Wine Analysis

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Pau Vila Cases Ph. D.

Director General
BioSystems S.A.
Index
Acetaldehyde 4 Potassium 17

Ammonia 4 Primary Amino Nitrogen (PAN) 17

Acetic Acid 5 Pyruvic Acid 18

Acetic Acid (liquid) 5 Sorbic Acid 18

Anthocyanins 6 Sucrose / D-Glucose / D-Fructose 19

Ascorbic Acid 6 Tartaric Acid 20

Calcium 7 Total Acidity 20

Catechins 7 Total Sulfite 21

Citric Acid 8 Multical / Ions Multical 22

Color 8 Control Wine (white and red) 23

Copper 9 Sulfite Control 23

CO2 9 High Glucose / Fructose Control 23

D-Gluconic Acid / D-Gluconolactone 10 Casein 24

D-Glucose / D-Fructose 11 Histamine “High Sensitivity” 24

D-Lactic Acid 12 Lysozyme 25

Glycerol 12 Ovalbumin 25

Free Sulfite 13 Ochratoxin-A 26

Iron 14 Y350 27

L-Lactic Acid 14 Y15 — Y15c 28

L-Malic Acid 15 Y25 29

pH 16 Y200 30

Polyphenols 16 Y400 31
Acetaldehyde | Ref. 12820 Ammonia | Ref. 12809
Enzymatic analysis for Enzymatic method
acetaldehyde determination for ammonia determination

Advantages Advantages
• Stable working reagent for 3 weeks • Stable liquid reagent until the expiration date
• Ready-to-use dedicated reagent • Ready-to-use dedicated reagent
• Liquid calibrator included in the kit • Liquid calibrator included in the kit

Acetaldehyde is one of the components of the oxidative chain Low nitrogen levels have been related to slow fermentation or
of alcoholic fermentation. Acetaldehyde is also formed in the sulfide production. Conversely, high levels can lead to micro-
wine aging process by ethanol oxidation. Acetaldehyde con- bial instability and production of ethyl carbonate.
centration is closely related to SO2 content. This combination
is responsible for antioxidant activity. Ammonia in the sample consumes NADH (according to the fol-
lowing reaction), which is then assayed by spectrophotometry.
Acetaldehyde in the sample yields NADH (by the following
reaction), which can be measured by spectrophotometry. GLDH
NH4+ + NADH + H+ + 2 - Oxoglutarate Glutamate + NAD+
ALDH
Acetaldehyde + NAD+ Acetic Acid + NADH + H+

Kit volume: 100 mL

Kit volume: 50 mL Method: Two-reagent differential, reading at 340 nm

Method: Two-reagent differential, reading at 340 nm Limit of linearity:: 200 mg/L

Limit of linearity:: 200 mg/L Limit of detection: 3 mg/L

Limit of detection: 0,1 mg/L

4
Acetic Acid | Ref. 12810 Acetic Acid (liquid) | Ref. 12930
Enzymatic method New enzymatic method
for acetic acid determination for acetic acid determination

Advantages Advantages
• Stable working reagent for 1 month • Stable liquid reagent until the expiration date
• Ready-to-use dedicated reagent • Ready-to-use dedicated reagent
• Liquid calibrator included in the kit • 5 liquid calibrators included in the kit

Acetic acid is produced during both alcoholic and malolactic Acetate in the sample consumes NADH (by the following
fermentations and helps enhance flavors and aromas. When reaction), which can be measure by spectrophotometry.
the wine is aerated or remains in contact with air, acetic acid
bacteria can multiply, leading to a problem known as “acetic
spoilage”. The characteristic aroma of this spoilage is due to ACS
Acetate + ATP + CoA Acetyl-CoA + AMP + Pyrophosphate
ethyl acetate. CS
Acetyl-CoA + Oxaloacetate + H2O Citrate + CoA
Acetate in the sample consumes NADH (by the following L-MDH
L-Malate + NAD+ Oxaloacetate + NADH + H+
reaction), which can be measured by spectrophotometry.

AK
Acetate + ATP Acetyl Phosphate + ADP Kit volume: 100 mL

PK Method: Two-reagent differential, reading at 340 nm

ADP+ PEP ATP+ Pyruvate
PK Linealidad: 1,3 g/L
Pyruvate + NADH Lactate + NAD+
Limit of detection: 0,02 g/L

Kit volume: 100 mL

Method: Fixed Time two-reagent, reading at 340 nm

Linealidad: 1,3 g/L

Limit of detection: 0,03 g/L

5
Anthocyanins | Ref. 12831 Ascorbic Acid | Ref. 12828
Colorimetric analysis Enzymatic method
for the assay of anthocyanins for ascorbic acid determination

Advantages Advantages
• Stable liquid reagent until the expiration date • Stable working reagent for 10 days
• Ready-to-use dedicated reagent • Ready-to-use dedicated reagent
• Calibrator included in the kit. Once reconstituted,
stable for 20 days
Anthocyanins are the tinted pigments in grapes, with the word
coming from the Greek root “antos” (flower) and “kyanos”
(blue). These pigments are found in both the skin and the
Ascorbic acid is a compound found in ripe grapes at very low
pulp.
levels compared with other acids (30-60 mg/L). It disappears
rapidly when grapes are crushed, leading to early oxidation
Anthocyanins are water-soluble pigments that provide the of must. Due to its reducing properties, ascorbic acid is used
characteristic red color of wine. At 520 nm and under cer- as an antioxidant.
tain conditions, the color is proportional to anthocyanin con-
centrations. The proposed method determines ionized and Ascorbic acid in the sample lowers MTT in the presence
ionizable anthocyanins present in the sample. Anthocyanins of PMS electron carrier, forming dehydroascorbic acid and
polymerized with tannins or other compounds cannot be MTT-formazan that can be assayed by spectrophotometry.
assayed with this method. To eliminate interferences ascorbic acid is eliminated from
the sample by oxidation to dehydroascorbic acid (ascorbate
oxidase [AO]).
PMS
Kit volume: 100 mL Ascorbic acid + Xred + MTT Dehydroascorbic Acid + Xox + MTT-formazan
AO
Ascorbic acid + ½ O2 Dehydroascorbic Acid
Method: End point with reading at 520 nm

Limit of linearity:: 1386 mg/L Kit volume: 90 mL

Limit of detection: 15 mg/L Method: Two-reagent differential, reading at 560 nm

Limit of linearity:: 150 mg/L

Limit of detection: 1,4 mg/L

6
Calcium | Ref. 12824 Catechins | Ref. 12834
Colorimetric analysis Colorimetric analysis
for calcium determination for the assay of catechins

Advantages Advantages
• Stable two-reagent liquid until the expiration date • Stable liquid reagent until the expiration date
• Ready-to-use dedicated reagent • Stable working reagent for 4 months
• Liquid calibrator included in the kit • Ready-to-use dedicated reagent
• Liquid calibrator included in the kit

Calcium is present in wine at concentrations of 6 to 165 Catechins reduce and prevent anthocyanin oxidation, keeping
mg/L. Instability due to calcium tartrate appears at 4 to 7 them from being precipitated. They are also responsible for
months of fermentation. the bitterness, astringency, yellow hue, structure and stabil-
ity of the wine. When catechins are polymerized, they form
Calcium in the sample reacts with 2,7-[bis(2-arsonophe- procyanidins that gradually form complexes with proteins,
nylazo)]-1,8-dihydroxynaphthalene-3,6-disulfonic acid (Ar- peptides and polysaccharides.
senazo III). The color increase is directly proportional to the
calcium concentration of the sample. Catechins in the sample react with the chromogen 4-(dime-
thylamino)-cinnalmaldehyde in the presence of ethanol and
an acidic medium, forming a colored complex that can be
pH=6,5 assayed by spectrophotometry.
Calcium (Ca) + Arsenazo III [Ca-(Arsenazo III)]

Catechins + DMACA [Catechins - DMACA]

Kit volume: 80 mL

Method: Two-reagent differential, reading at 635 nm Kit volume: 100 mL

Limit of linearity: 180 mg/L Method: Two-reagent differential, reading at 620 nm

Limit of detection: 2 mg/L Limit of linearity: 500 mg/L

Limit of detection: 12 mg/L

7
Citric Acid | Ref. 12825 Color | Ref. 12816
Enzymatic method Colorimetric analysis
for citric acid determination for color determination

Advantages Advantages
• Stable working reagent for 1 month • Stable liquid reagent until the expiration date
• Ready-to-use dedicated reagent • Ready-to-use dedicated reagent
• Liquid calibrator included in the kit

Citric acid is an organic acid naturally present in wine that con- Wine color plays a major role in the impression of quality.
tributes to total wine acidity. Its content is higher in white wine, Color is also an important indicator in many winemaking pro-
as the content in red wine drops during malolactic fermentation cesses. Regular use of this test allows enologists to docu-
yielding volatile acids. The permissible legal limit is 1 g/L, and its ment and confirm their own impressions.
concentration must be controlled by wine exporters.

Citrate in the sample yields oxaloacetate due to the action The wine sample is diluted in a buffer solution that does not
of the enzyme known as lyase citrate. All oxaloacetate from alter color-related properties. Absorbance reading at 420
citrate in the sample is converted into L-malic acid by the nm, 520 nm and 620 nm allows the chromatic characteris-
enzyme L-malate dehydrogenase. The disappearance of tics to be calculated.
NADH may be read by spectrophotometry.

CL
Citrate Oxaloacetate + Acetate
PK Kit volume: 80 mL
Oxaloacetate + NADH + H+ L-malate + NAD+
Method: End point monoreagent reading at
Kit volume: 50 mL 420, 520 y 620

Method: Two-reagent differential, reading at 340 nm Limit of linearity: 18 (A420, A520 y A620)

Limit of linearity:: 400 mg/L Limit of detection: 0,113 (A420), 0,144 (A520) y 0,121 (A620)

Limit of detection: 11 mg/L

8
Copper | Ref. 12814 CO2 | Ref. 12832
Colorimetric analysis Enzymatic method for CO2 determination
for copper determination

Advantages Advantages
• Stable liquid reagent until the expiration date • Stable liquid reagent until the expiration date
• Ready-to-use dedicated reagent • Ready-to-use dedicated reagent
• Liquid calibrator included in the kit • Liquid calibrator included in the kit

Copper is a metal that clearly originates in the process of Carbon dioxide is a natural gas produced during fermenta-
vinegrowing. The main source is phytosanitary treatments tion that is dissolved in wines. The addition of CO2 during
of vineyards to prevent mildew. During harvest, the copper preparation directly affects the aroma and taste of wine and
content may be 4 to 6 mg/L. During fermentation its concen- can enhance freshness and acidity in the mouth, softening
tration decreases to 0.2-0.3 mg/L due to the formation of the sweetness. However, it can also intensify bitterness and
copper sulfides or the presence of yeasts that fix the copper astringency.
contained in the medium.The OIV has set a maximum accept-
able limit of copper of 1 mg/L. According to the coupled reactions described below, carbon
dioxide (CO2) in the sample consumes NADH analogue co-
Any copper in the sample reacts with 4-(3,5-dibromo-2- pyr- factors that can be assayed by spectrophotometry at 405 nm.
idylazo)-N-ethyl-N-sulfopropylaniline (PAESA). The color in-
crease is directly proportional to the copper concentration PEPC
of the sample. Phosphoenolpyruvate +HCO3 Oxaloacetate + H2PO4-

pH=4,7 MDH
Copper (Cu) + 2PAESA [Cu(PAESA)2] Oxaloacetate + Analogue + NADH Malate + NAD+ Analogue
reducing agent

Kit volume: 100 mL

Kit volume: 50 mL
Method: Two-reagent differential, reading at 560 nm
Method: Single-reagent fixed time, reading at 405 nm
Limit of linearity: 7 mg/L
Limit of linearity: 1500 mg/L
Limit of detection: 0,4 mg/L
Limit of detection: 55 mg/L

9
D-Gluconic Acid /
D-Gluconolactone | Ref. 12811
Enzymatic method for D-gluconic acid /
D-gluconolactone determination
D-gluconic acid is an indicator of grape deterioration and san-
Advantages itary condition
D-gluconic acid in the sample yields NADPH (by the fol-
• Stable liquid reagent until the expiration date lowing reaction), which can be measured by spectropho-
• Ready-to-use dedicated reagent tometry.
• Liquid calibrator included in the kit
GK
D-Gluconate + ATP D-Gluconate-6-P + ADP

6-PGDH
Kit volume: 100 mL D-Gluconate-6-P + NADP+ Ribulose-5-P + NADPH + CO2 + H+

Method: Two-reagent differential, reading at 340 nm

D-gluconolactone can be determined according to the same
Limit of linearity: 2 g/L principle after alkaline hydrolysis.
pH11
Limit of detection: 0,003 g/L D-Gluconolactone + H2O D-Gluconate

10
 D-Glucose / D-Fructose | Ref. 12800
Enzymatic method for D-glucose
/ D-fructose determination
This test indicates the best moment for grape harvesting and
Advantages allows alcoholic fermentation to be monitored. It is widely
used to determine the dryness of the wine before bottling.
• Stable liquid reagent until the expiration date
D-fructose and D-glucose in the sample generate NADH (by
• Working reagent stable until the expiration date the following reaction), which can be measured by spectro-
• Ready-to-use dedicated reagent photometry. The configuration of these reagents allows D-glu-
• Liquid calibrator included in the kit cose/D-fructose (total sugars) to be determined if the enzyme
PGI is added or D-glucose to be determined if it is not.
HK
D-Fructose + ATP Fructose-6-Phosphate + ADP
HK
D-Glucose + ATP Glucose-6-Phosphate + ADP
PGI
Fructose-6-Phosphate Glucose-6-Phosphate
G6P-DH
Glucose-6-Phosphate + NADP+ Gluconate-6-Phosphate +NADPH + H+

Kit volume: 120 mL

Method: Two-reagent differential, reading at 340 nm

Limit of linearity: 8 g/L

Limit of detection: D-Glucose: 0,03 g/L

D-Glucose / D-Fructose: 0,02 g/L

11
D-Lactic Acid | Ref. 12801 Glycerol | Ref. 12812
Enzymatic method Colorimetric analysis
for D-lactic acid determination for glycerol determination

Advantages Advantages
• Stable liquid reagent until the expiration date • Stable one-reagent liquid until expiration date
• Ready-to-use dedicated reagent • Ready-to-use dedicated reagent
• Liquid calibrator included in the kit • Liquid calibrator included in the kit

The excess of bacteria that are producing D-Lactic acid can Glycerol is an indicator of the quality of finished wine and is
inhibit alcoholic fermentation, converting some sugars into extremely important for the mouthfeel. High glycerol concen-
D-lactic acid. This is one of the main problems in the wine- trations add sweetness, body and fullness to the wine.
making process. Levels above 0.3 g/L of D-lactic acid indi-
cate bacterial contamination. Glycerol in the sample yields (by the following reaction),
a colored complex that is assayed by spectrophotometry.
D-lactic acid in the sample yields NADH (by the following
reaction), which can be measured by spectrophotometry.
glycerol kinase
Glycerol + ATP Glycerol-3-P + ADP
D-LDH G-3P-oxidase
D-Lactate + NAD+ Piruvate + NADH Glycerol-3-P + O2 Dihydroxyacetone-P + H2O

peroxidase
2 H2O2 + 4-Minoantipyrine Quinone-Imine + 4 H2O

Kit volume: 100 mL

Method: Two-reagent differential, reading at 340 nm

Kit volume: 100 mL
Limit of linearity: 0,25 mg/L
Method: Two-reagent differential, reading at 500 ± 20 nm
Limit of detection: 0,004 g/L
Limit of linearity: 20 g/L

Limit of detection: 0,24 g/L

12
Free Sulfite | Ref. 12813
Colorimetric analysis
for free sulfite determination
Most sulfur dioxide added to the must or wine combines with
Advantages different organic compounds. This is the predominant frac-
tion in wine; however, there is another fraction that is not
combined, namely, free SO2. Although it is present in lower
• Stable liquid reagent until the expiration date
amounts, its antiseptic and antioxidant properties are superior
• Ready-to-use dedicated reagent to those of combined sulfite.
• Liquid calibrator included in the kit
Free sulfite in the sample reacts with 4,4’-(4-iminocyclo-
hexa-2,5-dienylidenemethylene) dianiline chromogen
(pararosaniline; PR) and formaldehyde (F) in acid medium.
Formaldehyde
SO2 + PR + I [PR-F-SO2] + [PR-F-I] In a second reaction, free sulfite is removed by oxidation and
the rest of substances (I) that are able to react with the chro-
Formaldehyde
SO2 + PR + I [PR-F-I] mogen are measured. The difference between the results ob-
+Oxidizer-SO2 tained from the two reactions is the free sulfite concentration.

Kit volume: 250 mL

Method: End point Two-reagent, reading at 560 nm

Limit of linearity: 100 mg/L

Limit of detection: 0,7 mg/L

13
Iron | Ref. 12817 L-Lactic Acid | Ref. 12802
Colorimetric analysis Enzymatic method
for iron determination for L-lactic acid determination

Advantages Advantages
• Stable liquid reagent until the expiration date • Stable liquid reagent until the expiration date
• Ready-to-use dedicated reagent • Ready-to-use dedicated reagent
• Liquid calibrator included in the kit • Liquid calibrator included in the kit

Metal components in wine can originate in grapes or the ma- L-lactic acid is the product of the metabolism of malic acid
chinery used to make wine. A high iron content can cause during the malolactic fermentation. L-lactic acid is perceived
clouding due to a lack of solubilization, thus affecting the color as less acidic and softer on the palate compared to malic acid.
and clarity of the wines.
L-lactic acid in the sample yields NADH (by the following
Any iron in the sample reacts with 3-(2-pyridyl)-5,6-bis reaction), which can be measured by spectrophotometry.
(4-phenylsulfonic)-1,2,4-triazine (ferrozine) sodium salt in
acidic medium and in the presence of a reducing agent. The
L-LDH
color increase is directly proportional to the iron concentra- L-Lactato + NAD+ Piruvato + NADH
tion of the sample.

Kit volume: 100 mL

pH=4,1
Iron (Fe) + Ferrozine [Fer-Ferrozine]
reducing agent Method: Two-reagent differential, reading at 340 nm

Limit of linearity: 3 g/L

Kit volume: 100 mL Limit of detection: 0,02 mg/L

Method: Two-reagent differential, reading at 560 nm

Limit of linearity: 30 mg/L

Limit of detection: 0,1 mg/L

14
L-Malic Acid | Ref. 12803
Enzymatic method
for L-malic acid determination
L-malic acid is responsible for the sharply acidic, green apple
Advantages flavor in wine. It’s fermentation yields L-lactic acid and causes
perceived acidity to soften.
• Stable liquid reagent until the expiration date
• Stable working reagent for 4 months L-malic acid in the sample yields NADH (by the following
• Ready-to-use dedicated reagent reaction), which can be measured by spectrophotometry.
The equilibrium of this reaction moves toward L-malic acid
• Liquid calibrator included in the kit
formation. The enzyme glutamate-oxaloacetate transami-
nase (GOT) causes the equilibrium to shift by eliminating
oxaloacetate, which is converted into L-aspartate in the
L-MDH presence of L-glutamate.
L-Malate + NAD+ Oxaloacetate + NADH
GOT
Oxaloacetate + NADH + H+ Aspartate + 2-Oxoglutarate

Kit volume: 100 mL

Method: Two-reagent differential, reading at 340 nm

Limit of linearity:: 4 g/L

Limit of detection: 0,016 g/L

15
pH | Ref. 12876 Polyphenols | Ref. 12815
Colorimetric method for Colorimetric analysis
the determination of pH for polyphenols determination

Advantages Advantages
• Liquid reagent stable until expiry date • Stable liquid reagent until the expiration date
• Dedicated reagents ready for use • Ready-to-use dedicated reagent
• Liquid calibration standards included in the kit • Liquid calibrator included in the kit

In musts and wines the pH varies depending on the ripe- Phenol components significantly enhance the antioxidant
ness of the grapes, the concentration of organic acids at the properties, color and mouthfeel of red wines. The importance
time of harvest, the variety of the grape, the presence and of these phenol components in sensory perception requires
metabolism of micro-organisms and the fermentation tem- assay at all stages of the winemaking process.
perature etc. The appearance of tartrate precipitates during
the wine-making process will alter the final pH of the wine. Any polyphenols in the sample react with Folin-Ciocalteu’s
reagent in basic medium. The color increase is directly pro-
The hydrogen ions in the sample alter the pH of the sample/ portional to the polyphenols concentration of the sample.
buffer mix and can be measured spectrophotometrically with
the bromophenol blue indicator. pH=10,9
Polyphenols + Folin-Ciocalteu’s Reagent (RF) [Polyphenols - FC]

Kit volume: 100 mL Kit volume: 80 mL

Method: Two-reagent differential, reading at 600 nm Method: Two-reagent endpoint, reading at 670 or 750 nm

Measurement range: 3,00 a 4,40 Limit of linearity: 3000 mg/L

Limit of detection: 60 mg/L

16
Potassium | Ref. 12823 Primary Amino Nitrogen
Enzymatic method (PAN) | Ref. 12807
for potassium determination
Colorimetric analysis
for primary amino nitrogen determination
Advantages
• Stable liquid reagent until the expiration date
Advantages
• Ready-to-use dedicated reagent
• Stable liquid reagent until the expiration date
• 5 liquid calibrators included in the kit
• Stable working reagent for 12 months
• Ready-to-use dedicated reagent
The amount of potassium in grape must varies between 600 • Liquid calibrator included in the kit
and more than 2500 mg/L in certain varieties of red wine.
During véraison, soil potassium moves toward the fruit where
it forms soluble potassium bitartrate. Alcohol and low tem- Nitrogen compounds (molecules containing a primary amine
peratures can reduce its solubility, leading to precipitation. nitrogen) in must and wine play a key role in fermentation and
the potential of microbial stability.
Potassium in the sample consumes NADH (by the following
reaction), which can be measured by spectrophotometry. Any molecules in the sample that contain a primary amino
nitrogen react with o-phthaldialdehyde (OPA) in the pres-
ence of a reducing agent in basic medium, yielding a chro-
Phosphoenolpyruvate + ADP
K+
Piruvate + ATP mogen that is assayed spectrophotometrically.
PK
LDH
Piruvate + NADH + H+ Lactate + NAD+ pH=9,5
OPA + NH2R [OPA-NH2R]
reducing agent

Kit volume: 80 mL Kit volume: 100 mL

Method: Two-reagent differential, reading at 340 nm Method: Two-reagent differential, reading at 340 nm

Limit of linearity: 1500 mg/L wine | 4000 mg/L Most Linealidad: 400 mg/L

Limit of detection: 8 mg/L Limit of detection: 1 mg/L

17
Pyruvic Acid | Ref. 12826 Sorbic Acid | Ref. 12880
Enzymatic method Colorimetric method
for pyruvic acid determination for sorbic acid determination

Advantages Advantages
• Stable liquid reagent until the expiration date • Stable liquid reagent until the expiration date
• Stable working reagent for 2 months • Liquid calibrator included in the kit
• Ready-to-use dedicated reagent
• Liquid calibrator included in the kit
Sorbic Acid is a natural organic compound used as a food
preservative in its mineral salt form (Potassium sorbat). The
Pyruvic acid is an organic acid naturally present in wine and maximum limit permitted by OIV in wine for sorbic acid is
one of the acids that most influences its body and mouthfeel. 200 mg/l. Sorbic acid inhibits with SO2 yeast populations in
Pyruvic acid is a result of the fermentation process and con- pre-bottled wine and it is used to avoid undesired fermenta-
tributes to the organoleptic properties of wine, but must be tions in wine bottles, especially in off-dry and sweet wines.
controlled because selective sulfite-binding shortens the life
of the wine. Sorbic acid is oxidized to obtain malondialdehyde (MD)
that reacts with thiobarbituric acid (TBA) generating a com-
Pyruvate in the sample yields oxalacetate due to the ac- pound that can be measured spectrophotometrically.
tion of the enzyme known as D-lactate dehydrogenase.
This reaction consumes NADH that is oxidized to NAD+ Oxidizer
and the disappearance of the latter can be measured Sorbic Acid 2 Malondialdehyde
by spectrophotometry. MD + 2TBA [TBA-MD-TBA]

D-LDH
Piruvate + NADH Oxaloacetate + NAD+
Kit volume: 50 mL

Kit volume: 100 mL Method: Kynetic monoreagent, reading at 520 nm

Method: Two-reagent differential, reading at 340 nm Limit of linearity:: 300 mg/L

Limit of linearity:: 400 mg/L Limit of detection: 2,19 mg/L

Limit of detection: 6 mg/L

18
 Sucrose / D-Glucose /
D-Fructose | Ref. 12819
Enzymatic method for sucrose or Sucrose
/ D-Glucose / D-Fructose determination
Precise analysis of sucrose or total sugar is important for
Advantages many winecellars in two winemaking operations. Sparkling
wine (cava, champagne, etc.) production: adding sucrose
• Stable liquid reagent until the expiration date once alcoholic fermentation has been carried out in order to
achieve a secondary fermentation that produces CO2, which
• Stable working reagent for 3 months
is retained in the wine.
• Ready-to-use dedicated reagent
• Liquid calibrator included in the kit Sucrose, D-fructose and D-glucose in the sample generate
NADPH (by the following reaction), which can be measured
by spectrophotometry.
β-Fructosidasa
Sacarose + H2O D-glucose + D-fructose
HK
D-fructose + ATP Fructose-6-phosphate + ADP
HK
D-glucose + ATP Glucosa-6-phosphate + ADP
PGI
Fructose-6-phosphate Glucosa-6-phosphate
G6P-DK
Glucosa-6-phosphate + NADP+ Gluconate-6-phosphate + NADPH + H+

Kit volume: 60 mL

Method: End-point/two-reagent differential, reading at 340 nm

Limit of linearity: Sucrose 4 g/L, Suc./D-Gluc./D-Fruc. 8 g/L

Limit of detection: Sucrose 0,08 g/L, Suc./D-Gluc./D-Fruc. 0,07 g/L

19
Tartaric Acid | Ref. 12808 Total Acidity | Ref. 12846
Colorimetric analysis Colorimetric analysis
for tartaric acid determination for the assay of total acidity

Advantages Advantages
• Stable liquid reagent until the expiration date • Stable liquid reagent until the expiration date
• Ready-to-use dedicated reagent • Ready-to-use dedicated reagent
• Liquid calibrator included in the kit • Liquid calibrator included in the kit

Total acidity should be determined in must to ensure good

Tartaric acid is the main acid of wine that can become in- fermentation, as well as in wine after fermentation because it
soluble, forming various salts. This acid produces the fruity is a key factor for the storage and stability of wine over time.
aromas and freshness of wines and is the most commonly Low acidity means that microbial alterations and wine with
used acidifier. defects and of poorer quality is more likely. Low acidity can
cause microbial instability that results in wine defects and ove-
Any tartaric acid in the sample reacts with vanadium salt in rall decrease in quality. Wine should have an adequate total
acidic medium, forming a colored complex that is assayed acidity value consistent with the other components to achieve
by spectrophotometry. good balance. This value can be between 3 and 7 g/L.

Total acidity is the sum of assayable acids in wine or must,

pH=2,5 such as malic acid, tartaric acid, lactic acid, etc., except for
Tartaric Acid (TART) + Vanadium Salt (V) [TART-V]
carbonic acid and sulfurous acid. Th is reagent determines
the total acidity, expressed as g/L of tartaric acid.

Kit volume: 100 mL Acids in the sample modify the pH in the reaction mixture
that, in the presence of the bromothymol blue (BTB) indica-
Method: Two-reagent differential, reading at 520 nm
tor, can be measured spectrophotometrically.
Measurement range: 0,06 a 6 g/L
Kit volume: 100 mL
Limit of detection: 0,06 g/L
Method: Two-reagent differential, reading at 620 nm

Linealidad: 12 g/L

20
Total Sulfite | Ref. 12806
Colorimetric analysis for
total sulfite determination
Sulfite is the main preservative of wines and musts, due to
Advantages its antiseptic properties on yeasts and bacteria; it also has
antioxidant properties. According to Council Regulation (EC)
No 1493/1999 and Council Regulation (EC) Nº 1622/2000,
• Stable liquid reagent until the expiration date
the sulfur dioxide content of wine is limited, as it is considered
• Ready-to-use dedicated reagent to be a slightly toxic substance from the point of view of its
• Liquid calibrator included in the kit effects on human physiology.

Total sulfites in the sample react with 5-5’-dithio-2-nitroben-

zoic (DTNB) acid in basic medium. Cleavage of the disulfide
pH=1,0
SO2 + R-S-S-R (DTNB) R-S-SO2 + S-R bond (R-S-S-R) of DTNB by a sulfite molecule yields the
5-mercaptan- 2-nitrobenzoate molecule, which absorbs at
405 nm. The color increase of the sample is directly propor-
Kit volume: 200 mL tional to the total sulfite concentration of the sample.
Method: Two-reagent differential, reading at 405 nm

Limit of linearity: 400 mg/L

Limit of detection: 1 mg/L

21
Multical | Ref. 12818 Ions Multical | Ref. 12841
Multiparameter calibrator Multiparameter calibrator

MULTICAL is a multiparameter calibrator with five synthetic IONS MULTICAL. 5 levels with 10 mL. Multiparameter calibra-
matrix liquid levels (5 x 10 mL). It contains various analaytes tor with five synthetic matrix liquid levels that contain various
at adequate concentrations for the calibration of the measu- metals at adequate concentrations to calibrate the measure-
rement procedures. ment procedures.

The traceability of the results in samples to reference ma- The concentration values assigned to each component and
terials or systems of higher metrological hierarchy is only their traceability is ensured by using the reagents and mea-
ensured when the reagents and measurement procedures surement procedures recommended by BioSystems.
recommended by BioSystems are used.

Parameter U 1 2 3 4 5 Parameter U 1 2 3 4 5

Acetic acid g/L 0,15 0,30 0,60 0,90 1,20 Calcium mg/L 20,3 40,5 81,0 121,5 162,0

Ammonia mg/L 23 45 90 135 180 Copper mg/L 0,8 1,6 3,2 4,7 6,3

Citric acid mg/L 45 90 180 270 360 Iron mg/L 3,4 6,8 13,5 20,3 27,0

D-Gluconic Acid g/L 0,20 0,40 0,80 1,20 1,60 Potassium mg/L 188 375 750 1125 1500

D-Glucose g/L 0,90 1,80 3,60 5,40 7,20 Magnesium mg/L 4,5 9,0 18,0 27,0 36,0

D-Glucose/D-fructose g/L 0,90 1,80 3,60 5,40 7,20 Traceability: aqueous reference standard

Glicerol g/L 0,113 0,225 0,450 0,675 0,900

Ácido D-láctico mg/L 0,028 0,056 0,113 0,169 0,225
Ácido L-láctico g/L 0,34 0,68 1,35 2,03 2,70
Ácido L-málico g/L 0,45 0,90 1,80 2,70 3,60
PAN mg/L 45 90 180 270 360
Suc./D-gluc./D-fruc. g/L 0,90 1,80 3,60 5,40 7,20

Traceability: aqueous reference standard

22
Control Wine Sulfite Control | Ref. 12827
(white and red) | Ref. 12821 / 12822 Sulfite (I and II) Control is a synthetic liquid material that con-
tains stabilized sulfite at adequate concentrations for quality
Multiparameter calibrator control in laboratories. It does not contain preservatives that
could interfere with the measurements.

Control Wine (white and red) is a wine (10 x 5 mL) that con- The concentration values assigned to each level are shown
tains various components at adequate concentrations for in the attached tables. The values are traceable to the unit
quality control in laboratories. The product is designed for of mass. Traceability is ensured only by using the measure-
intra-laboratory quality control and is supplied with acceptable ment reagents and procedures recommended by BioSyste-
value intervals. ms. The acceptable ranges suggested have been prepared
based on prior experience in interlaboratory variability and
Traceability is only ensured when the reagents and measure- are provided only as a guideline, as each laboratory should
ment procedures recommended by BioSystems are used. establish its own precision parameters.

Component U Component Level Value Limits Units

Acetic acid
g/L Sulfite I 40 36-44 mg/L

Ammonia
mg/L (free & total) II 80 72-88 mg/L

Iron
mg/L
D-Gluconic acid g/L
D-glucose / D-fructose g/L
High Glucose / Fructose
D-glucose
Glycerol
g/L
g/L
Control | Ref. 18069
L-Lactic acid g/L BioSystems offers a 200 g/L aqueous standard in order to
L-Malic acid
g/L facilitate work with D-Glucose/D-Fructose techniques that
include a pre-dilution.
Primary Amine Nitrogen mg/L
Polyphenols
mg/L
Tartaric acid
g/L
Citric acid mg/L
Histamina

23
Casein | Ref. 14113 Histamine “High Sensitivity” | Ref. FCE3100
ELISA method ELISA method

Advantages Advantages
• Fast, standard method • High sensitivity
• High sensitivity • Liquid reagent, stable until the expiration date
• Liquid reagent, stable until the expiration date • Easy sample preparation
• Easy sample preparation

Casein is an allergenic protein present in cow’s milk and dairy Histamine is a biogenic amine present in certain food with
products made from cow’s milk. The presence of traces of high concentrations of protein or foods exposed to fermenta-
these proteins must be labeled due to the risk it poses to tion processes. Histamine is created by certain microorgan-
the health of people with allergies, as set forth in the legisla- isms that affect the amino acid histidine. Histamine intake by
tion. In addition to foods that naturally contain casein, there sensitive individuals produces undesireable effects, such as
may be traces of these proteins in processed foods due to headaches or skin reactions; hence, it should be controlled.
cross-contamination or the use of additives. Caseins are used
as clarifier or fining agent in the winemaking process.
High-sensitivity ELISA of histamine is a competitive en-
Casein reagent is a sandwich enzyme-linked immunosor- zyme-linked immunoabsorbent assays for the quantitative
bent assay (ELISA) for the quantitative analysis of casein analysis of histamine in wine, fish, cheese and meat.
traces in samples of wine, juice, cookies, meat products,
chocolate and other food products. Histamine in the sample is quantitatively derivatized to N-acyl-
histamine by using an acylating reagent. The microplate wells
are coated with histamine. In a first incubation, acylated hista-
mine in the sample or reference standard competes with fixed
histamine to bind to anti-histamine antibodies.
Presentation: 96 wells

Method: Sandwich ELISA, reading at 450 nm Presentation: 96 wells

Limit of detection: 0,04 ppm Method: Competitive ELISA, reading at 450 nm

STD Concentration: 0 - 0,2 - 0,6 - 2 - 6 ppm Limit of detection: 0,15 ppb

STD Concentration: 0 - 25 - 100 - 250 - 500 ppb

24
Lysozyme | Ref. 14122 Ovalbumin | Ref. 14125
ELISA method ELISA method

Advantages Advantages
• Fast, standard method • Fast, standard method
• High sensitivity • High sensitivity
• Liquid reagent, stable until the expiration date • Liquid reagent, stable until the expiration date
• Easy sample preparation • Easy sample preparation

Lysozyme is an allergenic protein contained in eggs and egg Ovalbumin is an allergenic protein contained in eggs and
products. As set forth by law, the presence of traces of this egg products. As set forth by law, the presence of traces of
protein should be labeled due to the risk posed to the health of this protein should be labeled due to the risk posed to the
allergic individuals. In addition to foods that naturally contain ly- health of allergic individuals. In addition to foods that natu-
sozyme, there may be traces of this protein in processed foods rally contain ovalbumin, there may be traces of this protein
due to cross-contamination or the use of additives. Lysozyme in processed foods due to cross-contamination or the use of
is used as a preservative in the winemaking process. additives. Ovalbumin is used as a clarifier finding agent in the
winemaking process.
Lysozyme reagent is a sandwich enzyme-linked immuno-
sorbent assay (ELISA) for the quantitative analysis of casein Ovalbumin reagent is a sandwich enzyme-linked immuno-
traces in wine and cheese samples. sorbent assay (ELISA) for the quantitative analysis of casein
traces in wine and food samples.

Presentation: 96 wells Presentation: 96 wells

Method: Sandwich ELISA Method: Sandwich ELISA

Limit of detection: 2 ppb Limit of detection: 4 ppb

STD Concentration: 0 - 25 - 100 - 250 - 500 ppb STD Concentration: 0 - 25 - 100 - 250 - 500 ppb

25
Ochratoxin-A ELISA | Ref. 14108 Rapid Test | Ref. 14203

ELISA Method / Rapid Test

Ochratoxin-A is a nephrotoxic and hepatocarcinogenic mi-
crotoxin produced by Penicillium verrucosum and P. viridica-
tum as well as by several species of Aspergillus, such as A.
ochraceus in warm tropical areas of the world. Ochratoxin-A
has been found in several cereals and other plant products,
in coffee, wine and animal feed.
The legally-established maximum levels in Europe for ochra-
toxin-A depend on whether the food is being used directly
for human consumption or as a raw material for prepared
products, varying between 2 and 20 μg/kg (ppb).

Advantages of ELISA Advantages of Rapid Test

• High sensitivity • Results in 10 minutes
• Validated for wine • Easy to use
• Includes everything necessary for on-site analysis
The ELISA Ochratoxin-A kit is a competitive enzyme im- • Low cost
munoassay for quantitative analysis of the Ochratoxin-A in • High sensitivity
foods and feeds (corn, rice, wheat, sorghum, barley, oat, • Detection limits in line with current legislation
rye, coffee, cacao, pulses and wine).

The Ochratoxin-A Rapid Test is a competitive enzyme im-

munoassay on nitrocellulose for screening Ochratoxin-A in
Presentation: 96 wells wines (white, rosé and red)..
Method: Competitive ELISA, reading at 450 nm Presentation: 10 test

Limit of detection: 0,3 ppb Method: Rapid Test (Ezyme immunoassay)

STD Concentration: 0 - 0,0625 - 0,125 - 0,25 - 0,5 - 1 ppb Limit of detection: 0,3 ppb

26
Y15 / Y25 / Y350 are open analyzers.
In conjunction with the reagent line, the BioSystems Analyzers make it possible to monitor the entire vinification process.
The system adjusts to the various sample types that the enologist needs to analyze.

Technical Specifications | Code 80176

Optical System Calibration
Range of measurment: 0-3.5 A Factor
all wavelengths Calibrador
Wavelengths: 280, 340, 405, 420, 505, 520, Curva de calibración
560, 620, 635, 670, 750 (nm)
Light Source: LEDs Calibration Curve
Settings: monochromatic and bichromatic Up to 8 Calibration points
Up to 3 replicates per point

Thermostat System Quality Control

Peltier system from 25-40 °C 2 controls per test
Levey-Jennings control chart
Fluidic System Westgard´s Rules
Continuous flow system with peristaltic
pump incorporated Installation Characteristics
Stepper motor pump operation Voltage: 100V-240 V
Sipping volume can be programmed Frequency: 50/60 Hz
from 100 μL to 5 mL Maximum power: 30 W
Automatic adjustment of sample volumen Temperature: 10-35 °C
Automatic adjustment of sample position Max Rel humidity: 75 %
Height: <2000 m
Printer Screen and Keyboard Dimensions: 420 x 350 x 216 mm
Thermic printer Weight: 4 kg
Screen: graphic LCD lighted screen 320 x
240 px Accessories
Keyboard: tactile membrane Battery Pack
- Capacity 2000 mAh
Methods of Calculation - Duration: 2 hrs
Absorbance 0,2, 1 and 10 mm flow quartz cuvette
End Point 10 mm flow glass cuvette
Differential Mode 1 mm glass cuvette + adapter
Fixed Time 10 mm quartz cuvette
Kinetic

27
 Technical Specifications ||| Cód.
Code 83106
Code83106 /
83106c

Random Access automatic analyser.

Photometric reading directly on the reaction rotor.

Test rate 150 tests/hour (75 results/hour)

Number of rack positions — Y15 4 (samples and/or reagents)

Number of rack positions — Y15c 2 (samples and/or reagents)

Number of samples per rack 24 (multiuse racks)

Number of samples per rack 10 (20 and 50 mL bottles)

Number of cooled reagents — Y15c 10 (20 mL bottles) and 10 (50 mL bottles)

Maximum number of samples/reagents — Y15 72 samples / 30 reagents

Maximum number of samples/reagents — Y15c 48 samples / 30 reagents

Sample tubes ø13 mm, ø15 mm (maximum height 100 mm)

Standard vial ø13 mm

Programmable reagent volume — A / B 10 μL - 600 μL / 10 μL - 200 μL

Programmable sample volume 2 μL - 80 μL

Removable methacrylate rotor

Number of wells in the rotor 120

Automatic pre- and post-dilutions

Permissible reaction volumes 180 μL - 800 μL

Measurement range from -0,05 A to 3,6 A

Filter drum configuration 340, 405, 420, 520, 560,

600, 620, 635, 670 (nm)

Dimensions 840 x 670 x 615 (mm) (length x deep x height)

Weight 45 kg

28
 Technical Specifications | Code 83107
Random Access automatic analyser.
Photometric reading directly on the reaction rotor.

Test rate 240 tests/hour (120 results/hour)

Number of cooled reagents 30

Positions for uncooled rack 3 (multipurpose rack)

Number of samples per rack 24

Maximum number of samples 10 (20 mL and 50 mL bottles)

Sample tubes ø13 mm, ø15 mm (maximum height 100 mm)

ø13 mm

Number of reagents per rack 10

Max. number of uncooled reagents 20

Reagent bottles 20 mL y 50 mL

Programmable reagent volume — A / B 10 μL - 440 μL / 10 μL - 200 μL

Programmable sample volume 2 μL - 40 μL

Removable methacrylate rotor

Number of wells 120

Automatic pre- and post-dilutions

Reaction volume range 180 μL - 680 μL

Measurement range from -0,05 A to 3,6 A

Filter drum configuration 340, 405, 420, 520, 560,

600, 620, 635, 670 (nm)

Dimensions 1080 x 695 x 510 (mm) (length x deep x height)

Weight 73 kg

29
 Technical Specifications | Code 83020
Speed
200 tests/hour (200 results/hour)

Capacity
88 positions:
· 44 sample or 60 mL positions
(tube or pediatric well)
· 44 sample or 20 mL positions
(tube or pediatric well)

Sistema Fluídico
RA volume from 90 μL to 300 μL
RB volume from 10 μL to 100 μL
Sample volume from 2 μL to 40 μL
Reaction volume from 180 μL to 440 μL
Level and clot detector

Optical System
LED + Hard Coating Filter
Main photodiode + reference photodiode
Wavelenghts
340, 405, 420, 430, 505, 520, 560, 600, 620
635, 750 (nm)

Other Characteristics
Dimensions 1077 x 690 x 680 (mm)
166 Kg

30
Technical Specifications | Code 83040
Speed
400 tests/hour (400 results/hour)

Capacity
135 sample positions
(90 with automatic barcode reading)

88 reagent bottles (refrigerated)

Removable blade with 120 reaction cuvettes
(autowashable)

Fluid System
RA volume from 90 μL to 450 μL
RA volume from 10 μL to 300 μL
Sample volume from 2 μL to 40 μL
Reaction volume from 180 μL to 600 μL
Level and clot detector

Optical System
LED + Hard Coating Filter
Main photodiode + reference photodiode
Wavelenghts
340, 405, 420, 430, 505, 520, 560, 600, 620
635, 750 (nm)

Other Characteristics
Dimensions 1200 x 720 x 1258 (mm)
210 Kg

31
99826

Manufactured by: BioSystems S.A.

BioS/06-21

Costa Brava 30, 08030 Barcelona (Spain) | Tel. (+34) 93 311 00 00

enology@biosystems.es | www.biosystems.es