARISON OF THE ANALYSIS DATA , THE MEMBER STATES MAY , IN ORDER TO ASSESS WHETHER A PRODUCT HAS BEEN SUBJECTED TO TREATMENTS OR PRACTICES WHICH ARE NOT IN ACCORDANCE WITH COMMUNITY PROVISIONS :

- DETERMINE THE LEVELS OF CONSTITUENTS IN THAT PRODUCT ,

- STIPULATE THE USE OF TABLES ENABLING A COMPARISON OF ANALYSIS DATA .

5 . WHEREVER MENTION IS MADE OF WATER FOR SOLUTION , DILUTION OR WASHING PURPOSES , THIS SHALL MEAN DISTILLED WATER , OR DEMINERALIZED WATER OF EQUIVALENT PURITY .

ALL CHEMICALS SHALL BE OF ANALYTICAL REAGENT QUALITY EXCEPT WHERE OTHERWISE SPECIFIED .

ARTICLE 2

REGULATION ( EEC ) NO 2984/78 IS HEREBY REPEALED .

ARTICLE 3

THIS REGULATION SHALL ENTER INTO FORCE ON 1 JUNE 1982 .

THIS REGULATION SHALL BE BINDING IN ITS ENTIRETY AND DIRECTLY APPLICABLE IN ALL MEMBER STATES .

DONE AT BRUSSELS , 21 APRIL 1982 .

FOR THE COMMISSION

POUL DALSAGER

MEMBER OF THE COMMISSION

ANNEX

1 . DENSITY AND SPECIFIC GRAVITY AT 20* C

1.1 . DEFINITIONS

THE DENSITY AND THE SPECIFIC GRAVITY OF WINE AND OF MUST ARE DETERMINED AT 20* C .

THE DENSITY ( * ) IS THE MASS PER UNIT VOLUME OF WINE OR MUST AT 20* C . IT IS EXPRESSED IN GRAMS PER CUBIC CENTIMETRE ( 1 ) , SYMBOLIZED BY * .

THE SPECIFIC GRAVITY , * OR D , IS THE RATIO OF THE DENSITY OF WINE ( OR OF MUST ) AT 20* C TO THE DENSITY OF WATER AT THE SAME TEMPERATURE .

CONVENTIONALLY , THE RELATIVE DENSITY AND THE SPECIFIC GRAVITY SHALL BE SUBJECT TO CORRECTION FOR THE EFFECT OF SULPHUR DIOXIDE AND ANY PRESERVATIVE ADDED AT THE TIME OF SAMPLING FOR STABILIZING THE WINE OR MUST SAMPLE .

1.2 . PRINCIPLE OF METHODS

REFERENCE METHOD : MEASUREMENT BY PYCNOMETRY , RESULTS ACCURATE TO 0.0001 .

FIRST USUAL METHOD : MEASUREMENT BY HYDROMETRY , RESULTS ACCURATE TO 0.0003 .

SECOND USUAL METHOD : MEASUREMENT BY DENSIMETRY BY THE HYDROSTATIC BALANCE , RESULTS ACCURATE TO 0.0003 .

PRELIMINARY TREATMENT

ANY EXCESS CARBON DIOXIDE IN THE WINE OR MUST MAY BE REMOVED BY STIRRING 250 ML IN A ONE LITRE FLASK OR BY FILTRATION AT REDUCED PRESSURE THROUGH 2 G OF COTTON WOOL .

IF THE WINE OR MUST IS CLOUDY IT SHOULD EITHER BE FILTERED OR CENTRIFUGED IN SEALED TUBES . THE USE OF EITHER PROCEDURE SHOULD BE STATED ON THE CERTIFICATE OF ANALYSIS .

1.3 . REFERENCE METHOD - MEASUREMENT BY A PYCNOMETER - APPARATUS

A PYREX PYCNOMETER , OF APPROXIMATELY 100 ML CAPACITY , WITH A GROUND GLASS JOINT THERMOMETER CALIBRATED IN 10THS OF A DEGREE FROM 10 TO 30* C , IS USED .

THE PYCNOMETER COMPRISES A SIDE TUBE OF 15 MM LENGTH AND 1 MM ( MAXIMUM ) INSIDE DIAMETER ENDING IN A CONICAL GROUND JOINT . THIS SIDE TUBE MAY BE CAPPED BY A " RECEIVER PLUG " CONSISTING OF A CONICAL GROUND JOINT TUBE TERMINATING IN A DRAWN-OUT PORTION . THIS PLUG SERVES AS AN EXPANSION CHAMBER . THE THERMOMETER SHOULD BE CAREFULLY CHECKED AND THE TWO GROUND JOINTS OF THE APPARATUS SHOULD BE PREPARED WITH GREAT CARE . THEY MAY BE GREASED LIGHTLY WITH A TRACE OF SILICONE .

A VESSEL OF THE SAME OUTSIDE VOLUME AS THE PYCNOMETER IS USED AS A TARE . IT IS FILLED ALMOST COMPLETELY WITH A SOLUTION OF A METAL SALT SUCH AS SODIUM CHLORIDE , POTASSIUM CHROMATE , ETC . , OF A SPECIFIC GRAVITY SUCH THAT THE MASS OF THIS STANDARD IS EQUAL TO THE MASS OF THE PYCNOMETER FILLED WITH A LIQUID OF DENSITY 1.01 ( 2 ) . THIS VESSEL IS SEALED IN A FLAME . ITS OUTSIDE CAPACITY SHOULD BE EQUAL TO THE OUTSIDE CAPACITY OF THE FILLED PYCNOMETER TO WITHIN 1 ML .

TARING OF THE PYCNOMETER

THE EMPTY PYCNOMETER SHALL BE TARED USING A BALANCE WITH A SENSITIVITY OF 0.1 MG AND A MAXIMUM RANGE OF AT LEAST 200 G , USING PRECISION WEIGHTS .

ADD THE MASS OF AIR CONTAINED IN THE EMPTY AND DRY PYCNOMETER ( 3 ) TO THE WEIGHTS , P , PLACED BESIDE THE PYCNOMETER . THE CORRECTED TARE OF THE EMPTY PYCNOMETER IS THUS OBTAINED .

1.2 G PER LITRE MAY BE ASSUMED AS THE DENSITY OF AIR ( 4 ) FOR THE CALCULATION OF THE MASS OF AIR M CONTAINED IN THE PYCNOMETER .

THE MASS OF THE AIR M CONTAINED IN THE APPARATUS IS OBTAINED BY MULTIPLYING THE VOLUME OF THE PYCNOMETER IN MILLILITRES BY 0.0012 . AS IT IS NOT NECESSARY TO KNOW M WITH AN ACCURACY GREATER THAN TO THE NEAREST 0.5 MG , THIS VOLUME IS NUMERICALLY EQUAL TO P - P' , P' BEING THE WEIGHT REQUIRED TO BALANCE THE PYCNOMETER FILLED WITH WATER .

TARE THE WATER-FILLED PYCNOMETER AT T* C , T BEING THE TEMPERATURE AT WHICH THE OPERATION IS CARRIED OUT AS SHOWN ON THE PYCNOMETER THERMOMETER , MEASURED TO 0.02* C , AND P' BEING THE WEIGHT IN GRAMS REQUIRED TO ESTABLISH EQUILIBRIUM .

CALCULATE THE VOLUME OF THE PYCNOMETER AT 20* C ( 5 ) BY MULTIPLYING THE MASS OF WATER AT T* C CONTAINED IN THE PYCNOMETER AT T* C BY THE FACTOR F GIVEN IN TABLE I . THIS TABLE GIVES THE RECIPROCALS OF THE DENSITIES OF WATER AT T* C CORRECTED FOR THE EXPANSION OF THE PYREX PYCNOMETER BETWEEN T AND 20* C .

THE KNOWLEDGE OF THIS VOLUME IS NECESSARY FOR THE MEASUREMENT OF THE DENSITIES OF LIQUIDS .

IF THE SPECIFIC GRAVITY IS TO BE CALCULATED , IT IS NECESSARY TO KNOW THE MASS OF THE WATER AT 20* C CONTAINED IN THE PYCNOMETER AT 20* C . THIS IS OBTAINED BY MULTIPLYING THE VOLUME AT 20* C BY 0.998203 , THE DENSITY OF WATER AT 20* C .

THE CALIBRATION OPERATION SHOULD BE CARRIED OUT IN TRIPLICATE TO CHECK THAT THERE IS NO ERROR IN THIS FUNDAMENTAL OPERATION ( 6 ) .

THE TARE OF THE EMPTY PYCNOMETER AND THE VOLUME AT 20* C ( AND CONSEQUENTLY THE MASS OF THE WATER AT 20* C ) ARE CHARACTERISTIC OF THE EQUIPMENT . BECAUSE A TARE OF THE SAME VOLUME AS THE FULL PYCNOMETER IS USED IT IS NOT NECESSARY TO DETERMINE THEM AT EACH MEASUREMENT . IT IS RECOMMENDED , HOWEVER , TO CHECK THEM PERIODICALLY E . G . ANNUALLY .

TABLE I

F FACTORS

BY WHICH THE MASS OF THE WATER CONTAINED IN THE PYREX PYCNOMETER AT T* C HAS TO BE MULTIPLIED TO CALCULATE THE VOLUME OF THE PYCNOMETER AT 20* C ( SEE OJ NO L 133 OF 14 . 5 . 1982 ) .

PROCEDURE

CAREFULLY FILL THE PYCNOMETER WITH THE LIQUID AT AMBIENT TEMPERATURE AND PLACE IN THE THERMALLY INSULATED ENCLOSURE ; SHAKE THE LIQUID TWO OR THREE TIMES BY INVERTING THE CONTAINER UNTIL THE TEMPERATURE READING ON THE THERMOMETER IS CONSTANT . ACCURATELY ADJUST THE LEVEL TO THE UPPER RIM OF THE LATERAL TUBE . WIPE THE LATERAL TUBE DRY , PUT THE RECEIVER PLUG ON AND MEASURE THE TEMPERATURE T* C WITH THE SAME CARE AS PREVIOUSLY ; CAREFULLY WIPE THE PYCNOMETER DRY AND THEN WEIGH , P'' BEING THE WEIGHT TO GIVE EQUILIBRIUM .

THE DIFFERENCE BETWEEN THE CORRECTED TARE AND THE WEIGHT RECORDED GIVES THE MASSES OF THE LIQUID IN THE APPARATUS ( CORRECTED FOR BUOYANCY OF THE AIR ) .

CALCULATE THE APPARENT DENSITY AT T* C BY DIVIDING THIS MASS BY THE VOLUME AT 20* C , AND THE APPARENT SPECIFIC GRAVITY * BY DIVIDING THIS SAME MASS BY THE MASS OF THE WATER CONTAINED AT 20* C IN THE PYCNOMETER .

1 . CORRECTION FOR TEMPERATURE

CORRECT * AND * USING ONE OF THE FOLLOWING CORRECTION TABLES IN ACCORDANCE WITH THE NATURE OF THE LIQUID MEASURED : DRY WINE ( TABLE II ) , MUST OR CONCENTRATED MUST ( TABLE III ) , WINE CONTAINING RESIDUAL SUGARS ( TABLE IV ) , ALCOHOL-FREE RESIDUE ( WINE FREED OF ALCOHOL BY DISTILLATION AND THE RESIDUE FROM THE DISTILLATION MADE UP TO THE ORIGINAL VOLUME ) ( TABLE II ) . IN THE COMPILATION OF THESE TABLES THE EXPANSION OF THE LIQUIDS AND OF THE CONTAINER BETWEEN T AND 20* C HAS BEEN TAKEN INTO ACCOUNT .

( TABLES : SEE OJ )

2 . CORRECTIONS MADE NECESSARY BY THE PRESENCE OF PRESERVATIVES

THE DENSITY MUST SUBSEQUENTLY BE CORRECTED FOR THE EFFECT OF SULPHUR DIOXIDE BY USING THE FORMULA : ( SEE OJ ) .

1.4 . FIRST USUAL METHOD

MEASUREMENT BY A HYDROMETER - APPARATUS

THE APPARATUS USED SHALL COMPLY WITH THE FOLLOWING CHARACTERISTICS .

THE APPARATUS SHALL HAVE A CYLINDRICAL BULB , A STEM OF CIRCULAR SECTION OF MINIMUM DIAMETER 8 MM . FOR DRY WINES THE APPARATUS SHALL BE GRADUATED FROM 0.9830 TO 1.0030 WITH GRADUATION MARKS EVERY 0.0010 AND 0.0002 . EACH MARK AT 0.0010 SHALL BE SEPARATED BY AT LEAST 5 MM FROM THE NEXT CORRESPONDING MARK . FOR THE MEASUREMENT OF THE SPECIFIC GRAVITY OF DE-ALCOHOLIZED WINES , SWEET WINES AND MUSTS , A SET OF FIVE HYDROMETERS GRADUATED FROM 1.0000 TO 1.0300 , 1.0300 TO 1.0600 , 1.0600 TO 1.0900 , 1.0900 TO 1.1200 AND 1.1200 TO 1.1500 SHALL BE USED . THESE HYDROMETERS SHALL BE GRADUATED IN DENSITIES AT 20* C WITH MARKS AT 0.0010 AND 0.0005 , EACH MARK AT 0.0010 BEING SEPARATED BY AT LEAST 3 MM FROM THE NEXT CORRESPONDING MARK .

THESE HYDROMETERS SHALL BE GRADUATED SO AS TO BE READ AT THE " TOP OF THE MENISCUS " . TO AVOID ANY CONFUSION WITH HYDROMETERS GRADUATED AT 15* C AND NORMALLY READ AT THE LIQUID SURFACE , AN INDICATION OF THE GRADUATION IN DENSITY AT 20* C AND OF THE READING AT THE " TOP OF THE MENISCUS " SHALL BE CARRIED EITHER ON THE GRADUATED SCALE OR ON A STRIP OF PAPER ENCLOSED IN THE BULB .

THESE HYDROMETERS SHALL BE CALIBRATED BY A GOVERNMENT DEPARTMENT .

THE RESULT SHALL BE EXPRESSED TO FOUR DECIMAL PLACES TO THE NEAREST THREE UNITS OF THE FOURTH DECIMAL PLACE ( MORE OR LESS 0.0003 ) .

A CALIBRATED THERMOMETER GRADUATED IN MINIMUM STEPS OF 0.5* C SHALL BE USED .

DETERMINATION OF DENSITY - PROCEDURE

POUR 250 ML OF WINE INTO A LEVEL GLASS CYLINDER OF 36 MM INTERNAL DIAMETER AND 320 MM HEIGHT . PLACE A HYDROMETER AND THERMOMETER IN THE CYLINDER .

READ THE OBSERVED TEMPERATURE T* C ONE MINUTE AFTER MIXING THE SAMPLE WELL . REMOVE THE THERMOMETER AND READ THE APPARENT DENSITY FROM THE HYDROMETER AFTER A FURTHER MINUTE .

THE DENSITY AT 20* C IS THEN CALCULATED USING TABLE V , VI OR VII ( 7 ) . ( SEE OJ )

CORRECT THE DENSITY FOR THE EFFECT OF ADDED SULPHUR DIOXIDE AND OTHER PRESERVATIVES IN THE SAME MANNER AS DESCRIBED FOR THE REFERENCE METHOD .

1.5 . SUPPLEMENT TO THE REFERENCE METHOD - MEASUREMENT BY PYCNOMETER ON SINGLE-PAN BALANCE

PRINCIPLE

THE REFERENCE METHOD FOR THE DETERMINATION OF THE DENSITY OF MUSTS AND WINES AND FOR THE DETERMINATION OF THE ALCOHOLIC STRENGTH OF WINES REQUIRES THE USE OF A TWO-PAN BALANCE AND A PYCNOMETER WITH ITS TARE BOTTLE OF THE SAME EXTERNAL VOLUME TO ELIMINATE THE EFFECT OF AIR . THIS SIMPLE TECHNIQUE MAY ALSO BE APPLIED WHEN A SINGLE-PAN BALANCE IS USED SUBJECT TO A SUPPLEMENTARY WEIGHING OF THE TARE BOTTLE TO TAKE ACCOUNT OF THE VARIATIONS OF THE BUOYANCY OF AIR :

- WHEN ESTABLISHING THE PYCNOMETER CONSTANTS : WEIGHT OF THE EMPTY PYCNOMETER , VOLUME AT 20* C , THE WEIGHT T0 OF THE TARE BOTTLE IS DETERMINED ,

- WHEN DETERMINING DENSITY , THE WEIGHT T1 OF THE TARE BOTTLE IS AGAIN DERIVED AND THE WEIGHT OF THE PYCNOMETER FILLED WITH LIQUID IS CORRECTED FOR THE MATHEMATICAL VALUE OF THE DIFFERENCE BETWEEN T0 AND T1 . THE REMAINDER OF THE CALCULATIONS ARE UNCHANGED .

PROCEDURE

1 . STANDARDIZATION OF THE PYCNOMETER

ESTABLISH THE FOLLOWING CONSTANT CHARACTERISTICS OF THE PYCNOMETER :

- ITS WEIGHT EVACUATED , P0 ,

- ITS VOLUME AT 20* C , V20 ,

- AT THE SAME TIME , RECORD ACCURATELY THE WEIGHT OF THE TARE BOTTLE T0 .

THE WEIGHT OF THE TARE BOTTLE T0 IS OBTAINED BY WEIGHING TO THE NEAREST 0.1 MG .

THE WEIGHT OF THE PYCNOMETER EVACUATED , P0 , IS THE WEIGHT P OF THE CLEAN AND DRY PYCNOMETER , FILLED WITH AIR LESS THE MASS M OF THE AIR WHICH IT CONTAINS . OBTAIN THE MASS M OF THE AIR BY MULTIPLYING THE DENSITY OF THE AIR BY THE VOLUME OF THE PYCNOMETER . UNDER AVERAGE CONDITIONS OF TEMPERATURE AND PRESSURE ASSUME A VALUE OF 0.0012 G/ML FOR THE DENSITY OF AIR . FOR THE CALCULATION OF M , CONSIDER THAT THE VOLUME OF THE PYCNOMETER IS NUMERICALLY EQUAL TO P1 - P , P1 BEING THE WEIGHT OF THE PYCNOMETER FILLED WITH WATER OBTAINED AT THE TIME OF THE DETERMINATION OF THE WEIGHT OF WATER IN THE PYCNOMETER AT T* C .

HENCE : P0 = P - M , WHERE M = 0.0012 ( P1 - P ) .

TO OBTAIN THE VOLUME AT 20* C OF THE PYCNOMETER , DETERMINE THE WEIGHT P1 OF THE PYCNOMETER , FILLED UP TO THE MARK WITH DEGASSED WATER AT T* C . THIS TEMPERATURE MUST BE MEASURED TO 0.02* C .

THE MASS MT OF THE WATER AT T* C CONTAINED IN THE PYCNOMETER IS EQUAL TO :

MT = P1 - P0

MULTIPLY MT BY THE FACTOR F ( PRODUCT OF THE RECIPROCAL OF THE DENSITY OF THE WATER AT T* C AND THE RATIO OF THE VOLUMES OF THE PYCNOMETER AT T* C AND AT 20* C ) GIVEN IN TABLE I ( PAGE 5 ) TO OBTAIN THE VOLUME OF THE PYCNOMETER AT 20* C :

V20 = F ( P1 - P0 )

2 . DETERMINATION OF A DENSITY

WEIGH THE TARE BOTTLE TO THE NEAREST 0.1 MG ; LET THIS WEIGHT BE T1 .

FILL THE PYCNOMETER WITH LIQUID ( MUST , WINE OR DISTILLATE ) AT T* C AND WEIGH ; LET THIS BE P2 .

CALCULATE THE DIFFERENCE DT = T1 - T0 ; THIS REPRESENTS THE VARIATION IN THE BUOYANCY OF AIR .

THE MASS LT OF THE LIQUID CONTAINED IN THE PYCNOMETER AT T* C IS EQUAL TO LT = P2 - ( P0 + DT ) .

NOTE MUST BE TAKEN OF THE SIGN OF DT . THE APPARENT DENSITY * IS EQUAL TO THE RATIO OF LT TO V20 .

ONCE THE APPARENT DENSITY AT T* C IS KNOWN , THE DENSITY AT 20* C OF THE LIQUID UNDER TEST ( DRY WINE , MUST OR CONCENTRATED MUST , SWEET WINE CONTAINING RESIDUAL SUGARS , ALCOHOL-FREE RESIDUE ) IS DETERMINED AS SPECIFIED ON PAGE 6 .

NUMERICAL EXAMPLE

1 . STANDARDIZATION OF THE PYCNOMETER :

WEIGHT OF TARE BOTTLE : T0 = 171.9160

WEIGHT OF PYCNOMETER FILLED WITH AIR : P = 67.7913

WEIGHT OF PYCNOMETER FILLED WITH WATER AT 21.65* C : P1 = 169.2715

MASS OF AIR CONTAINED IN THE PYCNOMETER : M = 0.0012 ( 169.2715 - 67.7913 ) = 0.1218

WEIGHT OF EMPTY PYCNOMETER : P0 = 67.7913 - 0.1218 = 67.6695

VOLUME OF THE PYCNOMETER AT 20* C : V20 = 1.002140 ( 169.2715 - 67.6695 ) = 101.8194 ML OR 0.1018194 DM3

2 . DETERMINATION OF THE APPARENT DENSITY OF A WINE :

WEIGHT OF TARE BOTTLE AT THE TIME OF THE DETERMINATION : T1 = 171.9178

WEIGHT OF PYCNOMETER FULL OF WINE AT 18* C : P2 = 169.2799

VARIATION IN THE BUOYANCY OF AIR : DT = 171.9178 - 171.9160 = + 0.0018

MASS OF THE WINE AT 18* C : LT = 169.2799 - ( 67.6695 + 0.0018 ) = 101.6086

APPARENT DENSITY OF THE WINE AT 18* C : *

3 . CALCULATION OF THE DENSITY OF A WINE AT 20* C :

PROCEED AS SPECIFIED ON PAGE 5 .

SECOND USUAL METHOD

DENSIMETRY BY THE HYDROSTATIC BALANCE

APPARATUS AND CALIBRATION OF THE FLOAT

THE HYDROSTATIC BALANCE , OF MAXIMUM CAPACITY NOT LESS THAN 100 G , MUST HAVE A SENSITIVITY OF 0.1 MG .

UNDER EACH PAN IS FIXED A PYREX ( 8 ) GLASS FLOAT OF VOLUME AT LEAST 20 ML . A VOLUME OF 50 OR 100 ML IS PREFERABLE . THESE TWO IDENTICAL FLOATS MUST BE SUSPENDED BY A THREAD OF DIAMETER NOT EXCEEDING 0.1 MM .

THE FLOAT SUSPENDED BENEATH THE RIGHT-HAND PAN MUST BE ABLE TO BE INTRODUCED INTO A CYLINDRICAL TEST-TUBE BEARING A MARK INDICATING THE LEVEL . THIS TEST-TUBE MUST HAVE AN INTERNAL DIAMETER AT LEAST 6 MM LARGER THAN THAT OF THE FLOAT . THE FLOAT MUST BE CAPABLE OF BEING CONTAINED COMPLETELY IN THE VOLUME OF THE TEST-TUBE SITUATED BELOW THE MARK ; THE SURFACE OF THE LIQUID TO BE MEASURED MUST BE TRAVERSED ONLY BY THE SUSPENSION THREAD . THE TEMPERATURE OF THE LIQUID PRESENT IN THE TEST-TUBE IS MEASURED BY A THERMOMETER GRADUATED IN DIVISIONS OF 0.2* C .

CALIBRATION OF THE APPARATUS

WITH THE TWO FLOATS IN AIR ONE FINDS THE BALANCED POSITION BY PLACING WEIGHTS MARKED P ON THE RIGHT-HAND PAN .

THE TEST-TUBE IS FILLED WITH PURE WATER UP TO THE MARK AND THE TEMPERATURE , T* C , IS MEASURED AFTER SHAKING AND ALLOWING TO STAND FOR TWO TO THREE MINUTES . THE BALANCED POSITION IS FOUND ONCE AGAIN BY MEANS OF MARKED WEIGHTS PLACED ON THE RIGHT-HAND PAN . LET THESE WEIGHTS BE P' .

THE VOLUME ( 9 ) OF THE FLOAT AT 20* C IS EQUAL TO V20 = ( P' - P ) ( F + 0.0012 ) ;

WHERE :

F IS THE FACTOR GIVEN IN TABLE I ,

P AND V20 ARE THE CHARACTERISTICS OF THE FLOAT , BECAUSE THEY DO NOT VARY WITH THE AIR PRESSURE , WHICH IS COMPENSATED FOR BY THE CALIBRATED FLOAT OF THE LEFT-HAND PAN .

PROCEDURE

THE RIGHT-HAND FLOAT IS IMMERSED IN THE TEST-TUBE FILLED WITH WINE ( OR MUST ) UP TO THE MARK . LET T* C BE THE TEMPERATURE OF THE WINE ( OR MUST ) AND P'' BE THE MARKED WEIGHTS WHICH RE-ESTABLISH THE BALANCED POSITION . THE APPARENT DENSITY , * , IS GIVEN BY THE FORMULA : ( SEE OJ )

THIS DENSITY IS CORRECTED TO 20* C BY USING ONE OF TABLES II TO IV , IF THE FLOAT IS OF PYREX GLASS , OR ONE OF TABLES V TO VII IF THE FLOAT IS OF ORDINARY GLASS .

THE USE OF AN AUTOMATIC HYDROSTATIC BALANCE IS AUTHORIZED IF THIS LEADS TO MEASUREMENT OF THE DENSITY AT 20* C WITH THE SAME ACCURACY .

2 . ALCOHOLIC STRENGTH BY VOLUME ( DEGREES ALCOHOL BY VOLUME )

2.1 . DEFINITION

THE ALCOHOLIC STRENGTH BY VOLUME IS EXPRESSED BY THE SYMBOL " % VOL " . IT IS EQUAL TO THE NUMBER OF LITRES OF ETHANOL ( 10 ) IN 100 LITRES OF PRODUCT AT A TEMPERATURE OF 20* C ( 11 ) .

2.2 . PRINCIPLE OF METHODS

1 . FOR WINES :

( A ) REFERENCE METHOD : DOUBLE DISTILLATION AND MEASUREMENT OF THE SPECIFIC GRAVITY OF THE DISTILLATE WITH A PYCNOMETER . STRENGTH EXPRESSED TO THE NEAREST 0.05 % VOL ;

( B ) USUAL METHODS :

- FIRST USUAL METHOD : SINGLE DISTILLATION OF THE SAMPLE WHICH HAS BEEN MADE ALKALINE AND MEASUREMENT OF ALCOHOLIC STRENGTH WITH A HYDROMETER : THE ALCOHOLIC STRENGTH TO BE EXPRESSED TO THE NEAREST 0.1 % VOL ,

- SECOND USUAL METHOD : SINGLE DISTILLATION OF THE SAMPLE WHICH HAS BEEN MADE ALKALINE AND MEASUREMENT OF ALCOHOLIC STRENGTH BY THE HYDROSTATIC BALANCE : THE ALCOHOLIC STRENGTH TO BE EXPRESSED TO THE NEAREST 0.1 % VOL .

2 . FOR MUSTS AND FERMENTING MUSTS WITH AN ALCOHOLIC STRENGTH NOT EXCEEDING 6 % VOL :

( A ) REFERENCE METHOD : MEASUREMENT OF ALCOHOL BY CHEMICAL MEANS ;

( B ) USUAL METHODS : THE METHODS SHALL BE THOSE GIVEN UNDER 1 ( B )

2.3 . REFERENCE METHOD FOR WINES

EQUIPMENT

1 . DISTILLATION EQUIPMENT : THIS CONSISTS OF A 1 000 ML ROUND-BOTTOMED FLASK WITH GROUND JOINTS , FITTED WITH A RECTIFYING COLUMN OF 20 CM LENGTH , OR , IF NOT AVAILABLE , A KJELDAHL BULB TO PREVENT SPLASHING . TO AVOID ANY OVERHEATING OF THE VOLATILE MATERIAL , THE FLASK , WHICH SHOULD BE HEATED BY GAS , SHOULD BE PLACED ON A PLATE PROVIDED WITH A HOLE OF 8 CM DIAMETER . THE VOLATILE SUBSTANCES ARE PASSED INTO A WEST ( 12 ) CONDENSER PLACED VERTICALLY . THE DISTILLATE IS DIRECTED TOWARDS THE BOTTOM OF THE RECEIVING FLASK USING A DRAWN-OUT TUBE . IN SUMMER THIS FLASK SHOULD BE IMMERSED IN ICE-WATER .

ANY OTHER TYPE OF DISTILLATION APPARATUS AND ANY OTHER METHOD OF COLLECTION OF THE VOLATILE ALCOHOLIC MATERIAL MAY BE USED , OR THE ALCOHOL MAY BE STEAM-DISTILLED , PROVIDED THE APPARATUS USED CORRESPONDS TO THE FOLLOWING TEST : 200 ML OF A 10 % VOL ETHANOL-WATER MIXTURE DISTILLED FIVE TIMES IN SUCCESSION SHOULD HAVE AN ALCOHOLIC STRENGTH OF AT LEAST 9.9 % VOL AFTER THE LAST DISTILLATION , I . E . THE LOSS OF ALCOHOL OCCURRING DURING EACH DISTILLATION MUST NOT BE GREATER THAN 0.02 % VOL .

2 . PYCNOMETER : THIS PIECE OF APPARATUS IS IDENTICAL WITH THAT SPECIFIED FOR THE MEASUREMENT OF THE DENSITY ; THE TARE SHALL HAVE A WEIGHT WHICH IS JUST GREATER ( NO MORE THAN 1 OR 2 G ) THAN THAT OF THE PYCNOMETER FILLED WITH WATER . IT SHALL BE STANDARDIZED AS SPECIFIED FOR THAT MEASUREMENT IN " DENSITY AND SPECIFIC GRAVITY " ( PAGE 3 ) .

PROCEDURE

REMOVE THE BULK OF ANY CARBON DIOXIDE FROM YOUNG AND SPARKLING WINES BY STIRRING 250 ML OF THE WINE IN A 500 ML BOTTLE WHICH HAS BEEN PREVIOUSLY TREATED INTERNALLY WITH THREE DROPS OF A 1 % SILICON SOLUTION AND DRIED .

1 . DISTILLATION : MEASURE OUT THE WINE TO BE ANALYZED INTO A 200 ML GRADUATED FLASK WITH A NECK OF MAXIMUM 12 MM DIAMETER AND RECORD ITS TEMPERATURE . SMALLER OR LARGER VOLUMES OF WINE MAY BE USED , BUT AT LEAST EQUAL TO 150 ML .

TRANSFER THE WINE TO THE DISTILLATION FLASK INTO WHICH HAS BEEN PLACED SOME INERT POROUS MATERIAL ( UNGLAZED PORCELAIN , GLASS FRIT , PUMICE , ETC . ) . RINSE THE FLASK FOUR TIMES WITH 5 ML OF WATER . ADD 10 ML OF MILK OF LIME ( 120 G OF CAO PER LITRE ) . THE COLOURING MATTER OF THE WINE SHOULD BE CHANGED BY THE ALKALINITY . IN THE CASE OF VERY ACID , SOUR , ETC . , WINES ADD THE LIME UNTIL THE WINE IS CLEARLY ALKALINE TO PHENOLPHTHALEIN USED EXTERNALLY . THE DISTILLATE IS COLLECTED IN A 200 ML FLASK , TO WHICH APPROXIMATELY 10 ML OF PURE WATER HAS BEEN PREVIOUSLY ADDED AND INTO WHICH SHOULD DIP THE DRAWN-OUT TUBE CONNECTED TO THE CONDENSER .

A VOLUME EQUAL TO APPROXIMATELY THREE QUARTERS OF THE INITIAL VOLUME SHOULD BE DISTILLED AND COLLECTED .

REDISTILL THE DISTILLATE AFTER 1 ML OF 10 % SULPHURIC ACID AND SOME FRAGMENTS OF DRY INERT POROUS MATERIAL HAVE BEEN ADDED TO THE FLASK AND THE FLASK HAS BEEN RINSED FOUR MORE TIMES WITH 5 ML OF WATER .

COLLECT THE SECOND DISTILLATE IN THE SAME FLASK WHICH WAS USED FOR MEASURING THE VOLUME OF THE WINE AND INTO WHICH 10 ML OF DISTILLED WATER HAVE BEEN PLACED . AFTER SHAKING , MAKE THE DISTILLATE UP TO VOLUME AT A TEMPERATURE IDENTICAL WITH THE INITIAL TEMPERATURE TO WITHIN MORE OR LESS 2* C .

2 . MEASUREMENT OF THE DENSITY OF THE DISTILLATE : CAREFULLY FILL THE PYCNOMETER WITH THE DISTILLATE AT AMBIENT TEMPERATURE ; PLACE THE PYCNOMETER IN THE THERMALLY INSULATED ENCLOSURE ; SHAKE THE LIQUID TWO OR THREE TIMES BY INVERTING THE CONTAINER UNTIL THE TEMPERATURE READING ON THE THERMOMETER IS CONSTANT . ADJUST THE LEVEL ACCURATELY TO THE UPPER RIM OF THE SIDE TUBE . WIPE THE SIDE TUBE DRY , PUT ON THE RECEIVER PLUG AND MEASURE THE TEMPERATURE T* C WITH THE SAME CARE AS PREVIOUSLY ; CAREFULLY WIPE THE PYCNOMETER DRY AND WEIGH , P'' BEING THE WEIGHT .

THE DIFFERENCE P + M - P'' BETWEEN THE CORRECTED TARE AND THE WEIGHT P'' GIVES THE MASS OF THE LIQUID CONTAINED IN THE PYCNOMETER ( CORRECTED FOR BUOYANCY OF AIR ) .

THE APPARENT DENSITY * IS THEN CALCULATED BY DIVIDING THIS MASS BY THE VOLUME AT 20* C .

3 . USE OF THE TABLE OF APPARENT DENSITY * OF WATER-ALCOHOL MIXTURES TAKEN FROM THE INTERNATIONAL TABLE : FIND THE SMALLEST DENSITY GREATER THAN * IN THE TABLE ON THE HORIZONTAL LINE CORRESPONDING TO THE TEMPERATURE IN WHOLE UNITS IMMEDIATELY BELOW T* C . USE THE TABULAR DIFFERENCE READ BELOW THIS DENSITY * FOR CALCULATING THE DENSITY * AT THIS TEMPERATURE IN WHOLE UNITS .

SUBTRACT THE CALCULATED DENSITY FROM THE DENSITY * OF THE TABLE IMMEDIATELY ABOVE * . DIVIDE THIS DIFFERENCE BY THE TABULAR DIFFERENCE READ ON THE RIGHT OF THE DENSITY * . THE QUOTIENT GIVES THE DECIMAL PORTION OF THE ALCOHOLIC STRENGTH , WHEREAS THE WHOLE UNIT PORTION OF THIS STRENGTH IS SHOWN AT THE HEAD OF THE COLUMN WHERE THE DENSITY * IS LOCATED .

NOTE :

CALCULATIONS MAY BE AVOIDED DURING ROUTINE DETERMINATIONS BY CALCULATING ONE PARTICULAR TABLE FOR EACH PYCNOMETER GIVING THE WEIGHTS P'' WHICH RE-ESTABLISH EQUILIBRIUM ON THE BALANCE FOR EACH DEGREE OF ALCOHOLIC STRENGTH AND EACH TEMPERATURE ( IN 1* C UNITS ) .

CALCULATE THIS TABLE USING THE TABLE OF APPARENT DENSITIES BY WHOLE DEGREES OF ALCOHOLIC STRENGTH AND WHOLE DEGREES OF TEMPERATURE BY MULTIPLYING THE DENSITIES OF THE TABLE BY VOLUME OF THE PYCNOMETER AT 20* C AND BY SUBTRACTING THESE DENSITIES FROM THE WEIGHT P + M .

IF NECESSARY THE WEIGHTS P'' FOR 0.1 % VOL OF ALCOHOLIC STRENGTH AND 0.1* C UNITS OF TEMPERATURE MAY BE OBTAINED BY INTERPOLATION FROM THE TABLE WHICH HAS BEEN CALCULATED FOR WHOLE % VOL OF ALCOHOLIC STRENGTH AND FOR 1* C UNITS OF TEMPERATURE .

EXAMPLE OF THE CALCULATION OF THE ALCOHOLIC STRENGTH OF A WINE ( SEE OJ ) .

FOR MUSTS AND FERMENTING MUSTS , MEASUREMENT OF ALCOHOL BY CHEMICAL PROCEDURE

THIS METHOD SHOULD ONLY BE USED WITH LIQUIDS OF LOW ALCOHOLIC CONTENT ( LESS THAN 6 % VOL ) AND WITH SAMPLES WHERE ONLY A SMALL VOLUME IS AVAILABLE .

SOLUTIONS

( A ) POTASSIUM DICHROMATE SOLUTION : DISSOLVE 33.600 G OF POTASSIUM DICHROMATE ANALYTICAL REAGENT QUALITY IN DISTILLED WATER AND MAKE TO ONE LITRE AT 20* C . ( THIS SOLUTION CORRESPONDS TO AN INTERNATIONAL ALCOHOLIC STRENGTH ( 13 ) . )

1 ML OF THIS SOLUTION OXIDIZES 7.8924 MG OF ALCOHOL .

( B ) AMMONIUM FERROUS SULPHATE SOLUTION : DISSOLVE 135 G OF AMMONIUM FERROUS SULPHATE AND 20 ML OF CONCENTRATED SULPHURIC ACID IN WATER AND MAKE TO ONE LITRE . THIS SOLUTION IS EQUIVALENT TO APPROXIMATELY HALF ITS VOLUME OF DICHROMATE SOLUTION WHEN FRESHLY PREPARED ; IT WILL THEN SLOWLY OXIDIZE .

FREQUENTLY RESTANDARDIZE THIS SOLUTION AGAINST THE DICHROMATE SOLUTION WHICH KEEPS WELL IN A GROUND GLASS STOPPERED BOTTLE . CARRY OUT THE TITRATION IN THE SAME WAY AS FOR THE ALCOHOL DETERMINATION , EXCEPT THAT THE 10 ML OF ALCOHOL DILUTION IS REPLACED BY AN EQUAL VOLUME OF PURE WATER . THIS COMPENSATES FOR THE SMALL QUANTITIES OF ORGANIC SUBSTANCES WHICH SOMETIMES ARE PRESENT IN THE SULPHURIC ACID .

( C ) POTASSIUM PERMANGANATE SOLUTION : DISSOLVE 1.088 G OF POTASSIUM PERMANGANATE IN WATER AND MAKE UP TO ONE LITRE WITH WATER .

( D ) DILUTE SULPHURIC ACID : GRADUALLY ADD , WITH STIRRING , 500 ML OF CONCENTRATED SULPHURIC ACID TO 500 ML OF DISTILLED WATER . AFTER COOLING MAKE UP TO ONE LITRE WITH DISTILLED WATER .

( E ) FERROUS O-PHENANTHROLINE REAGENT : DISSOLVE 0.695 G OF FESO4 , 7H2O IN 100 ML OF WATER AND ADD 1.485 G OF O-PHENANTHROLINE MONOHYDRATE . HEAT TO ASSIST DISSOLUTION . THIS SOLUTION , BRIGHT RED IN COLOUR , KEEPS VERY WELL .

PROCEDURE

1 . DISTILLATION : SEPARATE THE ALCOHOL BY DISTILLATION , USING THE EQUIPMENT DESCRIBED ABOVE ( PAGE 16 ) . CALCULATE THE VOLUMES OF THE SAMPLE AND OF THE DISTILLATE SO THAT THE ALCOHOLIC STRENGTH OF THE LATTER SHOULD BE OF THE ORDER OF 1 % VOL ; IT MUST BE LESS THAN 1.8 % VOL .

TRANSFER A VOLUME V OF LIQUID , WINE OR MUST TO BE ANALYZED , WHICH HAS BEEN MADE ALKALINE BY THE ADDITION OF A SLIGHT EXCESS OF 4N MILK OF LIME AND THE PRESCRIBED VOLUME OF WATER , TO THE STEAM-DISTILLATION FLASK DESCRIBED PREVIOUSLY . COLLECT THE DISTILLATE IN A MEASURING FLASK OF EQUAL VOLUME TO THAT SPECIFIED IN THE LAST COLUMN OF THE ABOVE TABLE . FILL UP TO THE MARK WITH DISTILLED WATER .

THE DISTILLATE MAY ALSO BE DILUTED BY ONE OF THE ROUTINE METHODS ( PAGE 27 ) , BY TAKING 50 , 25 OR 20 ML OF THE DISTILLATE AND MAKING UP TO 100 , 200 OR 250 ML .

2 . OXIDATION : PIPETTE 20 ML OF STANDARD POTASSIUM DICHROMATE SOLUTION AND 20 ML OF 50 % V/V SULPHURIC ACID INTO A 250 ML WIDE-NECKED FLASK FITTED WITH A GROUND-GLASS STOPPER . MIX WITH STIRRING . PIPETTE 10 ML OF DISTILLATE INTO THE FLASK . STOPPER THE FLASK , THE STOPPER HAVING BEEN PREVIOUSLY MOISTENED WITH A DROP OF CONCENTRATED SULPHURIC ACID , SHAKE AND ALLOW TO STAND FOR AT LEAST 30 MINUTES WITH OCCASIONAL SHAKING .

3 . TITRATION : RINSE THE STOPPER AND NECK OF THE FLASK AND TITRATE THE EXCESS DICHROMATE WITH STANDARD FERROUS AMMONIUM SULPHATE FROM A CALIBRATED BURETTE USING FOUR DROPS OF O-PHENANTHROLINE AS INDICATOR ADDED WHEN THE GREEN COLORATION OF THE SOLUTION TURNS TO GREEN-BLUE . THE END-POINT IS INDICATED WHEN THE COLOUR CHANGES FROM BLUE-GREEN TO A RED-BROWN .

AS THE COLOUR CHANGE IS FREQUENTLY SLIGHTLY OVERSHOT , RETURN TO THE EXACT END-POINT USING A POTASSIUM PERMANGANATE SOLUTION OF 1.088 G/L . SUBTRACT ONE TENTH OF THE VOLUME OF THIS SOLUTION USED FROM THE VOLUME OF THE FERROUS SULPHATE SOLUTION , THE DIFFERENCE BEING N .

CARRY OUT A BLANK EXPERIMENT USING A SIMILAR BOTTLE CONTAINING THE SAME QUANTITIES OF THE SAME REAGENTS EXCEPT THAT THE 10 ML OF DISTILLATE HAVE BEEN REPLACED BY 10 ML OF PURE WATER . THE VOLUME OF THE FERROUS SOLUTION USED IS N' .

CALCULATIONS ( SEE OJ )

NOTES :

1 . THE 10 AND 20 ML PIPETTES AND THE BURETTE USED FOR THE MEASUREMENT MUST BE CERTIFIED AND ACCURATE TO AT LEAST ONE PART PER 1 000 .

2 . THE MEASUREMENT OF THE ACETIC ACID FORMED PERMITS CHECKING THE CHROMOMETRIC DETERMINATION OF THE ALCOHOL AND THE IDENTIFICATION OF THAT ALCOHOL . METHANOL , IN PARTICULAR , IS OXIDIZED TO CARBON DIOXIDE AND WATER UNDER THE CONDITIONS WHERE ETHANOL IS CONVERTED TO ACETIC ACID ; THE OXIDATION OF THE FORMER REQUIRES AT LEAST 12 HOURS HOWEVER , WHEREAS THAT OF ETHANOL IS COMPLETE IN 30 MINUTES ( 14 ) .

IT SHOULD BE NOTED THAT 1 ML OF K2CR2O7 OF CONCENTRATION 33.600 G/L OXIDIZES 3.6596 MG OF METHANOL .

TO MEASURE THE ACETIC ACID , STEAM-DISTILL AND COLLECT THE DISTILLATE . THE VOLUME OF THE DISTILLATE SHOULD BE EQUAL TO AT LEAST 10 TIMES THE VOLUME OF THE SOLUTION IN WHICH THE CHROMOMETRIC TITRATION HAS BEEN CARRIED OUT AND TO WHICH A SMALL EXCESS OF FERROUS SULPHATE HAS BEEN ADDED .

1 ML 0.1N NAOH CORRESPONDS TO 4.607 MG OF ETHANOL .

1 ML K2CR2O7 OF CONCENTRATION 33.600 G/L THUS CORRESPONDS TO 1.71 ML OF 0.1N NAOH .

2.4 . USUAL METHODS FOR WINES AND MUSTS

FIRST USUAL METHOD

MEASUREMENT BY A HYDROMETER

APPARATUS

1 . DISTILLATION APPARATUS : THE APPARATUS IS SIMILAR TO THAT DESCRIBED FOR THE REFERENCE METHOD .

AS FOR THE REFERENCE METHOD OTHER TYPES OF EQUIPMENT MAY BE USED . IN PARTICULAR STEAM DISTILLATION MAY BE EMPLOYED FOR THE DISTILLATION OF THE ALCOHOL , WHICH SPEEDS THE OPERATION , PROVIDED THAT THE APPARATUS AND PROCEDURE FULFIL THE CONDITIONS SPECIFIED FOR THE REFERENCE METHOD ( SEE PAGE 16 ) .

2 . HYDROMETER : THE HYDROMETER USED SHALL COMPLY WITH THE SPECIFICATIONS OF COUNCIL DIRECTIVE 76/765/EEC OF 27 JULY 1976 RELATING TO ALCOHOLOMETERS AND ALCOHOL HYDROMETERS .

THIS APPARATUS SHALL BE CALIBRATED BY A GOVERNMENT DEPARTMENT .

A CALIBRATED THERMOMETER GRADUATED IN 1/10TH DEGREES FROM 0 TO 30* C CERTIFIED TO 1/20TH DEGREE SHALL BE USED .

CARRY OUT THE TEST IN AN UPRIGHT CYLINDRICAL TUBE OF 36 MM DIAMETER AND 320 MM HEIGHT . THE INSIDE DIAMETER OF THE CYLINDER MUST EXCEED THE BULB DIAMETER OF THE HYDROMETER BY AT LEAST 6 MM .

PROCEDURE

1 . DISTILLATION : FREE YOUNG OR SPARKLING WINES OF THE BULK OF THEIR CARBON DIOXIDE BY SHAKING 260 ML OF THE WINE IN A 500 ML BOTTLE WHICH HAS PREVIOUSLY BEEN SILICONIZED INTERNALLY .

MEASURE OUT THE WINE TO BE ANALYZED INTO A 250 ML FLASK WITH AN INTERNAL NECK DIAMETER NOT GREATER THAN 12 MM . TRANSFER TO A DISTILLATION FLASK . RINSE THE FLASK FOUR TIMES WITH 5 ML OF WATER . ADD 10 ML OF MILK OF LIME ( 120 G OF CAO PER LITRE ) . THE COLOURING MATTER OF THE WINE SHOULD BE CHANGED BY THE ALKALINITY . IN THE CASE OF VERY ACID , SOUR WINES , ETC . , ADD THE LIME UNTIL THE WINE IS CLEARLY ALKALINE TO PHENOLPHTHALEIN USED EXTERNALLY . ADD , IF NECESSARY , SOME INERT POROUS MATTER AND A DROP OF AN AQUEOUS SOLUTION , OF ONE PART PER 100 OF WATER-SOLUBLE SILICONE TO PREVENT FROTHING . COLLECT THE DISTILLATE IN THE SAME FLASK AS THAT USED FOR MEASURING OUT THE WINE , AND INTO WHICH HAS BEEN PLACED 10 ML OF DISTILLED WATER .

DISTILL AT LEAST 200 ML . SHAKE THE FLASK AND MAKE UP TO THE MARK AT THE SAME INITIAL TEMPERATURE ( TO WITHIN 5* C ) .

2 . MEASUREMENT OF SPECIFIC GRAVITY OF THE DISTILLATE : POUR THE DISTILLATE INTO A GLASS CYLINDER STANDING ON A LEVEL SURFACE OF 36 MM INTERNAL DIAMETER AND 320 MM HEIGHT . KEEP THE CYLINDER VERTICAL . PLACE A THERMOMETER AND HYDROMETER IN THE CYLINDER . READ THE THERMOMETER ONE MINUTE AFTER SHAKING , SO ENSURING EQUILIBRIUM TEMPERATURE OF CYLINDER , THERMOMETER , HYDROMETER AND DISTILLATE HAS BEEN REACHED . WITHDRAW THE THERMOMETER AND READ THE APPARENT DEGREE OF ALCOHOLIC STRENGTH AFTER ONE MINUTE . TAKE AT LEAST THREE READINGS USING A MAGNIFYING GLASS . CORRECT THE APPARENT DEGREE MEASURED AT T* C FOR THE EFFECT OF THE TEMPERATURE WITH THE FOLLOWING TABLE .

WHEN MAKING THE OBSERVATIONS THE TEMPERATURE SHOULD NOT DIFFER FROM AMBIENT BY MORE THAN 5* C . IF A GREATER DIFFERENCE IS OBSERVED IT MAY BE AVOIDED BY USING A VACUUM FLASK .

SECOND USUAL METHOD

DENSIMETRY BY THE HYDROSTATIC BALANCE

THE DISTILLATION APPARATUS AND ITS METHOD OF OPERATION ARE THE SAME AS FOR THE FIRST USUAL METHOD .

THE APPARATUS , THE CALIBRATION OF THE FLOAT AND THE PROCEDURE HAVE BEEN DESCRIBED IN CONNECTION WITH THE MEASUREMENT OF THE DENSITY , BUT THE MEASUREMENT OF DENSITY OF THE DISTILLATE MUST BE MORE ACCURATE THAN THAT OF WINE . IT SHOULD BE OBTAINED AT LEAST TO THE NEAREST 5 BY 10-5 G/ML . IN ORDER TO ACHIEVE THIS RESULT ONE SHOULD USE A FLOAT OF AT LEAST 50 ML AND TAKE ALL NECESSARY PRECAUTIONS TO AVOID ANY EVAPORATION OF THE ALCOHOL AND TO KNOW THE TEMPERATURE , T* C , OF THE DISTILLATE TO AT LEAST THE NEAREST 0.1* C .

THE FORMULAE FOR THE CALCULATION HAVE ALSO BEEN MENTIONED ALREADY .

KNOWING PT AT T* C , ONE DETERMINES THE ALCOHOLOMETRIC TITRE BY MEANS OF ONE OF THE DENSITY TABLES , WHICH ONE SELECTS ACCORDING TO THE NATURE OF THE GLASS USED IN THE FLOAT , FOLLOWING THE SAME INTERPOLATION PROCEDURE AS FOR THE REFERENCE METHOD .

2.5 . SUPPLEMENT TO THE REFERENCE METHOD FOR THE MEASUREMENT OF THE DENSITY OF THE DISTILLATE

MEASUREMENT BY PYCNOMETER ON SINGLE-PAN BALANCE

PRINCIPLE

THE PRINCIPLE OF THE METHOD OF DETERMINING THE DENSITY OF THE DISTILLATE IS GIVEN IN " DENSITY AND SPECIFIC GRAVITY " .

NUMERICAL EXAMPLE

DETERMINE THE CONSTANTS OF THE PYCNOMETER AND CALCULATE AS DESCRIBED IN " DENSITY AND SPECIFIC GRAVITY " .

CALCULATIONS ( SEE OJ )

3 . TOTAL DRY EXTRACT

TOTAL DRY MATTER

3.1 . DEFINITION

THE TOTAL DRY EXTRACT OR THE TOTAL DRY MATTER INCLUDES ALL MATTER WHICH IS NON-VOLATILE UNDER SPECIFIED PHYSICAL CONDITIONS . THESE PHYSICAL CONDITIONS MUST BE SUCH THAT THE MATTER FORMING THE EXTRACT IS NOT MATERIALLY ALTERED WHILE THE TEST IS BEING CARRIED OUT .

THE SUGAR-FREE EXTRACT IS THE DIFFERENCE BETWEEN TOTAL DRY EXTRACT AND THE TOTAL SUGARS .

THE REDUCED EXTRACT IS THE DIFFERENCE BETWEEN THE TOTAL DRY EXTRACT AND THE TOTAL SUGARS IN EXCESS OF 1 G/L , POTASSIUM SULPHATE IN EXCESS OF 1 G/L , ANY MANNITOL PRESENT AND ANY OTHER CHEMICAL SUBSTANCES WHICH MAY HAVE BEEN ADDED TO THE WINE .

THE RESIDUAL EXTRACT IS THE SUGAR-FREE EXTRACT LESS THE FIXED ACIDITY EXPRESSED AS TARTARIC ACID .

THE EXTRACT IS EXPRESSED IN GRAMS PER LITRE AND IT SHOULD BE DETERMINED TO WITHIN THE NEAREST 0.5 G .

3.2 . PRINCIPLE OF THE METHOD

SINGLE METHOD : MEASUREMENT BY A DENSIMETER

THE TOTAL DRY EXTRACT IS CALCULATED INDIRECTLY FROM THE SPECIFIC GRAVITY OF THE ALCOHOL-FREE RESIDUE .

THIS DRY EXTRACT IS EXPRESSED IN TERMS OF THE QUANTITY OF SUCROSE WHICH , WHEN DISSOLVED IN WATER AND MADE UP TO A VOLUME OF ONE LITRE , GIVES A SOLUTION OF THE SAME SPECIFIC GRAVITY AS THE ALCOHOL-FREE RESIDUE . THIS QUANTITY IS SHOWN IN TABLE I .

METHOD OF CALCULATION ( SEE OJ )

4 . REDUCING SUGARS

4.1 . DEFINITION

THE REDUCING SUGARS , COMPRISING ALL OF THE SUGARS EXHIBITING KETONIC AND ALDEHYDIC FUNCTIONS , ARE DETERMINED USING THEIR REDUCING ACTION ON AN ALKALINE COPPER SALT SOLUTION .

THEIR DETERMINATION REQUIRES TWO SUCCESSIVE OPERATIONS : CLARIFICATION AND TITRATION .

4.2 . PRINCIPLE OF THE METHODS

1 . CLARIFICATION

REFERENCE METHOD : AFTER NEUTRALIZATION AND ALCOHOL REMOVAL THE WINE IS PASSED THROUGH AN EXCHANGE COLUMN IN WHICH ITS ANIONS ARE EXCHANGED BY ACETATE IONS , FOLLOWED BY CLARIFICATION WITH NEUTRAL LEAD ACETATE .

USUAL METHOD : THE WINE IS TREATED WITH ONE OF THE FOLLOWING REAGENTS :

( A ) NEUTRAL LEAD ACETATE ;

( B ) WINE FERROCYANIDE .

2 . DETERMINATION

SINGLE METHOD : AFTER THE CLARIFIED MUST OR WINE HAS REACTED WITH A SPECIFIC QUANTITY OF ALKALINE COPPER SALT SOLUTION , THE EXCESS COPPER IONS ARE DETERMINED IODOMETRICALLY .

3 . EXPRESSION OF RESULTS

THE QUANTITY OF REDUCING SUGARS IS EXPRESSED AS GRAMS OF INVERT SUGAR PER LITRE , TO WITHIN 0.1 G .

4.3 . CLARIFICATION

THE SUGAR CONTENT OF THE LIQUID IN WHICH SUGAR IS TO BE DETERMINED MUST LIE BETWEEN 0.5 AND 5 G/L .

DRY WINES SHOULD NOT BE DILUTED DURING CLARIFICATION ; SWEET WINES SHOULD BE DILUTED DURING CLARIFICATION IN ORDER TO BRING THE SUGAR LEVEL WITHIN THE PRESCRIBED LIMITS , ACCORDING TO THE FOLLOWING TABLE ( 15 ) ( SEE OJ ) .

IF THE QUANTITY OF SUGARS AND THE ROTATING POWER ARE TO BE DETERMINED , IT IS FIRST NECESSARY TO OBTAIN A COLOURLESS LIQUID AS CONCENTRATED AS POSSIBLE TO ENSURE THE BEST POLARIMETRIC MEASUREMENT , AND THEN TO DILUTE IT FOR THE CHEMICAL ASSAY OF THE SUGARS IN THE SAME LIQUID .

1 . REFERENCE METHOD

PREPARATION OF THE ANION EXCHANGE RESIN COLUMN

THE BASIC APPARATUS CONSISTS OF A 50 ML BURETTE WITH STOPCOCK , SURMOUNTED BY A FUSED-ON GLASS CYLINDER WHICH SERVES AS A RESERVOIR . A SMALL GLASS WOOL PAD AND 15 ML OF DOWEX 3 RESIN ( 20 - 50 MESH ) IS PLACED IN THE BOTTOM OF THE BURETTE .

BEGIN BY SUBJECTING THE RESIN TO TWO COMPLETE CYCLES OF REGENERATION , WITH ALTERNATE PASSAGES OF N SOLUTIONS OF HYDROCHOLORIC ACID AND SODIUM HYDROXIDE . AFTER RINSING WITH 50 ML DISTILLED WATER , TRANSFER THE RESIN TO A BEAKER , ADD 50 ML OF A 4N SOLUTION OF ACETIC ACID AND STIR FOR FIVE MINUTES . THEN FILL THE BURETTE AGAIN AND POUR 100 ML OF THE 4N SOLUTION OF ACETIC ACID THROUGH THE COLUMN . ( IT IS PREFERABLE TO HAVE A STOCK OF RESIN STORED IN A BOTTLE FILLED WITH THIS 4N ACETIC ACID SOLUTION : THIS AVOIDS HAVING TO WASH THE COLUMN WITH THE ACETIC ACID SOLUTION . ) WASH THE COLUMN WITH DISTILLED WATER UNTIL THE EFFLUENT IS NEUTRAL .

REGENERATION OF THE RESIN . - POUR 150 ML OF A 2N SODIUM HYDROXIDE SOLUTION OVER THE RESIN TO REMOVE ACID AND MOST OF THE PIGMENTS FIXED TO THE RESIN . RINSE WITH 100 ML WATER , THEN POUR OVER 100 ML OF 4N ACETIC SOLUTION . WASH THE COLUMN UNTIL THE EFFLUENT IS NEUTRAL .

SOLUTION

NEUTRAL LEAD ACETATE SOLUTION ( SATURATED ) :

NEUTRAL LEAD ACETATE 250 G ,

VERY HOT WATER TO 500 ML .

STIR UNTIL DISSOLVED .

PROCEDURE

( A ) DRY WINE : PLACE 50 ML OF WINE IN A BEAKER HAVING A DIAMETER OF APPROXIMATELY 10 TO 12 CM , WITH ( N - 0.5 ) ML OF A N SOLUTION OF SODIUM HYDROXIDE ( N BEING THE VOLUME OF A N/10 SOLUTION OF SODIUM HYDROXIDE USED FOR DETERMINING ACIDITY IN 5 ML WINE ) AND EVAPORATE OVER A BOILING WATER BATH IN A STREAM OF HOT AIR UNTIL THE LIQUID IS REDUCED TO ABOUT 20 ML .

ALLOW THIS LIQUID TO FLOW THROUGH AN ANION EXCHANGE COLUMN , DOWEX 3 ( 20 - 50 MESH ) , IN ACETATE FORM , AT A RATE OF 3 ML EVERY TWO MINUTES . THE ACIDS ARE BOUND BY THE RESIN AND EXCHANGED BY THE ACETATE ANIONS . COLLECT THE EFFLUENT IN A CALIBRATED 100 ML FLASK . WASH THE VESSEL AND COLUMN SIX TIMES WITH 10 ML DISTILLED WATER EACH TIME . WHILE STIRRING , ADD 1 ML OF SATURATED LEAD ACETATE SOLUTION AND 0.5 G OF CALCIUM CARBONATE ; SHAKE SEVERAL TIMES AND ALLOW TO STAND FOR 15 MINUTES ; MAKE UP TO THE MARK WITH WATER . FILTER .

1 ML OF THIS FILTRATE CORRESPONDS TO 0.5 ML OF WINE .

( B ) MUSTS , MISTELLES AND SWEET WINES ( DETERMINATION OF SUGAR WITHOUT DETERMINATION OF POLARIMETRIC ROTATION ) :

1 . MUSTS AND MISTELLES : PREPARE A 10 % DILUTION OF THE LIQUID TO BE ANALYZED AND TAKE 10 ML OF THE DILUTED SAMPLE .

2 . SWEET WINES , WHETHER OR NOT FORTIFIED , HAVING A DENSITY BETWEEN 1.005 AND 1.038 : PREPARE A 20 % DILUTION OF THE LIQUID TO BE ANALYZED AND TAKE 20 ML OF THE DILUTED SAMPLE .

3 . SEMI-SWEET WINES HAVING A DENSITY BETWEEN 0.997 AND 1.006 : TAKE 20 ML OF THE NON-DILUTED WINE .

ALLOW THE ABOVEMENTIONED VOLUME OF WINE OR MUST TO FLOW THROUGH AN ANION EXCHANGE COLUMN , DOWEX 3 ( 20 - 50 MESH ) IN ACETATE FORM , AT A RATE OF 3 ML EVERY TWO MINUTES . COLLECT THE EFFLUENT IN A 100 ML FLASK , RINSE THE COLUMN WITH WATER UNTIL 90 ML OF EFFLUENT IS OBTAINED . ADD 0.5 G CALCIUM CARBONATE AND 1 ML SATURATED LEAD ACETATE SOLUTION ; STIR AND ALLOW TO STAND FOR 15 MINUTES , STIRRING FROM TIME TO TIME ; MAKE UP TO THE MARK WITH WATER . FILTER .

1 . 1 ML OF FILTRATE CORRESPONDS TO 0.01 ML MUST OR MISTELLE .

2 . 1 ML OF FILTRATE CORRESPONDS TO 0.04 ML OF SWEET WINE .

3 . 1 ML OF FILTRATE CORRESPONDS TO 0.20 ML SEMI-SWEET WINE .

( C ) DETERMINATION OF OPTICAL ROTATION OF SWEET WINES ( WINES FROM BOTRYTIZED , EUDEMIZED GRAPES , ETC . , NATURALLY SWEET WINES AND LIQUEUR WINES ) : PROCEED AS IN THE FIRST PARAGRAPH OF ( A ) , BUT DILUTE THE ALCOHOL-FREE WINE TO 100 ML BEFORE ALLOWING IT TO PASS THROUGH THE ANIONIC EXCHANGE COLUMN . COLLECT THE EFFLUENT IN A 200 ML FLASK ; RINSE THE COLUMN WITH DISTILLED WATER TO OBTAIN ABOUT 180 ML OF EFFLUENT . ADD 1 ML OF SATURATED LEAD ACETATE SOLUTION AND 0.5 G OF CALCIUM CARBONATE WHILE STIRRING ; SHAKE SEVERAL TIMES AND LEAVE TO STAND AT LEAST 15 MINUTES ; MAKE UP TO THE MARK WITH WATER . FILTER .

1 ML OF THIS LIQUID CORRESPONDS TO 0.25 ML WINE .

2 . USUAL METHODS

A . CLARIFICATION BY NEUTRAL LEAD ACETATE

SOLUTION

SOLUTION OF NEUTRAL LEAD ACETATE ( SATURATED ) :

NEUTRAL LEAD ACETATE 250 G ,

VERY HOT WATER TO 500 ML .

STIR UNTIL DISSOLVED .

PROCEDURE

( A ) DRY WINE : PLACE 50 ML OF WINE IN A CALIBRATED 100 ML FLASK ; ADD ( N - 0.5 ) ML OF A N SOLUTION OF SODIUM HYDROXIDE ( N BEING THE VOLUME OF A N/10 SOLUTION OF SODIUM HYDROXIDE USED FOR DETERMINING ACIDITY IN 5 ML WINE ) . WHILE STIRRING ADD 2.5 ML OF SATURATED LEAD ACETATE SOLUTION AND 0.5 G CALCIUM CARBONATE ; SHAKE SEVERAL TIMES AND ALLOW TO STAND AT LEAST 15 MINUTES ; MAKE UP TO THE MARK WITH WATER . FILTER .

1 ML OF FILTRATE CORRESPONDS TO 0.5 ML WINE .

( B ) MUSTS , MISTELLES AND SWEET WINES ( DETERMINATION OF SUGAR WITHOUT DETERMINATION OF POLARIMETRIC ROTATION ) : PLACE A VOLUME OF WINE ( OR MUST OR MISTELLE ) DEFINED AS FOLLOWS IN A GRADUATED 100 ML FLASK :

1 . MUSTS AND MISTELLES : PREPARE A 10 % DILUTION OF THE LIQUID TO BE ANALYZED AND TAKE 10 ML OF THIS SOLUTION .

2 . SWEET WINES , WHETHER OR NOT FORTIFIED , HAVING A DENSITY BETWEEN 1.005 AND 1.038 : PREPARE A 20 % DILUTION OF THE LIQUID TO BE ANALYZED AND TAKE 20 ML OF THE DILUTED SAMPLE .

3 . SEMI-SWEET WINES HAVING A DENSITY BETWEEN 0.997 AND 1.006 : TAKE 20 ML OF THE NON-DILUTED WINE .

ADD 0.5 G CALCIUM CARBONATE , ABOUT 60 ML WATER AND 0.5 , 1 OR 2 ML SATURATED LEAD ACETATE SOLUTION ; STIR AND LEAVE TO STAND FOR 15 MINUTES , STIRRING FROM TIME TO TIME . MAKE UP TO THE MARK . FILTER .

1 . 1 ML OF FILTRATE CORRESPONDS TO 0.01 ML OF MUST OR MISTELLE .

2 . 1 ML OF FILTRATE CORRESPONDS TO 0.04 ML OF SWEET WINE .

3 . 1 ML OF FILTRATE CORRESPONDS TO 0.20 ML SEMI-SWEET WINE .

B . CLARIFICATION BY ZINC FERROCYANIDE

THIS CLARIFICATION PROCESS SHOULD BE USED ONLY FOR WHITE WINES , LIGHTLY COLOURED SWEET WINES AND MUSTS .

SOLUTIONS

SOLUTION I , POTASSIUM FERROCYANIDE :

POTASSIUM FERROCYANIDE 150 G ,

WATER TO 1 000 ML .

SOLUTION II , ZINC SULPHATE :

ZINC SULPHATE 300 G ,

WATER TO 1 000 ML .

PROCEDURE

PLACE A VOLUME OF WINE ( OR MUST OR MISTELLE ) DEFINED AS FOLLOWS IN A CALIBRATED 100 ML FLASK :

1 . MUSTS AND MISTELLES : PREPARE A 10 % DILUTION OF THE LIQUID TO BE ANALYZED AND TAKE 10 ML OF THE DILUTED SAMPLE .

2 . SWEET WINES , WHETHER OR NOT FORTIFIED , HAVING A DENSITY BETWEEN 1.005 AND 1.038 : PREPARE A 20 % DILUTION OF THE LIQUID TO BE ANALYZED AND TAKE 20 ML OF THE DILUTED SAMPLE .

3 . SEMI-SWEET WINES HAVING A DENSITY BETWEEN 0.997 AND 1.006 : TAKE 20 ML OF UNDILUTED WINE .

4 . DRY WINES : TAKE 50 ML OF UNDILUTED WINE .

ADD 5 ML OF SOLUTION I OF POTASSIUM FERROCYANIDE AND 5 ML OF SOLUTION II OF ZINC SULPHATE . MIX . MAKE UP TO THE MARK . WAIT 10 MINUTES , FILTER .

1 . 1 ML OF FILTRATE CORRESPONDS TO 0.01 ML OF MUST OR MISTELLE .

2 . 1 ML OF FILTRATE CORRESPONDS TO 0.04 ML OF SWEET WINE .

3 . 1 ML OF FILTRATE CORRESPONDS TO 0.20 ML OF SEMI-SWEET WINE .

4 . 1 ML OF FILTRATE CORRESPONDS TO 0.50 ML OF DRY WINE .

4.4 . DETERMINATION OF SUGARS

THE STANDARD SOLUTION OF INVERT SUGAR

ALL ANALYSTS MUST CHECK THEIR PROCEDURE USING A STANDARD SOLUTION OF INVERT SUGAR . PREPARE THIS SOLUTION ACCORDING TO THE FOLLOWING METHOD :

INVERT SUGAR SOLUTION CONTAINING 10 G/L .

PLACE THE FOLLOWING IN A ONE LITRE FLASK :

SUCROSE PURE AND DRY 9.50 G ,

WATER , APPROXIMATELY 100 ML ,

HYDROCHLORIC ACID , CONCENTRATED D = 1.16 TO 1.19 ) 5 ML .

LEAVE FOR SEVEN DAYS AT 10 TO 15* C , OR THREE DAYS AT 20 TO 25* C , AFTER WHICH THE SUGAR HAS INVERTED ; THEN MAKE UP THE SOLUTION TO 1 000 ML AT 20* C . THIS ACID SOLUTION KEEPS WELL FOR ONE MONTH . SHORTLY BEFORE USE , NEUTRALIZE THE BULK OF THE FREE ACID WITH SODIUM HYDROXIDE SOLUTION ( THE PREPARED SOLUTION IS APPROXIMATELY 0.06N ACID ) .

NOTE :

THE STANDARD INVERT SUGAR SOLUTION MAY BE PREPARED MORE QUICKLY AS FOLLOWS : HEAT THE SUCROSE SOLUTION , ACIDIFIED AS ABOVE , IN A 200 ML FLASK ON A WATERBATH MAINTAINED AT 60* C TO A TEMPERATURE OF 50* C ; MAINTAIN THE FLASK AND CONTENTS AT 50* C FOR 15 MINUTES ; ALLOW TO COOL FOR 30 MINUTES THEN IMMERSE IN COLD WATER ; TRANSFER TO A ONE LITRE VOLUMETRIC FLASK AND MAKE UP TO VOLUME AT 20* C .

TO TEST THE CHOSEN PROCEDURE , DETERMINE THE INVERT SUGAR AT SEVERAL DILUTIONS COVERING ALL THE CONCENTRATIONS OVER WHICH IT IS TO BE APPLIED . FOR INSTANCE , FOR THE STANDARD VOLUMETRIC METHOD , 20 ML SAMPLES CONTAINING 10 , 20 , 50 AND 100 MG OF INVERT SUGAR ARE OBTAINED BY DILUTING THE 10 G/L SOLUTION TO 1/20 , 1/10 , 1/4 AND 1/2 .

4.5 . DETERMINATION OF EXCESS OF COPPER IONS BY IODOMETRY

THE QUANTITY OF COPPER PRECIPITATED BY THE ACTION OF THE CLARIFIED SOLUTION ON THE ALKALINE COPPER SALT SOLUTION IS DETERMINED BY IODOMETRY .

SOLUTIONS

ALKALINE COPPER SALT SOLUTION :

COPPER SULPHATE , PURE , CUSO4 , 5H2O 25 G ,

CITRIC ACID 50 G ,

SODIUM CARBONATE CRYSTALLINE 388 G ,

WATER TO 1 000 ML .

DISSOLVE THE COPPER SULPHATE IN 100 ML OF WATER , THE CITRIC ACID IN 300 ML OF WATER AND THE SODIUM CARBONATE IN 300 TO 400 ML OF HOT WATER . MIX THE CITRIC ACID AND THE SODIUM CARBONATE SOLUTIONS . ADD THE COPPER SULPHATE SOLUTION AND MAKE UP TO ONE LITRE .

POTASSIUM IODIDE SOLUTION , 30 % :

POTASSIUM IODIDE 30 G ,

WATER TO 100 ML

STORE IN A BROWN BOTTLE .

SULPHURIC ACID , 25 :

SULPHURIC ACID , CONCENTRATED 25 G ,

DISTILLED WATER TO 100 ML .

ADD THE ACID SLOWLY TO THE WATER , ALLOW TO COOL AND MAKE UP TO 100 ML WITH DISTILLED WATER .

STARCH SOLUTION , 5 G/L , CONTAINING 200 G OF SODIUM CHLORIDE PER LITRE AS PRESERVATIVE . BOIL THIS SOLUTION FOR 10 MINUTES DURING PREPARATION .

SODIUM THIOSULPHATE 0.1N .

PROCEDURE

MIX 15 ML WATER , 10 ML OF CLARIFIED SOLUTION AND 25 ML OF ALKALINE COPPER SALT SOLUTION IN A 300 ML CONICAL FLASK . THIS VOLUME OF SUGAR SOLUTION MUST NOT CONTAIN MORE THAN 60 MG OF INVERT SUGAR .

ADD A FEW GRAINS OF PUMICE STONE AND BRING THE MIXTURE TO BOILING WITHIN TWO MINUTES . FIT A REFLUX CONDENSER TO THE FLASK AND BOIL FOR EXACTLY 10 MINUTES .

COOL THE FLASK IMMEDIATELY IN COLD RUNNING WATER . ADD , WHEN COOL , 10 ML OF 30 % POTASSIUM IODIDE SOLUTION , 25 ML OF 25 % SULPHURIC ACID AND 2 ML OF STARCH SOLUTION .

TITRATE WITH THE 0.1N SODIUM THIOSULPHATE SOLUTION , LET N' BE THE NUMBER OF ML USED .

CARRY OUT A BLANK TITRATION IN WHICH THE 25 ML OF SUGAR SOLUTION IS SUBSTITUTED BY 25 ML OF DISTILLED WATER . THE VOLUME OF THIOSULPHATE USED IN N' .

CALCULATIONS

THE QUANTITY OF SUGAR EXPRESSED AS INVERT SUGAR CONTAINED IN THE TEST SAMPLE IS GIVEN IN THE FOLLOWING TABLE AS A FUNCTION OF THE NUMBER ( N' - N ) ML OF THIOSULPHATE USED . ( SEE OJ )

THE SUGAR CONTENT OF THE WINE IS EXPRESSED IN GRAMS OF INVERT SUGAR PER LITRE , ACCOUNT BEING TAKEN OF THE DILUTIONS MADE IN THE COURSE OF CLARIFICATION AND OF THE VOLUME OF THE TEST SAMPLE .

5 . SUCROSE

5.1 . PRINCIPLE OF METHODS

1 . REFERENCE METHOD

( A ) FOR QUALITATIVE TESTING :

THIN LAYER CHROMATOGRAPHY : THE SUCROSE IS IDENTIFIED IN THE MUSTS AND IN THE WINES AFTER SEPARATION FROM GLUCOSE AND FRUCTOSE BY THIN LAYER CHROMATOGRAPHY ON A PLATE COATED WITH SILICA G POWDER BUFFERED WITH SODIUM ACETATE . THE DEVELOPING AGENT ( THIOBARBITURIC ACID AND TRICHLOROACETIC ACID ) IS INCORPORATED IN THE SOLVENT ( ETHYL ACETATE , ISO-PROPANOL AND WATER ) . WHEN SUCROSE IS HEATED IN ACID MEDIUM , HYDROXYMETHYLFURFURAL IS PRODUCED WHICH ON REACTING WITH THIOBARBITURIC ACID GIVES A YELLOW-ORANGE COLORATION .

( B ) FOR MEASUREMENT AFTER INVERSION :

THE SUCROSE IS DETERMINED BY COMPARISON OF THE REDUCING POWER BEFORE AND AFTER INVERSION WITH HYDROCHLORIC ACID OF THE SOLUTION , OBTAINED AFTER CLARIFICATION OF THE WINE . THE CLARIFIED SOLUTION IS PREPARED ACCORDING TO THE PROCEDURES DESCRIBED UNDER " REDUCING SUGARS " .

THE MEASUREMENT OF THE REDUCING SUGARS BEFORE AND AFTER INVERSION IS CARRIED OUT BY THE METHOD DESCRIBED UNDER " REDUCING SUGARS " .

2 . USUAL METHOD

( A ) FOR QUALITATIVE TESTING :

COLORIMETRIC METHOD : THE WINE IS CLARIFIED USING LEAD ACETATE , MAGNESIA AND POTASSIUM PERMANGANATE AT PH 8 TO 9 . THE SOLUTION OBTAINED IS REACTED AT 100* C WITH DIPHENYLAMINE IN A HYDROCHLORIC AND ACETIC ACID MEDIUM . THE CONDENSATION PRODUCT OBTAINED IN THE PRESENCE OF SUCROSE IS EXTRACTED INTO CHLOROFORM WHICH FORMS A BLUE COLOUR .

( B ) FOR MEASUREMENT AFTER INVERSION SEE UNDER ( 1 ) ( B ) ABOVE .

5.2 . QUALITATIVE TESTING

1 . REFERENCE METHOD

THIN LAYER CHROMATOGRAPHY METHOD

APPARATUS :

- APPARATUS REQUIRED FOR THIN LAYER CHROMATOGRAPHY :

- LAYER SPREADER ,

- PLATE HOLDER ,

- CHROMATOGRAPHY TANK ,

- MICROMETRIC SYRINGE OR MICRO-PIPETTE ,

- OVEN WITH AIR CIRCULATION .

SOLUTION AND REAGENTS :

- SILICA G POWDER ,

- SODIUM ACETATE IN 0.02 M SOLUTION ,

( PREPARATION OF PLATES : MIX 30 G OF SILICA G POWDER AND 60 ML OF 0.02 M SODIUM ACETATE TO OBTAIN A HOMOGENEOUS SUSPENSION . SPREAD ONTO PLATES TO GIVE A THICKNESS OF 0.30 MM . )

- SOLVENT WITH INCORPORATED DEVELOPER .

THE FOLLOWING MIXTURE IS PREPARED :

- ETHYL ACETATE 65 PARTS ,

- ISOPROPANOL 30 PARTS ,

- WATER FIVE PARTS .

JUST BEFORE USE ADD TO THE MIXED SOLVENT :

- 0.3 % OF 2-THIOBARBITURIC ACID ,

- 5.0 % OF CRYSTALLINE TRICHLOROACETIC ACID .

STANDARD SOLUTIONS :

- SUCROSE SOLUTION 0.05 % W/V ,

- SUCROSE AND REDUCING SUGAR SOLUTIONS .

THE FOLLOWING SOLUTIONS ARE PREPARED :

- " A " : SUCROSE SOLUTION 0.5 % ,

- " B " : SOLUTION CONTAINING 5 % OF GLUCOSE AND 5 % OF FRUCTOSE .

MIX 1 ML OF SOLUTION A WITH 1 , 2 , 3 , 4 AND 5 ML RESPECTIVELY OF SOLUTION B IN 10 ML VOLUMETRIC FLASKS . MAKE UP THE SOLUTIONS TO 10 ML WITH DISTILLED WATER . A SERIES OF REFERENCE SOLUTIONS CONTAINING RISING PERCENTAGES OF REDUCING SUGARS IS THUS OBTAINED .

PREPARATION OF SAMPLE

WHEN THE WINE IS STRONGLY COLOURED , TREAT IT WITH DECOLORIZING CHARCOAL BEFORE INOCULATING THE PLATE .

DEVELOPMENT OF THE CHROMATOGRAM

APPLY THE LIQUIDS TO BE EXAMINED TO A STARTING LINE SITUATED 2.5 CM FROM THE EDGE OF THE PLATE TO BE DIPPED INTO THE SOLVENT , AT 3 CM FROM THE LATERAL EDGE ( SIDES ) OF THE PLATE AND 3 CM FROM ONE ANOTHER .

DEPOSIT BETWEEN 1 TO 5 UL OF SAMPLE DEPENDING ON THE CONTENT OF REDUCING SUGARS . EACH SPOT SHOULD NOT CONTAIN MORE THAN 0.25 MG OF REDUCING SUGARS . ADDITIONALLY , APPLY 5 UL OF THE STANDARD SOLUTIONS TO THE PLATE . IN ORDER TO OBTAIN SPOTS OF SMALL DIAMETER IT IS RECOMMENDED THAT SOLUTIONS ARE APPLIED REPEATEDLY IN SMALL AMOUNTS , DRYING AFTER EACH APPLICATION WITH A WEAK JET OF COLD AIR .

THE SILICA LAYER SHOULD BE REMOVED OVER A WIDTH OF 0.5 CM ON THE LATERAL EDGES OF THE PLATE BEFORE PLACING THE PLATE INTO THE CHROMATOGRAPHY TANK CONTAINING THE SOLVENT . ENSURE THAT THE ATMOSPHERE IN THE TANK IS SATURATED WITH SOLVENT .

ALLOW THE SOLVENT TO RUN MORE THAN 16 CM FROM THE STARTING LINE .

DRY THE PLATE IN A CURRENT OF AIR , AFTER WITHDRAWING FROM THE TANK AND PLACE IN THE OVEN FOR 15 MINUTES AT 105* C PARALLEL WITH THE CURRENT OF AIR .

IN THE PRESENCE OF SUCROSE , A YELLOW-ORANGE SPOT APPEARS , THE RF OF WHICH IS IDENTICAL WITH THAT OF SUCROSE IN THE STANDARD SOLUTIONS . GLUCOSE AND FRUCTOSE GIVE A SIMILARLY COLOURED SPOT , THE RF OF WHICH IS GREATER THAN THAT OF SUCROSE .

2 . USUAL METHOD

COLORIMETRIC METHOD

SOLUTIONS :

- NEUTRAL LEAD ACETATE , CRYSTALLINE ,

- HEAVY MAGNESIUM OXIDE , POWDER ,

- POTASSIUM PERMANGANATE SOLUTION , 0.2 % ,

- CHLOROFORM ,

- DIPHENYLAMINE REAGENT . ADD 10 ML DIPHENYLAMINE 10 % IN ABSOLUTE ETHANOL TO 20 ML OF GLACIAL ACETIC ACID AND 70 ML OF CONCENTRATED HYDROCHLORIC ACID , ANALYTICAL REAGENT GRADE * : 1.16 TO 1.19 ) .

THE PURITY OF THE REAGENT SHOULD BE CHECKED ; PLACE 2 ML OF DIPHENYLAMINE REAGENT , TO WHICH HAS BEEN PREVIOUSLY ADDED 2 ML OF DISTILLED WATER , ON THE BOILING WATERBATH FOR FIVE MINUTES . COOL THE SOLUTION RAPIDLY BY IMMERSION IN COLD WATER AND EXTRACT WITH 1 ML OF CHLOROFORM . NO BLUE COLORATION SHOULD APPEAR .

PROCEDURE

DISSOLVE 100 TO 200 MG OF MAGNESIUM OXIDE AND APPROXIMATELY 200 MG OF NEUTRAL LEAD ACETATE IN 10 ML OF WATER AND PLACE ON THE WATERBATH , MAINTAINED AT 90 TO 95* C , FOR THREE TO FIVE MINUTES . ADD 2 ML OF WINE OR MUST , WHICH SHOULD BE CLEAR AND HAVE A TOTAL SUGAR CONTENT OF LESS THAN 1 % .

IF NECESSARY DILUTE THE SAMPLE TO BRING SUGAR CONTENT WITHIN THIS LIMIT .

CHECK THAT SUFFICIENT LEAD ACETATE HAS BEEN ADDED BUT THAT THERE IS NO EXCESS , WHICH WOULD REDUCE THE SENSITIVITY OF THE REACTION . ADD DROPWISE A CONCENTRATED SOLUTION OF NEUTRAL LEAD ACETATE TO THE SUPERNATANT CLEAR LIQUID UNTIL NO MORE PRECIPITATE IS FORMED . STIR THE MIXTURE AND MAINTAIN ON THE WATERBATH UNTIL THE SUPERNATANT LIQUID BECOMES CLEAR . THEN ADD A SATURATED SOLUTION OF SODIUM SULPHATE TO PRECIPITATE THE EXCESS LEAD .

ADD 0.5 ML OF POTASSIUM PERMANGANATE SOLUTION AND MAINTAIN THE MIXTURE ON THE WATERBATH ( 90 TO 95* C ) FOR 10 MINUTES .

COOL RAPIDLY BY IMMERSION IN COLD WATER AND FILTER .

A YELLOW COLORATION MAY DEVELOP , BUT THIS DOES NOT INTERFERE WITH THE REACTION .

PLACE 2 ML OF THE FILTRATE IN A TEST TUBE AND ADD 2 ML OF DIPHENYLAMINE REAGENT ; PLACE ON A BOILING WATERBATH FOR EXACTLY FIVE MINUTES . COOL BY IMMERSION IN COLD WATER , ADD 1 ML OF CHLOROFORM AND IMMEDIATELY EXTRACT THE COLOUR .

DRY WINES , TO WHICH NO SUCROSE HAS BEEN ADDED , GIVE , AT MOST , A VERY SLIGHT GREY-BLUE COLOUR ; A CLEAR BLUE COLOUR DEVELOPS IN THE PRESENCE OF SUCROSE .

SWEET WINES WITH SUGARS , MUSTS FROM MATURE GRAPES AND CONCENTRATED MUSTS PRODUCE , IN THE ABSENCE OF SUCROSE , A GOLDEN YELLOW COLOUR AND IN THE PRESENCE OF SUCROSE , A CLEAR GREEN COLOUR .

THE REACTION IS SENSITIVE TO 0.2 MG OF SUCROSE IN THE TEST SAMPLE .

5.3 . MEASUREMENT AFTER INVERSION

SINGLE REFERENCE AND USUAL METHOD

PROCEDURE

PLACE , IN TWO IDENTICAL FLASKS A AND B , THE SAME TEST SAMPLE OF CLARIFIED SOLUTION OBTAINED USING THE PROCEDURE DESCRIBED UNDER " REDUCING SUGARS " . ADD A VOLUME OF CONCENTRATED HYDROCHLORIC ACID TO FLASKS A AND B IN THE PROPORTION 0.3 ML PER 10 ML OF SUGAR-CONTAINING SOLUTION . IMMEDIATELY ADD THE SAME VOLUME OF 12N SODIUM HYDROXIDE SOLUTION , I . E . 0.3 ML PER 10 ML OF SUGAR SOLUTION , INTO FLASK A AND DETERMINE THE REDUCING SUGARS BY THE PROCEDURE DESCRIBED UNDER " REDUCING SUGARS " .

PLACE FLASK B , WITH THE ACIDIFIED SUGAR-CONTAINING SOLUTION , IN A BOILING WATERBATH FOR TWO MINUTES . ALLOW THE VESSEL TO COOL IN AIR FOR 15 MINUTES . THEN ADD THE SAME VOLUME OF 12N SODIUM HYDROXIDE AS ADDED TO FLASK A AND DETERMINE THE REDUCING SUGARS .

THE DIFFERENCE BETWEEN THE QUANTITIES OF REDUCING SUGARS OBTAINED FROM THESE TWO DETERMINATIONS IS MULTIPLIED BY 0.95 TO GIVE THE SUCROSE CONTENT OF THE TEST SAMPLE . THIS CONCENTRATION IS EXPRESSED IN GRAMS PER LITRE OF WINE , ACCOUNT BEING TAKEN OF THE DILUTIONS THAT MAY HAVE BEEN MADE IN THE COURSE OF CLARIFICATION AND OF THE VOLUME OF THE TEST SAMPLE OF SUGAR SOLUTION BEING EXAMINED .

6 . ASH CONTENT

6.1 . DEFINITION

THE ASH CONTENT IS DEFINED TO BE ALL OF THOSE PRODUCTS REMAINING AFTER IGNITING THE RESIDUE OBTAINED AFTER EVAPORATION OF THE WINE ; THE IGNITION IS CARRIED OUT SUCH THAT ALL OF THE CATIONS ( EXCLUDING AMMONIA ) ARE CONVERTED INTO CARBONATES OR OTHER ANHYDROUS MINERAL SALTS .

6.2 . PRINCIPLE OF THE METHOD

IGNITION OF THE WINE EXTRACT IS CARRIED OUT AT A TEMPERATURE OF 525 MORE OR LESS 25* C UNTIL COMPLETE OXIDATION OF THE CARBON IS OBTAINED .

THE QUANTITY OF ASH SHOULD BE EXPRESSED IN GRAMS PER LITRE AND BE DETERMINED TO THE NEAREST 0.03 G .

6.3 . SINGLE REFERENCE AND USUAL METHOD

PIPETTE 20 ML OF WINE INTO A FLAT-BOTTOMED PLATINUM DISH OF 70 MM DIAMETER AND 25 MM HEIGHT , PREVIOUSLY TARED ON A BALANCE SENSITIVE TO 0.1 MG . CAREFULLY EVAPORATE THE LIQUID , PREFERABLY ON A WATERBATH . HEAT THE RESIDUE GENTLY ON A HOT PLATE OR UNDER AN INFRARED EVAPORATOR ( FOR APPROXIMATELY 30 MINUTES ) UNTIL CARBONIZATION COMMENCES . WHEN EVAPORATION HAS CEASED , PLACE THE DISH IN AN ELECTRIC FURNACE WITH CONTINUOUS VENTILATION AND HEATED TO 525 MORE OR LESS 25* C . WITHDRAW THE DISH AFTER CARBONIZATION ( NORMALLY FIVE MINUTES ) , ADD 5 ML OF WATER , EVAPORATE THE MIXTURE ON A WATERBATH OR UNDER AN EVAPORATOR AND AGAIN HEAT THE RESIDUE TO 525* C .

IF COMBUSTION ( OXIDATION ) OF CARBONIZED PARTICLES IS NOT COMPLETE AFTER 15 MINUTES , REPEAT THE WASHING OPERATION , EVAPORATION OF THE WATER AND RE-IGNITION .

FOR WINES WITH A HIGH SUGAR CONTENT , ADD IF NECESSARY A FEW DROPS OF PURE VEGETABLE OIL TO THE EXTRACT BEFORE THE FIRST ASHING TO PREVENT EXCESSIVE FOAMING . IN THIS CASE PROLONG THE IGNITION FOR A FURTHER 15 MINUTES .

WEIGH THE DISH AFTER COOLING IT IN A DESICCATOR . THE WEIGHT OF THE ASH IS MULTIPLIED BY 50 AND REPORTED AS GRAMS PER LITRE .

7 . ALKALINITY OF THE ASH

7.1 . DEFINITIONS

TOTAL ALKALINITY OF THE ASH IS DEFINED AS THE SUM OF CATIONS OTHER THAN AMMONIUM COMBINED WITH THE ORGANIC ACIDS OF THE WINE .

PARTIAL ALKALINITY OF THE ASH IS THE ALKALINITY FROM THE CARBONATES , OXIDES AND SILICATES ONLY IN THE ASH .

ALKALINITY PER GRAM OF ASH ( OR ALKALINITY VALUE ) IS CALCULATED BY DIVIDING THE TOTAL ALKALINITY EXPRESSED IN GRAMS OF POTASSIUM CARBONATE BY THE WEIGHT OF THE ASH .

7.2 . PRINCIPLE OF METHODS

( A ) TOTAL ALKALINITY

TREATMENT WITH EXCESS SULPHURIC ACID AND BACK TITRATION OF THE EXCESS USING METHYL ORANGE AS INDICATOR .

( B ) PARTIAL ALKALINITY

TREATMENT WITH EXCESS SULPHURIC ACID AND BACK TITRATION OF THE EXCESS IN THE PRESENCE OF CEROUS IONS TO A PH OF 4.5 .

EXPRESSION OF RESULTS

THE ALKALINITY OF THE ASH IS EXPRESSED EITHER IN MILLIEQUIVALENTS PER LITRE AND DETERMINED TO THE NEAREST 0.5 , OR IN GRAMS OF POTASSIUM CARBONATE PER LITRE .

7.3 . TOTAL ALKALINITY

SINGLE REFERENCE AND USUAL METHOD

PROCEDURE

WEIGH THE ASH AND ADD 10 ML OF 0.1N SULPHURIC ACID AND PLACE THE DISH ON A BOILING WATERBATH FOR 15 MINUTES ; REPEATEDLY RUB THE BOTTOM OF THE DISH WITH A GLASS ROD IN ORDER TO ASSIST DISSOLUTION OF THE LESS READILY SOLUBLE MATERIAL . ADD TWO DROPS OF METHYL ORANGE , ONE PART PER 1 000 , AND TITRATE THE EXCESS SULPHURIC ACID AGAINST 0.1N SODIUM HYDROXIDE UNTIL THE COLOUR OF THE INDICATOR CHANGES TO YELLOW . LET THE VOLUME OF SODIUM HYDROXIDE USED BE N ML .

CALCULATIONS

THE ALKALINITY OF ASH IS EQUAL TO :

- 5 ( 10 - N ) EXPRESSED IN MILLIEQUIVALENTS PER LITRE ,

- 0.345 ( 10 - N ) EXPRESSED IN GRAMS OF POTASSIUM CARBONATE PER LITRE .

THE ALKALINITY OF 1 G OF ASH ( ALKALINITY VALUE ) IS EQUAL TO THE RATIO OF THE POTASSIUM CARBONATE ALKALINITY TO THE WEIGHT OF THE ASH .

7.4 . PARTIAL ALKALINITY

SINGLE REFERENCE AND USUAL METHOD

SOLUTIONS

( A ) MIXED INDICATOR : DISSOLVE 0.04 G OF METHYL ORANGE IN 20 ML OF DISTILLED WATER , AND DISSOLVE 0.2 G OF BROMOCRESOL GREEN AND 0.04 G OF METHYL RED IN 80 ML OF ETHANOL . MIX THE TWO SOLUTIONS . THE COLOUR CHANGE TO GREY TAKES PLACE AT PH 4.5 .

( B ) CERIUM CHLORIDE SOLUTION : DISSOLVE 10 G OF ANALYTICAL REAGENT GRADE CECL3.7H2O IN SUFFICIENT DISTILLED WATER ( IN A GRADUATED CYLINDER ) TO MAKE 12.5 ML OF SOLUTION .

PROCEDURE

DISSOLVE THE ASH FROM 20 ML OF WINE IN 10 ML OF 0.1N SULPHURIC ACID . BRING THE SOLUTION TO THE BOIL IN ORDER TO DRIVE OFF THE CO2 . ADD , AFTER COOLING , TWO DROPS OF MIXED INDICATOR , TWO DROPS ( OR FOUR IN THE CASE OF WINES RICH IN PHOSPHATES ) OF CERIUM CHLORIDE , AND TITRATE WITH 0.1N SODIUM HYDROXIDE UNTIL THE COLOUR OF THE INDICATOR CHANGES TO GREY . IN ORDER TO CHECK THAT SUFFICIENT CERIUM CHLORIDE HAS BEEN ADDED , ADD A FURTHER DROP OF CERIUM CHLORIDE SOLUTION , AND IF THE INDICATOR CHANGES FROM RED TO ORANGE , CONTINUE THE TITRATION WITH 0.1N SODIUM HYDROXIDE UNTIL THE COLOUR CHANGES TO GREY .

CALCULATIONS

AS ABOVE .

8 . TOTAL ACIDITY

8.1 . DEFINITION

THE TOTAL ACIDITY IS THE SUM OF THE TITRATABLE ACIDS WHEN THE WINE IS TITRATED TO PH 7 AGAINST A STANDARD ALKALINE SOLUTION . CARBON DIOXIDE AND FREE AND COMBINED SULPHUR DIOXIDE ARE NOT INCLUDED IN THE TOTAL ACIDITY . CARBON DIOXIDE IS REMOVED BY STIRRING WHEN COLD UNDER VACUUM .

8.2 . PRINCIPLE OF METHODS

REFERENCE METHOD : POTENTIOMETRIC TITRATION .

USUAL METHOD : TITRATION WITH BROMOTHYMOL BLUE AS INDICATOR .

EXPRESSION OF RESULTS

THE TOTAL ACIDITY IS EXPRESSED IN MILLIEQUIVALENTS PER LITRE TO THE NEAREST TWO MILLIEQUIVALENTS OR IN GRAMS OF TARTARIC ACID PER LITRE .

8.3 . REFERENCE METHOD

PROCEDURE

( A ) PRELIMINARY ELIMINATION OF CARBON DIOXIDE AND OF SULPHUR DIOXIDE .

FREE THE WINE OF DISSOLVED CARBON DIOXIDE AND SULPHUR DIOXIDE PRESENT . TO ACHIEVE THIS , PLACE 100 ML OF WINE IN A 500 ML FLASK FITTED WITH A REFLUX CONDENSER . REPLACE THE AIR IN THE FLASK BY NITROGEN AND THEN BOIL THE WINE FOR ONE HOUR WHILE PASSING A SLOW STREAM OF NITROGEN . COOL THE APPARATUS WITHOUT INTERRUPTING THE NITROGEN SUPPLY .

( B ) TITRATION .

PIPETTE 20 ML OF WINE INTO A BEAKER AND ADD 0.1N SODIUM HYDROXIDE UNTIL THE PH AS MEASURED AT 20* C WITH A GLASS ELECTRODE IS 7 . THE ADDITION OF THE SODIUM HYDROXIDE SOLUTION SHOULD BE CARRIED OUT SLOWLY AND THE SOLUTION SHOULD BE CONSTANTLY STIRRED . DURATION OF TITRATION MUST BE AT LEAST FIVE MINUTES .

EXPRESSION OF RESULTS

THE RESULT IS EXPRESSED IN MILLIEQUIVALENTS PER LITRE OR IN GRAMS OF TARTARIC ACID PER LITRE .

8.4 . USUAL METHOD

EXPEL THE CARBON DIOXIDE FROM THE WINE BY STIRRING 50 ML IN A ONE LITRE FLASK WHILE APPLYING A VACUUM PRODUCED BY A WATER-JET PUMP . STIRRING SHOULD TAKE PLACE FOR ONE TO TWO MINUTES OR UNTIL LIBERATION OF THE DISSOLVED GAS IS COMPLETED . TAKE A 10 ML ALIQUOT FOR THE TOTAL ACIDITY AND A 20 ML ALIQUOT FOR THE VOLATILE ACIDITY DETERMINATION .

NOTE :

AS THE TOTAL ACIDITY OF THE WINE IS MEASURED AT PH 7 WITH BROMOTHYMOL BLUE AS INDICATOR , A PRELIMINARY TEST ASSISTS IN THE PRODUCTION OF A SUITABLE END-POINT COLOUR AT PH 7 .

ALLOWANCE IS MADE FOR THE ACIDITY OF THE FREE AND COMBINED SULPHUR DIOXIDE BY USING THE FORMULA WHEREBY THE VOLUMES N' AND N'' 0.01N IODINE ARE REQUIRED FOR THE OXIDATION OF FREE AND COMBINED SULPHUR DIOXIDES FOUND IN THE VOLATILE ACIDITY DISTILLATE FROM 20 ML OF WINE ( SEE DETERMINATION OF VOLATILE ACIDITY ) .

SOLUTIONS

( A ) 0.05N SODIUM HYDROXIDE SOLUTION , FREE FROM CO2 .

( B ) BROMOTHYMOL BLUE INDICATOR SOLUTION :

- BROMOTHYMOL BLUE 4 G ,

- NEUTRAL ETHANOL 200 ML .

DISSOLVE AND ADD :

- WATER FREE OF CO2 200 ML ,

- N SODIUM HYDROXIDE SUFFICIENT TO PRODUCE BLUE-GREEN COLOUR ( PH 7 ) 7.5 ML APPROXIMATELY ,

- WATER TO 1 000 ML .

( C ) BUFFER SOLUTION PH 7 :

- MONOPOTASSIUM PHOSPHATE , KH2PO4 107.3 G ,

- SODIUM HYDROXIDE N 500 ML ,

- WATER TO 1 000 ML .

PROCEDURE

1 . PRELIMINARY TEST : END-POINT COLOUR DETERMINATION : PLACE 25 ML OF BOILED DISTILLED WATER INTO AN EVAPORATING DISH OF 12 CM DIAMETER , ADD 1 ML OF BROMOTHYMOL BLUE ( 4 G/L ) AND 5 ML OF CARBON DIOXIDE FREE WINE . NEUTRALIZE WITH SUFFICIENT 0.05N SODIUM HYDROXIDE SO THAT THE COLOUR CHANGES TO GREEN-BLUE . THEN ADD 5 ML OF THE PH 7 BUFFER SOLUTION .

2 . ACTUAL MEASUREMENT : PLACE 30 ML OF BOILED DISTILLED WATER , 1 ML OF BROMOTHYMOL BLUE ( 4 G/L ) , 5 ML OF CARBON DIOXIDE FREE WINE INTO AN EVAPORATING DISH OF 12 CM DIAMETER , AND ADD 0.05N SODIUM HYDROXIDE TO PRODUCE AN IDENTICAL COLOUR TO THAT IN THE CONTROL .

LET THE TITRATION BE N .

CALCULATIONS

THE TOTAL ACIDITY IS EXPRESSED IN MILLIEQUIVALENTS PER LITRE :

10 N - 0.35 N' - 0.25 N''

OR IN GRAMS OF TARTARIC ACID PER LITRE

0.75 ( N - 0.035 N' - 0.025 N'' ) .

NOTES :

1 . IN THE CASE OF WHITE , ROSE OR ALMOST COLOURLESS RED WINES THE QUANTITY OF WINE MAY BE DOUBLED . 10 ML OF WINE AND 0.1N SODIUM HYDROXIDE ARE THEN USED ; THE FORMULAE FOR THE CALCULATION REMAIN THE SAME .

2 . WHEN SEVERAL WINES OF THE SAME ORIGIN ARE TO BE EXAMINED A SINGLE END-POINT COLOUR MAY BE USED FOR ALL THE MEASUREMENTS . THE ABSENCE OF A CHANGE OF COLOUR ON THE ADDITION OF FURTHER BUFFER WILL VERIFY THAT THE END-POINT HAS BEEN REACHED .

9 . VOLATILE ACIDITY

9.1 . DEFINITION

THE VOLATILE ACIDITY DERIVES FROM THE FATTY ACIDS OF THE ACETIC SERIES PRESENT IN WINE EITHER IN THE FREE STATE OR COMBINED AS A SALT .

9.2 . PRINCIPLE OF THE METHOD

SINGLE REFEREE AND ROUTINE METHOD . SEPARATION OF THE VOLATILE ACIDS IS OBTAINED BY STEAM DISTILLATION AND REDISTILLATION OF THE DISTILLATE . THE WINE IS ACIDIFIED WITH TARTARIC ACID ( APPROXIMATELY 0.5 G FOR EACH 20 ML ) PRIOR TO STEAM DISTILLATION . PRECAUTIONS MUST BE TAKEN TO AVOID THE CARBON DIOXIDE IN THE DISTILLATE . THE INDICATOR USED IS PHENOLPHTHALEIN . THE ACIDITY OF DISTILLED FREE AND COMBINED SULPHUR DIOXIDE IS NOT CALCULATED AS VOLATILE ACIDITY AND SHOULD BE DEDUCTED FROM THE ACIDITY OF THE DISTILLATE AS WELL AS ANY ACIDITY FROM SORBIC ACID .

9.3 . EXPRESSION OF RESULTS

THE VOLATILE ACIDITY IS EXPRESSED IN MILLIEQUIVALENTS PER LITRE AND IS DETERMINED TO THE NEAREST 0.5 MILLIEQUIVALENT .

9.4 . SINGLE REFEREE AND USUAL METHOD

PRINCIPLE OF THE APPARATUS AND ITS CONTROL

THE APPARATUS CONSISTS OF A STEAM GENERATOR , A BUBBLER INTO WHICH THE WINE SAMPLE IS PLACED , A RECTIFYING COLUMN AND A CONDENSER .

IN VIEW OF THE LARGE VARIETY AND TYPE OF EQUIPMENT WHICH HAVE BEEN OR WHICH COULD BE PROPOSED , AND IN VIEW OF THE DIFFICULTIES WHICH THE ADOPTION OF A SINGLE TYPE OF EQUIPMENT FOR ALL USERS IN EVERY COUNTRY WOULD PRESENT , IT IS PREFERABLE AND RATIONAL TO SPECIFY IN THE FOLLOWING MANNER THE MINIMUM CONDITIONS WHICH MUST BE MET BY ANY EQUIPMENT OR PROCEDURE PERMITTING THE ACCURATE DETERMINATION OF VOLATILE ACIDITY AS DEFINED :

1 . THE STEAM PRODUCED BY THE GENERATOR MUST BE SUFFICIENTLY FREE OF CARBON DIOXIDE SUCH THAT 250 ML OF DISTILLATE TO WHICH HAS BEEN ADDED 0.1 ML OF N/10 SODIUM HYDROXIDE AND TWO DROPS OF PHENOLPHTHALEIN ( 1 : 100 ) EXHIBITS A STABLE COLORATION FOR AT LEAST 10 SECONDS .

2 . UNDER NORMAL CONDITIONS OF USE , THE RECOVERY OF AN AQUEOUS SOLUTION ACETIC ACID PLACED INTO THE BUBBLER INSTEAD OF WINE SHOULD BE AT LEAST 99.5 % IN THE DISTILLATE .

3 . NOT MORE THAN 5 % OF LACTIC ACID , ADDED AS A NORMAL SOLUTION TO REPLACE THE WINE IN A SIMILAR CONTROL EXPERIMENT , SHOULD APPEAR IN THE DISTILLATE UNDER THE CONDITIONS WHICH PRODUCE A 99.5 % RECOVERY OF ACETIC ACID .

ANY APPARATUS AND PROCEDURE WHICH PERMITS THESE THREE TESTS TO BE CARRIED OUT SUCCESSIVELY AND SUCCESSFULLY FULFILS THE REQUIREMENTS OF OFFICIAL INTERNATIONAL EQUIPMENT AND PROCEDURE .

THE TWO TYPES OF APPARATUS DESCRIBED BELOW MEET THE SPECIFIED CONDITIONS : THE FIRST IS SUITABLE FOR LABORATORIES DEALING WITH LIMITED NUMBERS OF SAMPLES , THE SECOND IS INTENDED FOR LABORATORIES WHERE A LARGE NUMBER OF DETERMINATIONS OF VOLATILE ACIDITY HAVE TO BE CARRIED OUT DAILY AND AS QUICKLY AS POSSIBLE .

1 . SMALL APPARATUS

STEAM IS PRODUCED IN A 1 500 ML FLASK . THE BUBBLER CONSISTS OF A CYLINDRICAL TUBE OF 3 CM DIAMETER AND 27 CM HEIGHT . THIS TUBE IS HEATED EITHER ELECTRICALLY OR BY A FLAME ; THE BASE OF THE BUBBLER TUBE IS INSERTED IN THE 29 MM DIAMETER HOLE CUT IN A SHEET-METAL DISC OF 15 CM DIAMETER AND UNDER WHICH THE FLAME IS PLAYED . THIS PREVENTS ANY OVERHEATING OF EXTRACTIVE MATERIAL IN THE WINE . CONTROL THE AUXILIARY HEATING SO THAT THE VOLUME OF WINE DOES NOT CHANGE BY MORE THAN 5 ML DURING THE COURSE OF THE STEAM-DISTILLATION . THE STEAM FEED TUBE SHOULD END 1 CM ABOVE THE BOTTOM OF THE BUBBLER . PASS VOLATILE ACIDS IN THE STEAM INTO A RECTIFIER COLUMN COMPRISING A CYLINDRICAL TUBE OF 20 MM DIAMETER AND 50 CM HEIGHT CONTAINING A HELIX OF CORRUGATED STAINLESS STEEL SHEET NO 100 AT A PITCH OF 15 MM .

PLACE A WEST CONDENSER , OF 40 CM EFFECTIVE LENGTH , VERTICALLY TO CONDENSE THE VAPOUR .

PROCEDURE

THE STEAM GENERATOR IS FILLED WITH CLEAR LIME WATER OR BARYTA WATER . PLACE 20 ML OF WINE , WHICH HAS BEEN FREED OF CARBON DIOXIDE BY STIRRING UNDER VACUUM AT ROOM TEMPERATURE , IN THE BUBBLER . ADD APPROXIMATELY 0.5 G OF TARTARIC ACID . DISTIL AND COLLECT 250 ML OF THE DISTILLATE IN 12 TO 15 MINUTES .

TITRATION

TITRATE THE ACIDITY AGAINST 0.1N SODIUM HYDROXIDE USING TWO DROPS OF PHENOLPHTHALEIN INDICATOR : ( 1 : 100 IN NEUTRAL ALCOHOL ) ; LET THE VOLUME OF TITRANT USED BE N ML . IMMEDIATELY RESTORE THE ACID CONDITIONS AFTER COMPLETION OF THE ACIDIMETRIC TITRATION BY ADDING A DROP OF CONCENTRATED HYDROCHLORIC ACID . THEN ADD 2 ML OF STARCH PASTE , A CRYSTAL OF POTASSIUM IODIDE AND TITRATE THE FREE SULPHUR DIOXIDE AGAINST AN 0.01N IODINE SOLUTION , LET THE VOLUME USED BE N' ML . NEXT ADD 20 ML OF A SATURATED BORAX SOLUTION ( THE SOLUTION BECOMES PALE PINK ) AND AGAIN ADD 0.01N IODINE SOLUTION UNTIL THE STARCH PASTE BECOMES COLOURED . LET THE VOLUME OF IODINE USED IN THIS SECOND TITRATION , WHICH REPRESENTS THE SULPHUR DIOXIDE COMBINED WITH ACETALDEHYDE , BE N'' .

THE VOLATILE ACIDITY EXPRESSED IN MILLIEQUIVALENTS PER LITRE IS EQUAL TO :

5 ( N - 0.1 N' - 0.05 N'' )

NOTES :

1.1 . WITH SAMPLES PRESERVED WITH SALICYLIC ACID , DISTIL THIS ACID , DETERMINE COLORIMETRICALLY AT A PH OF 3 MORE OR LESS 0.5 , AND SUBTRACT ITS VALUE FROM THIS FIGURE .

1.2 . COMPENSATION MUST BE MADE FOR THE PRESENCE OF SORBIC ACID IN WINE AS UNDER THE CONDITIONS OF THE EXPERIMENT IT IS ALMOST COMPLETELY DISTILLED . IT IS DETERMINED IN A SMALL PORTION OF THE DISTILLATE TAKEN BEFORE TITRATION . THE VOLUME OF THIS DISTILLATE ( 250 TO 300 ML ) MAY BE MEASURED IN A MEASURING CYLINDER ) .

TAKE 0.5 ML OF THE DISTILLATE AND TRANSFER TO A 1 CM SILICA SPECTROPHOTOMETRIC CELL AND ADD 1.5 ML OF SOLUTION A BELOW ( 16 ) ; AND MIX : ( SEE OJ )

MEASURE THE OPTICAL DENSITY AT 256 NM AFTER A FEW MINUTES AGAINST WATER AND A STANDARD SORBIC ACID SOLUTION OF 20 MG/L ( 17 ) , 0.5 ML OF WHICH IS PLACED IN A 1 CM CELL WITH 1.5 ML OF SOLUTION A AND MIXED .

THIS METHOD IS APPLICABLE TO CONCENTRATIONS OF 10 TO 300 MG/L SORBIC ACID . THE ACCURACY OF THE DETERMINATION IS APPROXIMATELY 1 % .

1 G OF SORBIC ACID IS EQUIVALENT TO 8.92 MILLIEQUIVALENTS AND 0.438 G OF SULPHURIC ACID . FOR ANY WINE TO WHICH 200 MG/L SORBIC ACID HAS BEEN ADDED , THE APPROPRIATE CORRECTION FOR THE VOLATILE ACIDITY IS 1.7 MILLIEQUIVALENTS PER LITRE OF WINE .

2 . RAPID DISTILLATION APPARATUS

THE STEAM GENERATOR CONSISTS OF A COPPER BOILER OF 20 LITRE CAPACITY FED CONTINUOUSLY WITH CLEAR DILUTE LIME WATER . IT IS HEATED MODERATELY VIGOROUSLY SO THAT ONE KILOGRAM OF STEAM IS PRODUCED IN 12 MINUTES AT A PRESSURE NOT GREATER THAN THAT OF A 50 CM WATER COLUMN .

THE BUBBLER HAS A RECTIFER COLUMN ATTACHED TO IT . IT CONSISTS OF A U-TUBE , ONE BRANCH OF WHICH IS THE STEAM SUPPLY TUBE WITH AN INSIDE DIAMETER OF 10 MM CONNECTED TO THE STEAM PIPE FROM THE BOILER . THE SECOND BRANCH CONSISTS OF A CYLINDRICAL VESSEL OF 26 MM DIAMETER AND 100 MM HEIGHT . THE STEAM IS INITIALLY DISPERSED AT THE BASE OF THE VESSEL THROUGH A FINNED PLATE . TO THIS CYLINDER ARE FUSED TWO BULBS , ONE OF 500 ML AND THE OTHER OF 1 000 ML , ON TOP OF ONE ANOTHER . THE WINE IS INTRODUCED THROUGH A LATERAL NOZZLE FITTED WITH A PLUG . THERE IS AN OPENING AT THE LOWEST POINT OF THE U-TUBE THROUGH WHICH RESIDUES ARE REMOVED AT THE COMPLETION OF A DISTILLATION . THROUGHOUT THE DISTILLATION THIS OPENING IS KEPT CLOSED WITH A RUBBER BUNG ATTACHED TO A LEVER . THE CYLINDRICAL PART IS HEATED ELECTRONICALLY BY A NICKEL-CHROME STEEL STRIP ( ELEMENT ) ; 350 W IS PRODUCED USING A 24 V SUPPLY . THE WATER IS EVAPORATED AT THE SAME RATE AS IT IS CONDENSING IN THE RECTIFIER COLUMN .

THE RECTIFIER COLUMN IS A COLUMN WITH A HELIX OF CORRUGATED STAINLESS STEEL GAUZE NO 100 AND IS 80 CM HIGH WITH AN INTERNAL DIAMETER OF 20 MM .

THE CONDENSER CONSISTS OF A STAINLESS STEEL TUBE WITH THIN WATER JACKET .

THIS APPARATUS MAY BE PROVIDED WITH DIFFERENT CONTROL AND INTERLOCKING DEVICES TO PREVENT FAULTY OPERATION AND TO ALLOW SEMI-AUTOMATIC FUNCTIONING IF CONSIDERED APPROPRIATE . 250 ML OF DISTILLATE CAN BE OBTAINED IN FOUR MINUTES USING THIS APPARATUS .

THE MEASUREMENT TAKES PLACE AS DESCRIBED PREVIOUSLY .

10 . FIXED ACIDITY

THE FIXED ACIDITY IS CALCULATED FROM THE DIFFERENCE BETWEEN TOTAL ACIDITY AND VOLATILE ACIDITY .

11 . TARTARIC ACID

11.1 . PRINCIPLE OF METHODS

( A ) REFERENCE METHOD

TARTARIC ACID IS PRECIPITATED IN THE FORM OF CALCIUM DL-TARTRATE AND DETERMINED GRAVIMETRICALLY . THIS DETERMINATION MAY BE COMPLETED BY A VOLUMETRIC PROCEDURE FOR COMPARISON . THE CONDITIONS FOR PRECIPITATION ( PH , TOTAL VOLUME USED , CONCENTRATIONS OF PRECIPITATING SOLUTIONS ) ARE SUCH THAT PRECIPITATION OF THE CALCIUM DL-TARTRATE IS COMPLETE WHEREAS THE CALCIUM ( - ) TARTRATE REMAINS IN SOLUTION .

WHEN METATARTARIC ACID HAS BEEN ADDED TO THE WINE , IT MUST FIRST BE HYDROLYZED .

( B ) USUAL METHOD

THE TARTARIC ACID , SEPARATED USING AN ION EXCHANGE COLUMN , IS DETERMINED COLORIMETRICALLY IN THE ELUATE BY MEASUREMENT OF THE RED COLOUR PRODUCED ON REACTION WITH VANADIC ACID . THE ELUATE ALSO CONTAINS LACTIC AND MALIC ACIDS WHICH DO NOT INTERFERE .

EXPRESSION OF RESULTS

THE TARTARIC ACID IS EXPRESSED IN MILLIEQUIVALENTS PER LITRE AND IN GRAMS OF TARTARIC ACID PER LITRE . ACCURACY OF DETERMINATION IS 0.5 MILLIEQUIVALENT PER LITRE .

11.2 . REFERENCE METHOD

REAGENTS

CALCIUM ACETATE SOLUTION CONTAINING 10 G/L OF CALCIUM :

- CALCIUM CARBONATE , ANALYTICAL REAGENT GRADE 25 G ,

- ACETIC ACID , GLACIAL 40 ML ,

- WATER TO 1 000 ML .

CALCIUM DL-TARTRATE , CRYSTALLIZED :

PLACE 20 ML OF ( + ) TARTARIC ACID SOLUTION ( 5 G/L ) INTO A 400 ML BEAKER . ADD 20 ML OF AMMONIUM ( - ) TARTRATE SOLUTION ( 6.126 G/L ) AND 6 ML OF CALCIUM ACETATE SOLUTION ( 10 G OF CALCIUM PER LITRE ) .

ALLOW TO STAND FOR TWO HOURS TO PRECIPITATE . COLLECT THE PRECIPITATE IN A SINTERED GLASS CRUCIBLE OF POROSITY NO 4 , AND WASH IT THREE TIMES WITH DISTILLED WATER , THE TOTAL WASHINGS TO AMOUNT TO ABOUT 30 ML . DRY TO CONSTANT WEIGHT IN THE OVEN AT 70* C . USING THE QUANTITIES OF REAGENT INDICATED ABOVE ABOUT 340 MG OF CRYSTALLIZED CALCIUM DL-TARTRATE IS OBTAINED . STORE IN A STOPPERED BOTTLE .

PRECIPITATION SOLUTION ( PH 4.75 ) :

- AMMONIUM ( - ) TARTRATE 150 MG ,

- CALCIUM ACETATE SOLUTION ( 10 G OF CALCIUM PER LITRE ) 8.8 ML ,

- WATER TO 1 000 ML .

DISSOLVE THE AMMONIUM ( - ) TARTRATE IN 900 ML OF WATER ; ADD 8.8 ML OF CALCIUM ACETATE SOLUTION AND MAKE UP TO ONE LITRE . SINCE CALCIUM DL-TARTRATE IS SLIGHTLY SOLUBLE IN THIS SOLUTION , ADD 5 MG OF CALCIUM DL-TARTRATE PER LITRE , STIR FOR 12 HOURS AND FILTER .

PROCEDURE

PLACE 500 ML OF PRECIPITATION SOLUTION AND 10 ML OF WINE INTO A 600 ML BEAKER . MIX BY RUBBING THE SIDES OF THE VESSEL WITH THE TIP OF A GLASS ROD . LEAVE TO PRECIPITATE FOR 12 HOURS ( OVERNIGHT ) .

FILTER THE LIQUID AND PRECIPITATE THROUGH A WEIGHED SINTERED GLASS CRUCIBLE OF POROSITY NO 4 FITTED ON A CLEAN VACUUM FLASK . RINSE THE VESSEL IN WHICH PRECIPITATION TOOK PLACE WITH THE FILTRATE TO ENSURE THAT ALL PRECIPITATE IS TRANSFERRED .

DRY TO CONSTANT WEIGHT IN AN OVEN AT 70* C . WEIGH : LET P BE THE WEIGHT OF CRYSTALLIZED CALCIUM DL-TARTRATE ( CAC4O6H4 , 4H2O ) OBTAINED .

WINES TO WHICH METATARTARIC ACID HAS BEEN ADDED

WHEN ANALYZING WINE TO WHICH METATARTARIC ACID HAS BEEN OR IS SUSPECTED OF HAVING BEEN ADDED , PROCEED BY FIRST HYDROLYZING THIS ACID AS FOLLOWS :

PLACE 10 ML OF WINE AND 0.4 ML OF GLACIAL ACETIC ACID INTO A 50 ML CONICAL FLASK . PLACE A REFLUX CONDENSER ON TOP OF THE FLASK AND BOIL FOR 30 MINUTES . ALLOW TO COOL AND THEN TRANSFER THE SOLUTION IN THE CONICAL FLASK TO A 600 ML BEAKER ; RINSE THE FLASK TWICE USING 5 ML OF WATER EACH TIME AND THEN CONTINUE AS DESCRIBED ABOVE .

METATARTARIC ACID IS CALCULATED AND INCLUDED AS TARTARIC ACID IN THE FINAL RESULT .

CALCULATION AND EXPRESSION OF RESULTS

ONE MOLECULE OF CALCIUM DL-TARTRATE CORRESPONDS TO HALF A MOLECULE OF ( + ) TARTARIC ACID IN THE WINE .

THE QUANTITY OF TARTARIC ACID PER LITRE OF WINE IS EQUAL TO :

384.5 P EXPRESSED IN MILLIEQUIVALENTS ,

28.84 P EXPRESSED IN GRAMS OF TARTARIC ACID ,

36.15 P EXPRESSED IN GRAMS OF POTASSIUM ACID TARTRATE .

COMPARATIVE VOLUMETRIC ANALYSIS

REAGENTS

- HYDROCHLORIC AND ( 1 : 5 V/V ) ( ABOUT 82 G/L )

- SOLUTION , EDTA 0.05 M :

- EDTA ( ETHYLENEDIAMINETETRAACETIC ACID DISODIUM SALT ) ON COMPLEXON III 18.61 G ,

- DISTILLED WATER TO 1 000 ML .

- SODIUM HYDROXIDE SOLUTION , 40 % ( M/V ) :

- SODIUM HYDROXIDE , ANALYTICAL REAGENT GRADE 40 G ,

- DISTILLED WATER TO 100 ML .

- COMPLEXOMETRIC INDICATOR ( 1 % ) :

- 2-HYDROXY-1 - ( 2-HYDROXY-4-SULPHO-1-NAPHTHYLAZO ) -3-NAPHTHOIC ACID ( PATTON AND REEDER'S INDICATOR ) 1 G ,

- SODIUM SULPHATE , ANHYDROUS 100 G .

PROCEDURE

WEIGH THE PRECIPITATE OF CALCIUM DL-TARTRATE CONTAINED IN THE CRUCIBLE AND THEN DISSOLVE IN 10 ML OF DILUTE HYDROCHLORIC ACID .

WASH THE SINTERED GLASS CRUCIBLE WITH 50 ML OF DISTILLED WATER .

ADD 5 ML OF 40 % SODIUM HYDROXIDE SOLUTION AND 30 MG OF INDICATOR . TITRATE WITH 0.05 M EDTA SOLUTION . LET THE NUMBER OF MILLILITRES USED BE N .

CALCULATION AND EXPRESSION OF RESULTS

THE QUANTITY OF TARTARIC ACID PER LITRE OF WINE IS EQUAL TO :

5N EXPRESSED IN MILLIEQUIVALENTS ,

0.375N EXPRESSED IN GRAMS OF TARTARIC ACID ,

0.470N EXPRESSED IN GRAMS OF POTASSIUM TARTRATE .

11.3 . USUAL METHOD

SOLUTIONS

1 . PRELIMINARY TREATMENT OF THE WINE

- AN ION EXCHANGE RESIN OF MERCK BASICITY STRENGTH III : THE EXCHANGE RESIN SHOULD BE IN THE ACETATE FORM . THIS IS OBTAINED BY KEEPING IT FOR AT LEAST 24 HOURS IN 30 % ACETIC ACID . IDEALLY , HOWEVER , A MORE SUBSTANTIAL STOCK IS MAINTAINED BY KEEPING THE RESIN UNDER 30 % ACETIC ACID ( 200 ML OF 30 % ACETIC ACID IS POURED OVER APPROXIMATELY 100 G OF EXCHANGE RESIN ) .

- ACETIC ACID 30 % : MAKE UP 300 ML OF GLACIAL ACETIC ACID TO ONE LITRE WITH DISTILLED WATER .

- ACETIC ACID 0.5 % : MAKE UP 5 ML OF GLACIAL ACETIC ACID TO ONE LITRE WITH DISTILLED WATER .

- SODIUM SULPHATE SOLUTION 7.1 % ( 0.5 M ) : A LARGE VOLUME OF THIS REAGENT IS REQUIRED FOR ELUTION . DISSOLVE 355 G OF ANHYDROUS SODIUM SULPHATE ( NA2SO4 ) IN WATER AND MAKE UP TO FIVE LITRES WITH WATER .

2 . DETERMINATION OF THE TARTARIC ACID

- VANADIC REAGENT : DISSOLVE 10 G OF AMMONIUM METAVANADATE IN 150 ML OF N SODIUM HYDROXIDE SOLUTION . TRANSFER THE SOLUTION TO A 500 ML VOLUMETRIC FLASK AND ADD 200 ML OF A 27 % SODIUM ACETATE SOLUTION . MAKE UP TO 500 ML WITH DISTILLED WATER .

- STANDARD 2N H2SO4 SOLUTION .

- STANDARD 0.1N H2SO4 SOLUTION .

- 0.05 M PERIODIC ACID SOLUTION ( 0.1N ) : PUT 10.696 G OF SODIUM PERIODATE AND 50 ML OF N SULPHURIC ACID INTO A ONE LITRE VOLUMETRIC FLASK AND MAKE UP TO ONE LITRE WITH DISTILLED WATER .

- GLYCEROL SOLUTION 10 % : TAKE 10 G OF ANALYTICAL REAGENT , TWICE DISTILLED GLYCEROL AND MAKE UP TO 100 ML WITH DISTILLED WATER .

- SODIUM SULPHATE SOLUTION , 7.1 % : DISSOLVE 71 G OF ANHYDROUS SODIUM SULPHATE IN DISTILLED WATER . MAKE UP TO ONE LITRE .

- TARTARIC ACID SOLUTION , 1 G/L :

TARTARIC ACID PURE FOR ANALYSIS 0.500 G ,

SODIUM HYDROXIDE SOLUTION , N 6.66 ML ,

SODIUM SULPHATE SOLUTION 7.1 % TO 500 ML .

PROCEDURE

PREPARATION OF THE ION EXCHANGE COLUMN

USE A GLASS TUBE OF 10 TO 11 MM INTERNAL DIAMETER AND APPROXIMATELY 300 MM LONG WITH A DRAIN TAP ATTACHED AT ITS END BY MEANS OF A DRILLED RUBBER BUNG . THE GLASS TUBE OF THE TAP MUST NOT EXTEND ABOVE THE INNER SURFACE OF THE BUNG SO AVOIDING ANY DEAD SPACE .

COVER THE RUBBER BUNG WITH A GLASS-WOOL PLUG OF ABOUT 2 TO 3 MM HEIGHT , FILL THE TUBE WITH WATER AND THEN ALLOW TO DRAIN TO APPROXIMATELY 5 MM ABOVE THE GLASS-WOOL . FILL THE TUBE WITH APPROXIMATELY 10 ML OF A SUSPENSION OF ION EXCHANGE RESIN IN ITS ACETATE FORM , AVOIDING AIR BUBBLES . TO ACHIEVE THIS , SUSPEND THE EXCHANGE RESIN , KEPT IN 30 % ACETIC ACID , BY STIRRING AND POUR RAPIDLY INTO THE TUBE THROUGH A FUNNEL . ALLOW THE ION EXCHANGE RESIN TO SETTLE . PLACE A SMALL PLUG OF GLASS-WOOL ON TOP OF THE RESIN TO PREVENT IT BEING DISTURBED DURING SUBSEQUENT WASHINGS . THE EXCHANGE RESIN MAY ONLY BE USED ONCE .

FOR SUBSEQUENT DETERMINATIONS REMOVE THE SPENT EXCHANGE RESIN BY BLOWING OUT WITH WATER FROM A TAP . THE GLASS-WOOL PLUG AT THE BOTTOM WILL NORMALLY BE RETAINED .

SEPARATION OF THE TARTARIC ACID USING AN ION EXCHANGE COLUMN

OPEN THE DRAIN TAP FULLY AND ALLOW 30 % ACETIC ACID TO FLOW DOWN THE COLUMN TO WITHIN APPROXIMATELY 2 TO 3 MM OF THE UPPER GLASS-WOOL PLUG . ADD APPROXIMATELY 10 ML OF 0.5 % ACETIC ACID AND MARK THE LIQUID LEVEL WITH A GREASE PENCIL . OPEN THE TAP WIDE AND ALLOW THE LIQUID TO DRAIN AGAIN DOWN TO WITHIN 2 TO 3 MM OF THE UPPER GLASS-WOOL PLUG . REPEAT THIS WASHING OPERATION FOUR MORE TIMES WITH APPROXIMATELY 10 ML 0.5 % ACETIC ACID , REFILLING THE COLUMN EACH TIME TO THE REFERENCE MARK .

AFTER THE LAST WASH CLOSE THE TAP AND POUR 10 ML OF THE WINE OR MUST ON TO THE EXCHANGE RESIN . ( THE PRESENCE OF SUGAR , ALCOHOL , COLOURING MATTER OR TANNINS DOES NOT AFFECT THE VOLUME OF WINE TO BE TAKEN . )

PASS THE WINE THROUGH DROP BY DROP SO THAT A FLOW RATE OF ONE TO 1.5 DROPS PER SECOND ( CORRESPONDING TO A FLOW OF 25 TO 30 ML IN 10 MINUTES ) IS NOT EXCEEDED ; STOP THE FLOW JUST ABOVE THE LEVEL OF THE UPPER GLASS-WOOL PLUG . FILL THE TUBE AGAIN WITH 0.5 % ACETIC ACID UP TO THE REFERENCE MARK . ALLOW TO DRAIN AT THE SAME RATE AS PREVIOUSLY AND SUBSEQUENTLY WASH SEVEN TIMES IN THE SAME MANNER USING 10 ML OF WATER EACH TIME , TAKING CARE TO WASH DOWN THE WALLS OF THE TUBE TO THE REFERENCE MARK ON EACH OCCASION .

CLOSE THE TAP AS SOON AS THE LIQUID LEVEL IS JUST BELOW THE UPPER GLASS-WOOL PLUG DURING THE LAST WASHING .

USE A GRADUATED FLASK OF 100 ML AS A RECEIVER FOR THE FOLLOWING OPERATION . ELUTE THE ACIDS ABSORBED TO THE EXCHANGE MATERIAL WITH A 0.5 M SODIUM SULPHATE SOLUTION . FOR THIS PURPOSE FILL A TUBULATED BOTTLE WITH 0.5 M SODIUM SULPHATE SOLUTION AND CONNECT ITS LOWER TUBE TO THE ION EXCHANGE COLUMN TUBE WITH RUBBER TUBING ( USING AN ADJUSTABLE SCREW-CLIP , A DRAWN-OUT TUBE AND A RUBBER BUNG ) . WHEN THE TWO PIECES OF APPARATUS ARE JOINED , OPEN THE SCREW-CLIP , RAISE THE RUBBER BUNG UNTIL THE TUBE CONTAINING THE ION EXCHANGE RESIN IS FILLED TO A HEIGHT OF APPROXIMATELY 10 CM WITH SODIUM SULPHATE SOLUTION . THE TWO PIECES OF APPARATUS WILL NOW BE JOINED . WHEN THE TAP IS OPENED SODIUM SULPHATE SOLUTION WILL RUN OUT AT THE SAME RATE AS THE TUBE REFILLS . THE FLOW IS REGULATED AT A RATE OF TWO TO THREE DROPS PER SECOND UNTIL THE RECEIVER FLASK HAS BEEN FILLED UP TO ITS CALIBRATION MARK .

DETERMINATION OF TARTARIC ACID

PLACE 20 ML OF THE ELUATE IN TWO 100 ML CONICAL FLASKS A AND B .

USE FLASK A FOR THE DETERMINATION ON SAMPLE AND FLASK B , FOR THE BLANK EXPERIMENT , IN WHICH THE TARTARIC ACID HAS BEEN DESTROYED BY PERIODIC ACID .

PIPETTE INTO FLASK A 2 ML OF 2N H2SO4 5 ML OF 1.1N H2SO4 AND 1 ML OF 10 % GLYCEROL .

PIPETTE INTO FLASK B 2 ML OF 2N H2SO4 5 ML OF 0.5N PERIODIC ACID ; ALLOW TO STAND FOR 15 MINUTES AND ADD 1 ML OF 10 % GLYCEROL SOLUTION TO DESTROY THE EXCESS OF PERIODIC ACID . ALLOW TO STAND FOR TWO MINUTES .

WITH STIRRING , PIPETTE 5 ML OF THE VANADIC REAGENT FIRST INTO FLASK B AND THEN IMMEDIATELY AFTERWARDS INTO FLASK A . MIX . IMMEDIATELY START A STOP-WATCH AND POUR PART OF THE CONTENTS OF FLASKS A AND B INTO TWO MATCHED 10 MM SPECTROPHOTOMETER CELLS . AFTER ONE MINUTE 30 SECONDS RECORD THE OPTICAL DENSITY OF THE LIQUID FROM FLASK A MEASURED AT 490 NM USING THAT FROM FLASK B AS THE BLANK .

PLOTTING THE CALIBRATION CURVE

PIPETTE 10 , 20 , 30 , 40 AND 50 ML ALIQUOTS OF THE STANDARD TARTARIC ACID SOLUTION PREPARATION ( SEE PAGE 49 ) ( 1 G/L ) INTO CALIBRATED 100 ML FLASKS AND MAKE UP TO VOLUME WITH THE 7.1 % SODIUM SULPHATE SOLUTION . THESE SOLUTIONS CORRESPOND TO WINE ELUATES CONTAINING 1 , 2 , 3 , 4 AND 5 G/L OF TARTARIC ACID .

PLACE 20 ML OF EACH OF THE STANDARD SOLUTIONS OF TARTARIC ACID IN TWO CONICAL 100 ML FLASKS A AND B AND TREAT AS DESCRIBED ABOVE FOR THE WINE ELUATE .

A GRAPH OF THE OPTICAL DENSITIES OF THESE SOLUTIONS AS A FUNCTION OF THEIR TARTARIC ACID CONCENTRATION IS A STRAIGHT LINE CURVING SLIGHTLY INWARDS TOWARDS THE ORIGIN .

DRAW THIS PART OF THE CALIBRATION CURVE MORE CAREFULLY BY REPEATING THE DETERMINATIONS USING SOLUTIONS CONTAINING 0.1 , 0.2 , 0.3 , 0.4 , 0.5 , 0.6 , 0.7 , 0.8 , 0.9 AND 1 G/L OF TARTARIC ACID .

12 . CITRIC ACID

SINGLE METHOD

12.1 . PRINCIPLE

CITRIC ACID IS PRECIPITATED BY THE BARIUM ION IN AN AQUEOUS ALCOHOLIC ALKALINE MEDIUM . AFTER DECOLORATION OF THE BARIUM PRECIPITATE IN AQUEOUS SOLUTION , BY ACTIVATED CHARCOAL , AND OXIDATION BY LEAD ( IV ) ACETATE , A YELLOW COMPOUND IS FORMED IN THE PRESENCE OF DIAZOSULPHANILIC ACID WHICH IS DETERMINED COLORIMETRICALLY AT 420 NM .

12.2 . REAGENTS ( 18 )

- CONCENTRATED AMMONIA : ( * = 0.910 ) .

- BARIUM CHLORIDE SOLUTION , 20 % : DISSOLVE 20 G BARIUM CHLORIDE , BACL2.2H2O IN DISTILLED WATER AND MAKE UP TO 100 ML .

- WASHING LIQUID : MIX 140 ML WATER WITH 300 ML ALCOHOL ( 96 % BY VOLUME ) .

- SODIUM SULPHATE SOLUTION , 7.1 % W/V : DISSOLVE 71 G ANHYDROUS SODIUM SULPHATE IN WATER AND MAKE UP TO ONE LITRE .

- ACTIVATED CHARCOAL .

- SODIUM ACETATE SOLUTION , 27 % : DISSOLVE 270 G ANHYDROUS SODIUM ACETATE IN WATER AND MAKE UP TO ONE LITRE .

- SODIUM NITRITE SOLUTION , 2 % : DISSOLVE 2 G CRYSTALLINE SODIUM NITRITE IN WATER AND MAKE UP TO 100 ML ( THIS SOLUTION MAY BE STORED FOR A LONG PERIOD IN A BROWN BOTTLE ) .

- SULPHANILIC ACID SOLUTION : DISSOLVE 1.5 G SULPHANILIC ACID IN 50 ML GLACIAL ACETIC ACID AND MAKE UP TO 250 ML WITH WATER . SHAKE FREQUENTLY AFTER THE PREPARATION AND ALLOW TO STAND OVERNIGHT . STORE IN BROWN BOTTLES . THIS SOLUTION KEEPS INDEFINITELY .

- SATURATED SOLUTION OF LEAD ( IV ) ACETATE : PLACE ABOUT 50 G LEAD ( IV ) ACETATE ( LEAD TETRA-ACETATE , MOISTENED WITH GLACIAL ACETIC ACID ) IN A BROWN BOTTLE AND ADD APPROXIMATELY 250 ML GLACIAL ACETIC ACID . SHAKE AND ALLOW THE NON-DISSOLVED PART OF THE LEAD TETRA-ACETATE TO SETTLE SO THAT A CLEAR SUPERNATANT LIQUID IS OBTAINED . PROVIDING IT IS STORED IN A BROWN BOTTLE , THE SOLUTION KEEPS INDEFINITELY . FURTHER SATURATED SOLUTION CAN BE PREPARED BY ADDING GLACIAL ACETIC ACID TO THE RESIDUE AND CONTINUING AS ABOVE .

- LEAD ( IV ) ACETATE SOLUTION , 1 % : INTRODUCE 5 ML OF THE SATURATED LEAD ( IV ) ACETATE SOLUTION INTO A 250 ML CONICAL FLASK AND ADD ALL TOGETHER 50 ML OF 10 % POTASSIUM IODIDE SOLUTION . TITRATE IMMEDIATELY WITH 0.1 N SODIUM THIOSULPHATE IN THE PRESENCE OF STARCH SOLUTION . LET N BE THE NUMBER OF MILLILITRES ADDED .

TAKE A VOLUME OF SATURATED LEAD ( IV ) ACETATE SOLUTION EQUAL TO * ML AND MAKE UP TO ONE LITRE WITH GLACIAL ACETIC ACID . STORE IN BROWN BOTTLES . THIS SOLUTION MAY BE KEPT INDEFINITELY .

12.3 . PROCEDURE

ISOLATION OF CITRIC ACID

PIPETTE INTO A 40 TO 50 ML CENTRIFUGE TUBE 5 ML WINE OR MUST , 1 ML CONCENTRATED AMMONIA AND 1 ML 20 % BARIUM CHLORIDE . MIX WITH A GLASS ROD . AFTER APPROXIMATELY TWO MINUTES ADD 15 ML ALCOHOL ( 96 % BY VOLUME ) AND MIX AGAIN WITH THE GLASS ROD . WAIT FIVE MINUTES AND THEN CENTRIFUGE FOR THREE TO FOUR MINUTES . DISCARD THE SUPERNATANT LIQUID AND THEN WASH THE PRECIPITATE AS FOLLOWS : TAKE 2 ML OF THE WASHING LIQUID USING A 2 ML PIPETTE ; APPLY THE POINT OF THE PIPETTE TO THE TOP OF THE TUBE WALL , EMPTY THE PIPETTE BY BLOWING OUT WHILE AT THE SAME TIME ROTATING THE TUBE ABOUT ITS AXIS . MIX THE PRECIPITATE AND WASHING LIQUID TO FORM A SUSPENSION USING A GLASS ROD FITTED WITH A RUBBER COVERED TIP . RINSE THE RUBBER TIP TWICE WITH A FURTHER 5 ML WASHING LIQUID , BLOWING FIERCELY DOWN THE PIPETTE SO THAT THE JET OF LIQUID REMOVES ANY PRECIPITATE ADHERING TO THE RUBBER . CENTRIFUGE AGAIN AND DISCARD THE CLEAR SUPERNATANT SOLUTION . WASH THE PRECIPITATE A SECOND TIME .

COVER THE PRECIPITATE , STILL WET WITH THE ALCOHOLIC WASHING LIQUID , WITH ABOUT 10 ML OF THE 7.1 % SODIUM SULPHATE SOLUTION . PLACE THE CENTRIFUGE TUBE IN A BOILING WATERBATH FOR 10 MINUTES AND CRUSH THE PRECIPITATE WITH A GLASS ROD SO THAT NO LUMPS REMAIN .

TRANSFER QUANTITATIVELY THE CONTENTS OF THE STILL-WARM TUBE TO A 50 ML VOLUMETRIC FLASK USING THE 7.1 % SODIUM SULPHATE SOLUTION . ALLOW TO COOL AND MAKE UP TO THE MARK WITH SODIUM SULPHATE SOLUTION , MIX AND POUR THE CONTENTS OF THE FLASK INTO A 100 ML CONICAL FLASK CONTAINING 0.2 G ACTIVATED CHARCOAL . STIR . ALLOW TO STAND FOR FIVE MINUTES AND FILTER THROUGH A SINTERED GLASS CRUCIBLE .

THE FILTRATE IS THE TEST SOLUTION CONTAINING THE CITRIC ACID FROM THE WINE OR MUST AT 10-FOLD DILUTION .

DETERMINATION OF CITRIC ACID

TAKE TWO 50 ML CONICAL FLASKS A AND B AND INTRODUCE 10 ML 27 % SODIUM ACETATE SOLUTION INTO EACH . PIPETTE 2 ML OF THE TEST SOLUTION TO EACH OF THE FLASKS .

PREPARE THE DIAZOTIZATION SOLUTION BY THE ADDITION , WITH SHAKING , OF 5 ML OF SULPHANILIC ACID TO 1 ML SODIUM NITRITE SOLUTION .

PIPETTE INTO FLASK A 2 ML OF THE DIAZOTIZATION SOLUTION WHILE SWIRLING THE FLASK CONTINUOUSLY AND THEN 5 ML GLACIAL ACETIC ACID ; PIPETTE INTO FLASK B 2 ML OF THE DIAZOTIZATION SOLUTION AND 5 ML OF 1 % LEAD ( IV ) ACETATE SOLUTION . IMMEDIATELY START A STOPCLOCK .

AFTER APPROXIMATELY FIVE MINUTES , FILTER THE CONTENTS OF FLASKS A AND B THROUGH FLUTED FILTER PAPERS TO AID RAPID FILTRATION . AFTER NEARLY 13 MINUTES , POUR THE FILTRATES INTO TWO PARALLEL SIDED CURVETTES WITH 30 MM PATH LENGTH AND PLACE IN THE SPECTROPHOTOMETER . AFTER EXACTLY 13 MINUTES , MEASURE THE ABSORBANCE AT 420 NM OF THE LIQUID FROM FLASK A USING THE LIQUID FROM FLASK B AS A BLANK .

IF IT IS FOUND THAT THE TEST SOLUTION CONTAINS A CITRIC ACID CONCENTRATION SUCH THAT THE ABSORBANCE IS TOO GREAT , DILUTE THE TEST SOLUTION FURTHER WITH THE 7.1 % SODIUM SULPHATE SOLUTION .

CALCULATE THE CITRIC ACID CONCENTRATION IN THE SOLUTION BY REFERENCE TO THE STANDARD CURVE GIVING THE CONCENTRATION OF CITRIC ACID C6H8O7 AS A FUNCTION OF ASORBANCE .

PREPARATION OF THE STANDARD CURVE

DISSOLVE 273.4 MG OF PURE CITRIC ACID MONOHYDRATE C6H8O7.H2O ( = 250 MG C6H8O7 ) IN A 7.1 % SODIUM SULPHATE SOLUTION . MAKE UP TO 100 ML WITH THIS SOLUTION . PIPETTE 10 ML OF THE SOLUTION INTO A FURTHER 100 ML VOLUMETRIC FLASK AND MAKE UP TO VOLUME WITH THE SODIUM SULPHATE SOLUTION . USING THIS SOLUTION , PREPARE THE SOLUTIONS DILUTED AS FOLLOWS , MAKING UP TO VOLUME WITH THE 7.1 % SODIUM SULPHATE SOLUTION IN EACH CASE :

5 ML MADE UP TO 200 ML ,

5 ML MADE UP TO 100 ML ,

5 ML MADE UP TO 50 ML ,

10 ML MADE UP TO 50 ML ,

20 ML MADE UP TO 50 ML .

THESE DIFFERENT SOLUTIONS CORRESPOND TO TEST SOLUTIONS FROM WINES CONTAINING 0.0625 , 0.125 , 0.250 , 0.500 AND 1 G/L CITRIC ACID ( C8H8O7 ) .

PIPETTE 2 ML OF EACH OF THESE DILUTIONS INTO THE FLASKS MARKED A AND B , REPEAT THE ABOVE OPERATIONS .

OBTAIN THE ZERO POINT OF THE CURVE BY PIPETTING 2 ML OF THE SODIUM SULPHATE SOLUTION INTO FLASKS A AND B AND REPEAT THE ABOVE OPERATIONS .

THE GRAPH OF THE ABSORBANCES OF THESE SOLUTIONS AS A FUNCTION OF THE CITRIC ACID CONTENT IS NOT LINEAR BUT SLIGHTLY CURVED .

13 . SULPHUR DIOXIDE

13.1 . DEFINITIONS

FREE SULPHUR DIOXIDE INCLUDES MOLECULAR SO2 AND THAT COMBINED AS HSO3 , SO3 AND H2SO3 .

COMBINED SULPHUR DIOXIDE IS THE DIFFERENCE BETWEEN TOTAL AND FREE SULPHUR DIOXIDE .

13.2 . PRINCIPLE OF METHODS

( A ) REFERENCE METHOD

THE SULPHUR DIOXIDE IS CARRIED OVER BY A CURRENT OF AIR OR NITROGEN ; IT IS FIXED AND OXIDIZED BY BEING BUBBLED INTO A DILUTED AND NEUTRAL SOLUTION OF HYDROGEN PEROXIDE . THE SULPHURIC ACID FORMED IS DETERMINED BY TITRATION WITH A STANDARD SOLUTION OF SODIUM HYDROXIDE .

ENTRAINMENT AT LOW TEMPERATURE ( 10* C ) ENSURES EXTRACTION AND DETERMINATION OF THE FREE SULPHUR DIOXIDE ONLY . AT A HIGHER TEMPERATURE ( APPROXIMATELY 100* C ) THE TOTAL SULPHUR DIOXIDE IS DETERMINED .

( B ) USUAL METHOD

( AA ) FREE SULPHUR DIOXIDE : DIRECT IODOMETRIC TITRATION WITH A BLANK TITRATION ON THE SAME WINE SAMPLE IN WHICH THE FREE SULPHUR DIOXIDE HAS BEEN COMBINED WITH AN EXCESS OF ACETALDEHYDE OR PROPIONALDEHYDE .

( BB ) COMBINED SULPHUR DIOXIDE : IODOMETRIC TITRATION AFTER DOUBLE ALKALINE HYDROLYSIS OF THE WINE WHERE THE FREE SULPHUR DIOXIDE HAS BEEN OXIDIZED DURING THE IODOMETRIC TITRATION .

EXPRESSION OF RESULTS

THE QUANTITY OF SULPHUR DIOXIDE IS EXPRESSED IN MILLIGRAMS OF SULPHUR DIOXIDE PER LITRE AND DETERMINED TO THE NEAREST 10 MG/L .

13.3 . REFERENCE METHOD

FREE AND COMBINED SULPHUR DIOXIDE

APPARATUS

THE APPARATUS USED SHOULD CONFORM TO THE DIAGRAM SHOWN BELOW WITH PARTICULAR ATTENTION PAID TO THE CONDENSER .

THE GAS FEED TUBE TO THE BUBBLER " B " ENDS IN A SMALL SPHERE OF 1 CM DIAMETER WITH 20 HOLES OF 0.2 MM DIAMETER . ALTERNATIVELY THE TUBE MAY ALSO END IN A GLASS FRIT PLATE WHICH PRODUCES A LARGE NUMBER OF VERY SMALL BUBBLES THUS ENSURING GOOD CONTACT BETWEEN THE GASEOUS AND LIQUID PHASES .

ENSURE THAT THE GAS FLOW THROUGH THE APPARATUS IS APPROXIMATELY 40 LITRES PER HOUR . THE BOTTLE ON THE RIGHT OF THE ILLUSTRATION IS INTENDED TO RESTRICT THE VACUUM EFFECT OF THE WATER VACUUM PUMP TO 20 TO 30 CM WATER . INSTALL A FLOWMETER WITH SEMI-CAPILLARY TUBE BETWEEN THE BUBBLER AND THE BOTTLE TO REGULATE THE VACUUM .

SOLUTIONS

- PHOSPHORIC ACID AT 25 % ( W/V ) .

- HYDROGEN PEROXIDE ( AT 0.3 VOLUME ) .

- INDICATOR REAGENT :

- METHYL RED 100 MG ,

- METHYLENE BLUE 50 MG ,

- ALCOHOL 50 % 100 ML .

- SODIUM HYDROXIDE 0.01 N .

PROCEDURE

MAINTAIN THE WINE AT 20* C IN A FULL AND STOPPERED BOTTLE FOR FOUR DAYS BEFORE THE DETERMINATION OF THE FREE AND COMBINED SULPHUR DIOXIDE . THIS PRECAUTION IS UNNECESSARY IF ONLY TOTAL SULPHUR DIOXIDE IS TO BE MEASURED .

DETERMINATION OF FREE SULPHUR DIOXIDE

INTRODUCE INTO THE 100 ML FLASK " A " OF THE ENTRAINMENT APPARATUS , 10 ML OF WINE AND 5 ML OF 25 % PHOSPHORIC ACID . ASSEMBLE THE APPARATUS IMMEDIATELY . IF A LARGER VOLUME ( 20 OR 50 ML ) OF A WINE OF VERY LOW SULPHUR DIOXIDE CONCENTRATION IS TO BE ANALYZED , USE FLASK OF 250 ML CAPACITY FOR A .

MAINTAIN FLASK A AT A TEMPERATURE OF 10* C .

PLACE 2 TO 3 ML OF HYDROGEN PEROXIDE AT 0.3 VOLUME ( PREVIOUSLY NEUTRALIZED WITH 0.01 N SODIUM HYDROXIDE ) AND TWO DROPS OF INDICATOR REAGENT INTO THE BUBBLER B .

CONNECT THE BUBBLER TO THE APPARATUS AND BUBBLE AIR ( OR NITROGEN ) THROUGH FOR 12 TO 15 MINUTES . THE FREE SULPHUR DIOXIDE CARRIED OVER IS OXIDIZED TO SULPHURIC ACID . REMOVE THE BUBBLER FROM THE APPARATUS AND TITRATE THE ACID WHICH HAS FORMED AGAINST THE 0.01 N SODIUM HYDROXIDE SOLUTION .

LET THE NUMBER OF MILLILITRES USED BE N .

DETERMINATION OF COMBINED SULPHUR DIOXIDE

WHEN THE DETERMINATION OF THE FREE SULPHUR DIOXIDE HAS BEEN COMPLETED , AGAIN PLACE 2 TO 3 ML OF NEUTRALIZED HYDROGEN PEROXIDE IN THE BUBBLER B . HEAT THE WINE CONTAINED IN FLASK A TO BOILING POINT WITH A SMALL FLAME OF 4 TO 5 CM HEIGHT WHICH SHOULD DIRECTLY TOUCH THE BASE OF THE FLASK . DO NOT PUT THE FLASK ON A METAL PLATE , BUT ON A DISC PROVIDED WITH A HOLE OF APPROXIMATELY 300 MM DIAMETER TO PREVENT ANY OVERHEATING OF THE EXTRACTIVE MATTER OF THE WINE ON THE WALLS OF THE FLASK .

SIMMER WHILST PASSING A CURRENT OF AIR ( OR NITROGEN ) . WITHIN 12 TO 15 MINUTES THE COMBINED SULPHUR DIOXIDE HAS BEEN CARRIED OVER AND OXIDIZED . DETERMINE THE SULPHURIC ACID WHICH HAS FORMED BY TITRATION WITH 0.01 N SODIUM HYDROXIDE SOLUTION .

LET THE NUMBER OF ML USED BE N' .

DETERMINATION OF TOTAL SULPHUR DIOXIDE

OBTAIN THE TOTAL SULPHUR DIOXIDE FROM THE SUM OF THE FREE AND COMBINED SULPHUR DIOXIDE .

THE TOTAL SULPHUR DIOXIDE MAY ALSO BE DETERMINED DIRECTLY BY HEATING THE TEST SAMPLE OF 10 ML OF WINE TO BOILING POINT WHILST A CURRENT OF AIR OR NITROGEN IS PASSED .

THE FREE AND COMBINED SULPHUR DIOXIDE ARE CARRIED OVER SIMULTANEOUSLY ; THE SULPHURIC ACID FORMED CORRESPONDS TO THE TOTAL SULPHUR DIOXIDE .

THE NUMBER OF MILLILITRES OF 0.01N SODIUM HYDROXIDE USED IS DESIGNATED N'' .

CALCULATION

FREE SULPHUR DIOXIDE IN MILLIGRAMS PER LITRE : 32 N .

COMBINED SULPHUR DIOXIDE IN MILLIGRAMS PER LITRE : 32 N' .

TOTAL SULPHUR DIOXIDE IN MILLIGRAMS PER LITRE : 32 ( N + N' ) OR 32 N'' .

NOTE :

IF THE WINE CONTAINS MORE THAN 2 G/L OF ACETIC ACID ( ESPECIALLY IN VINEGAR ) DETERMINE THE ACETIC ACID THAT MAY DISTIL INTO THE BUBBLER . THIS MAY OCCUR IF THE GAS FLOW IS TOO GREAT . AFTER TITRATING THE CONTENTS OF THE BUBBLER ADD A CRYSTAL OF TARTARIC ACID TO IT , PUT IN THE VOLATILE ACIDITY DISTILLATION APPARATUS AND PASS STEAM THROUGH . DEDUCT THE VOLATILE ACIDITY OBTAINED FROM THE ACIDIMETRIC DETERMINATION OF THE SULPHUR DIOXIDE .

13.4 . USUAL METHOD

FREE AND COMBINED SULPHUR DIOXIDE

TOTAL SULPHUR DIOXIDE

SOLUTIONS

( A ) SODIUM HYDROXIDE 4N ( 160 G/L ) .

( B ) SULPHURIC ACID 1 :