A project report on alcohol 2

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A PROJECT REPORT ON

ANALYSIS OF ALCOHOLIC BEVERAGES BY FLAME ATOMIC ABSORPTION SPECTROPHOTOMETER

(FAAS)

SUBMITTED TO DEPARTMENT OF CHEMISTRY, ST. JOHN’S COLLEGE, AGRA

FOR THE DEGREE OF MASTER OF SCIENCE (M Sc) IN PHYSICAL CHEMISTRY (2013-2014)

UNDER THE SUPERVISION OF:

Dr. SUSAN VERGHESE .P Associate Professor

Department of Chemistry St. John’s College, Agra

SUBMITTED BY: SAURAV K. RAWAT M Sc Final Physical Chemistry 2013-14

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CERTIFICATE

This is to certify that this project entitled “ANALYSIS OF ALCOHOLIC BEVERAGES BY FLAME ATOMIC ABSORPTION SPECTROPHOTOMETER (FAAS)” submitted to St. John’s College, Agra, for the fulfillment of the requirement for the Master degree is a bona fide project work carried out by SAURAV K. RAWAT student of M Sc Final (PHYSICAL CHEMISTRY) under my supervision and guidance during the session 2013-2014. The assistance and help rendered during the course of investigation and sources of literature have been acknowledged.

Dr. Susan Verghese .P Associate Professor Department of Chemistry St. John’s College, Agra

(Supervisor)

Dr. Hemant Kulshreshtha HEAD

Department of Chemistry St. John’s College, Agra

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ACKNOWLEDGEMENT

It is my proud privilege to express my profound sense of gratitude and sincere indebtedness to honorable Dr Alexander Lal, Principal of St. John’s College, Agra, for providing infrastructure for the completion of this project. I am thankful to Dr Hemant Kulshreshtha, Head of the Chemistry Department; he was always affectionate, pain taking and source of inspiration to me. I am highly obliged to him for their guidance, constructive criticism and valuable advice which they provided to me throughout the tenure of my project. The project work could not have been possible without his worthy suggestions and constant co-operation. I am also thankful to my supervisor Dr Susan Verghese to guide me on the various sides of this project and her help and guidance she provided to me for the initiation of this project. My heart is filled with deep sense of thankfulness and obeisance to my teachers Dr. R P Singh, Dr. H B Singh, Dr. P E Joseph, Dr. Raju V John, Dr. Shalini Nelson, Dr. Mohd. Anis, Dr. Anita Anand, Dr. Padma Hazra, and Dr. David Massey for their valuable suggestions and lively moral boosting during the progress of this investigation. I am also thankful to Ms. Nisha Siddhardhan (Instrumentation in-charge) for their kind support during the project work. I also place my sincere thanks to non-teaching staff for their support and co-operation. I am highly grateful to my parents for their affectionate and moral support. They have always been source of inspiration for me. Above all, I thank The Almighty for giving me strength to complete this project. Last but not the least I extend my sincere thanks to all those who have helped me in one or the other way during my project work.

Saurav K. Rawat M Sc Final (Physical Chemistry)

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ABBREVIATIONS

RDA = Recommended Dietary Allowance AI = Adequate Intake UL = Upper Limit DDI = Daily Dietary Intake DRI = Dietary Reference Intakes MAL = Maximum Acceptable Limit SAM = Standard Addition Method AA = Atomic Absorption FAAS = Flame Atomic Absorption Spectroscopy HCL = Hollow Cathode Lamp MIBK = Methyl isobutyl ketone APDC = Ammonium pyrrolidine dithiocarbamate ND = Non Detectable PMT = Photomultiplier tubes LPG = Liquefied petroleum gas ppm = Parts per million Cu = copper Cr = chromium Pb = Lead Ni = Nickel Na = Sodium Fe = Iron Ca = Calcium

UL = The maximum level of daily nutrient intake that is likely to pose no risk of adverse effects. Unless otherwise specified, the UL represents total intake from food, water, and supplements. ND = Non detectable.

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INDEX

1. Introduction 6 2. Review of Literature 8 3. Sources of metals in alcoholic beverages 9 3.1. Raw materials 9 3.2. Substances added during brewing 9 3.3. Process type 9 3.4. Process equipment 10 3.5. Bottling process 10 3.6. Aging/storage 11 3.7. Adulteration 11 4. Effects of metals present in alcoholic beverages 12 4.1. Effects on the beverages 12 4.1.1. Positive aspects 12 4.1.2. Negative aspects 13 4.2. Effects on humans 14 5. Metal concentrations and limits 17 6. Metal removal 18 7. Speciation 19 8. Determination of metals in alcoholic beverages 20 9. Results and Discussion 31 10. Conclusion 32 (TABLE-I to TABLE IX) 33-41 References 42

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1. Introduction

The concentration of metals in many alcoholic beverages can be a significant parameter affecting their consumption and conservation. This derives from the negative and positive effects caused directly or indirectly by the presence of metals. Negative effects include beverage spoilage and hazing, as well as sensorial and health consequences. Positive effects include the removal of bad odors and tastes, participation in fermentative processes, provision of pathways for dietary intake of some essential minerals, and usefulness for authentication purposes. Because of all this, many metals are carefully monitored and regulated, which has resulted in the development of a plethora of analytical techniques for their analysis. Metals in alcoholic beverages are often determined by atomic absorption or emission techniques; however, the high cost of the instruments involved and the long sample-preparation times required often preclude their widespread use. Electrochemical methods are an option for such analyses.

In the manufacturing process employed for most alcoholic beverages we distinguish between fermentation and distillation. In the second process, the distillate is sometimes obtained directly from a previously fermented product, which carries over the volatile compounds (water, alcohols, aromatic substances like acids, aldehydes, ketones, esters, etc.) and modifies the composition previously existing in the raw product. Due to the low volatility of the seven elements considered in this project (Fe, Cu, Ni, Cr, Pb, Ca, Na), their levels in the resulting distilled alcoholic beverages have to be lower. In the distillation process copper still is usually employed because copper is very malleable, a good heat conductor, and resistant to corrosion, as well as playing a role as catalyser in certain chemical reactions and of complexing molecules unpleasant from the organoleptical point of view. Consequently, an increase in the Cu levels of the resulting distilled brandy would be expected.

Additionally, it is known that in the ageing of brandies obtained via distillation, the activation of chemical reactions that require oxygen are probably enhanced by the existence of ions of heavy metals like Cu. Several techniques have been employed for the determination of minerals in wines and beverages:

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inductively coupled plasma mass spectrometry (ICP-MS) using a double-focusing sector fields; AAS with flame or electro thermal atomization; or electro analytical methods such as differential pulse anodic stripping voltammetry (DPASV). In the available food composition and nutrition tables, the levels of the essential minerals studied, especially those for the trace elements Cu and Ni are not usually collected. Therefore, the aim of this project was to review the various sources and concentrations of metals (e.g., Ca, Cr, Cu, Fe, Pb, Ni and Na) in 6 alcoholic beverages (Whisky, Vodka, Rum, Brandy, Spirit and Deshi liquors), most commonly consumed in India and over the world their effects, speciation, removal methods and detection by FAAS technique and flame photometer.

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2. Review of Literature Akrida-Demertzi, K., Koutinas, A.A., 1992 estimated the effect of copper,

potassium, sodium and calcium on alcoholic fermentation of raisin extract and

sucrose solution.

Almeida Neves, et al 2007. Studied about the elimination of copper(II) in

sugar-cane spirits.

Bakalov, N., Angelov, V., Gidov, G., Angelov, B., Bolgurov, S., 1989.

Worked on the technology for the removal of metals from fresh wine distillate

and brandy.

Dugo and Salvo, et al, 2004. Determined the concentration of Ni(II) in

beverages.

Cyro, T.N., 1976. Determined the level of calcium, magnesium, iron, copper,

and zinc levels in distilled, fermented beverages-using atomic absorption

spectrometry.

Mekhuzla, N.A., Panasyuk, A.L., Temkina, V.Y., 1978. Used the trisodium

salt of nitrilotrimethylphosphonic acid for the demetallization of brandies.

Mena, et al, 1997. Determined the level of lead contamination in wines and

other alcoholic beverages by atomic absorption spectrometry..

Pohl, et al. 2007. Studied the Fractionation analysis of metals in dietary

samples using ionexchange and adsorbing resins.

Reilly, C., 1972. Zinc, iron, and copper contamination in home-produced

alcoholic drinks.

Reilly, C., 1973. Heavy metal contamination in home-produced beers and

spirits

Servadio,et al studied the stabilization of vodka.1975

Shukla, J., Pitre, K.S., 1998 studied Simultaneous trace metal analysis in wine

samples.

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3. Sources of metals in alcoholic beverages

Metals find their way into alcoholic beverages at different stages and through various sources including raw materials, brewing, process type and equipment, bottling, aging/storage, and adulteration as discussed below.

3.1. Raw materials Several metal ions can be taken up from the surrounding soil by plants from which an alcoholic beverage is prepared. For instance, the type of soil (i.e., its geogenicity), its agrochemical treatments (e.g., the use of pesticides and fungicides), and the surrounding environmental pollution have implications in the mineral content of many beverages. In this way, wines from vineyards in coastal areas are richer in Na. Pesticides, fungicides, and fertilizers containing Cd, Cu, Mn, Pb, and Zn compounds can derive in increased contents of these metals in the alcoholic beverage. Most of the Mg found in beer can be introduced with the malt. Cu in beer comes mainly from raw materials; on the contrary, only a small percentage of the final Cu content in whiskey comes from the barely from which the spirit is distilled.

3.2. Substances added during brewing

Hops, acids, bases, silica gel, dilution water, flavoring agents, additives, and stabilizers are potential sources of metal ions in the brewing process. For example, the main source of Cu in wine is the CuSO4 added to remove sulfidic odors. The acidity of the liquor to be distilled may be important in this regard (e.g., in whiskies), since more acidic beverages tend to contain more Cu. Addition of fining and clarifying substances (e.g., flocculants) to reduce turbidity can bring about an increase in Na, Ca or Al in wine.

3.3. Process type Major differences in metal content (i.e., Ni) have been found among alcoholic beverages depending on their processing. In this way, certain fermented beverages (e.g., wine and beer) contain several times more Ni than distilled beverages (e.g., brandy, whiskey, and vodka).

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3.4. Process equipment This is frequently a key source of metal ions in the final products. Several examples follow.

The concentration of Cu coming from process equipment in vodka is twice as much that coming from the raw materials.

The main source of Cu in whiskey is the copper still used for its distillation. Corrosion of tequila-distillation equipment (made of Cu) provokes the presence

of this metal in the final product. Storage of vodka in metal containers (e.g., low-quality steel or Cu alloys) results

in their corrosion with the concomitant introduction of metals into the liquor. The Zn, Fe, and Cu contained in home-produced alcoholic drinks can be essentially unrelated to the material fermented as it primarily depends on the vessel materials. The temperature in the distillate and the degree of still utilization affect the Cu content in whiskey. The Fe-content in pulps and musts increases due to the Fe in concrete tanks used for the storage of raw materials. Contact of wine with process equipment, pipes, casks, and barrels is the usual source of Al, Cd, Cr, Cu, Fe, and Zn. The main sources of heavy metals in the production of an anistype beverage are the bronze pot stills. Lead plumbing can add Pb to beverages.

3.5. Bottling process Bottling water and equipment may also introduce metals in beverages. For example, the content of Ca, Mg, and Na in brandies depends on the quality of water used for dilution after distillation. The modification of certain imported alcoholic beverages ‘‘for the purpose of bottling and sale by the addition of distilled or otherwise purified water to adjust the beverage to a required strength’’ is sometimes allowed (DJC, 2005). It is noteworthy that when metallic capsules seal alcoholic beverage bottles, some Pb may be carried over (Mena et al., 1997).

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3.6. Aging/storage Possible effects caused by metals during these stages are multiple. For example, Fe(III) and Mn(II) affect the stability of old wines and modify their sensorial quality after bottling since they are believed to activate molecular oxygen by forming reactive oxygen species (e.g., hydroxyl radicals); this is possible due to their electronic configurations involving unpaired electrons that may interact quantum mechanically with the dioxygen triplet. Likewise, Fe catalyses the oxidation of polyphenolic substances and Mn facilitates acetaldehyde formation; the products of these reactions yield undesirable precipitates. Metal complex formation is also common at this aging/storage state, which may alter a significant number of beverage parameters. Customeraccessible containers for alcoholic beverages include metal cans, glass bottles, plastic containers, and paperboard cartons, and the containers themselves sometimes are a source of metal ions in the beverage. For example, Cu and Zn can be introduced into beer by welded cans (Mayer et al., 2003). In fact, the Zn concentration in a specific brand of bottled beer was measured at 0.33 ppm, whereas in canned beer it reached 0.87ppm (Weiner and Taylor, 1969). On the contrary, the Ni content in canned beer as compared with glass-bottled beer is not higher (Dugo et al., 2004).

3.7. Adulteration This term comes from the Latin word adulterare (i.e., to defile or falsify), and means to make something impure by the addition of extraneous, inferior, or improper ingredients. As far back as 1875, legislation has prohibited the adulteration of drinks (MacRae and Alden, 2002). Unfortunately, Pb and other metallic impurities can enter beverages during adulteration practices (Ijeri and Srivastava, 2001). For example, adulterated vodka has been found to contain an excess of Ca and Mn ions (Servadio et al., 1975).

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4. Effects of Metals Present in Alcoholic Beverages

These effects can be classified according to the final subject they act upon.

4.1. Effects on the beverages These include negative or positive aspects as described below.

4.1.1. Positive aspects Contrary to the above, some metals enhance the flavor of wine (Esparza et al., 2005). One plausible indirect pathway for this effect involves the binding of sulfur derivatives to reduce the sulfury flavor (e.g., Cu in whiskey, sugar cane spirits, and cognac), as confirmed by thermal desorption gas chromatography (Reaich, 1998; Richter et al., 2001). Cu is also involved in the reduction and formation of congeners throughout the distillation of whiskey. A higher concentration of Ca2+

ions results in their uptake by Saccharomyces cerevisiae cells, which reduces cell growth. Yeasts consume Ca, Cu, Fe, K, Mg and Zn and therefore their concentrations tend to decrease (Pohl, 2007b); a substantial cofactor role occurs in some cases. Another useful aspect of the presence of specific metals in alcoholic beverages involves their use in quality analysis and in authentication (fingerprinting) due to their typical stabilities. Chemometrics and pattern recognition methods are used for the distinction of beverages according to their origin, quality, variety, type, and other features. For example, the Cu concentration in a malt Scotch whiskey is 1.5–3.5 times greater than that in a blended (grain) Scotch whiskey. Another case in point is the metal content in zivania (a Cypriot traditional drink), which allows its differentiation from spirits produced in other countries. Likewise, the analysis of Na and Mg is often sufficient to discriminate among different wines; even the analysis of Mg alone may serve such a purpose (Kokkinofta et al., 2003). In the same manner, the metal profile of brandies depends on their elaboration process; accordingly, the content of metals is sometimes used as a chemical descriptor for classifying different kinds of brandies. Statistical techniques are most helpful here, since classification procedures based on artificial neural networks lead to a predictive ability up to 90%.

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4.1.2. Negative aspects Minerals may generate irreversible turbidities in liquors; the formation of sediments is a long lasting process that may depend on the contents of all metals and substances present, the redox potential, pH, and temperature. Thus, the establishment of absolute limiting values for metal contents is rather difficult. Turbidity typically increases with metal concentrations. For example, maximum turbidity in soplica liquor was observed with the addition of Ca after more than 2 months of storage, whereas a smaller Ca addition (i.e., 20mg Ca/L) barely changes a Winiak liquor’s clarity; Fe can likewise contribute to turbidity. Some metal ions promote the development of turbid colloids during cognac storage (Russu et al., 1985). Small amounts of Cu (ca. 3mg Cu/L) cause maximum turbidity after 5 months of storage of some beverages (Trawinska, 1977); it can also be a factor in the formation of hazes (called copper casse) in wine and in beer (Mayer et al., 2003; Green et al., 1997). Cu also contributes to the oxidation of beer and imparts a coppery, unpleasant metallic taste (Mayer et al., 2003); here, even small amounts of this metal (e.g., 0.15mg Cu/L) can cause gushing (Mayer et al., 2003). Radical formation in sugar cane spirits depends mainly on the Cu content, as detected by electron spin resonance (Bettin et al., 2002). Color changes in some alcoholic beverages (e.g., wine) can be attributed to different factors (Esparza et al., 2005): (a) complex formation of Fe, Cu, Al or Mg with anthocyanins and tannins, (b) the presence of metal–polyphenol complexes, (c) hyperchromic of batochromic effects originating from copigmentation processes, and (d) an exogenous contribution of selected metal cations that might result in favorable color modification. Examples of additional color changes include the following- Cu can increase the rate of oxidative spoilage of wine, which ultimately results in its browning (Pohl, 2007b). There is a relationship between the color of a brandy and its Fe and Cu contents (Varju, 1972); if the concentration of metals in brandy reaches a critical limit, they promote precipitation (Varju, 1972). Although a significant correlation is not found between the content of metals (e.g., Fe, Cu, and Zn) and the color development in sake (a traditional Japanese drink) during the first 2 years of storage, more extensive darkening is observed after 3 years (Kondo, 1966). Another effect involves the influencing of key organoleptic properties by high concentrations of certain metals. A case in point is the negative influence of Zn in wines (Salvo et al., 2003).

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4.2. Effects on humans Metal ion effects on humans can be quite varied, and their detailed coverage falls beyond the scope of the present review; key information can be found in toxicology treatises. To exemplify this issue, among the best known effects are those of the toxic Pb (II) and Cd (II) ions. In fact, Benjamin Franklin in the eighteenth century noticed that certain homemade rum was causing human paralysis; he successfully traced the problem to Pb-containing equipment (UMA, 2005). Chromium—specifically Cr (VI)—can also be toxic; fortunately, dangerous concentrations are not common in alcoholic beverages. For instance, in a wide sampling of beverages (including wine, beer, cider, brandy, rum, whiskey, gin, vodka, anisette, and liquors) its concentration was found to be below 0.025 mg/L (Lendinez et al., 1998). On the contrary, the presence of certain metal ions in alcoholic beverages is beneficial in that it may provide an intake path for necessary nutrients in consumers (Mayer et al., 2003). For example, moderate wine consumption provides important amounts of nutritional requirements of several essential metals like Ca, Co, Cr, Cu, Fe, K, Mg, Mn, Mo, Ni, and Zn (Pohl, 2007b). The effects and the functions of the metal determined during this project are given below- According to DRI the DDI and UL of the same metals are listed in table 1-5.

Impacts of Studied Metals in Biological System

Copper- copper is an essential constituent of many metallo-proteins and enzymes, involved in electron transfer, oxygenation and oxidation processes. Hence, deficiency of copper causes deactivation of these processes, leading to anaemia (ceruloplasmin deficiency), and loss of hair pigment (Tyrosine deficiency). Deficiency of Cu(II) containing enzyme, cytochrome C oxidase, causes reduced arterial elasticity and stunted growth in adults and Meneke’s disease in children, resulting in kinky hair, retarded growth, and respiratory problem, severely limiting life span.

If synthesis of ceruloplasmin is hindered, the mechanism of the control of copper level in the biological system is damaged. This leads to accumulation of copper in liver, kidney and brain. Thus the central nervous system (CNS) is damaged, leading to tremors, rigidity and abnormality of the brain. Accumulation of copper in liver leads to Cirrhosis and ultimate death. This physical abnormality is called Wilson’s disease.

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External intake of small excess of copper causes gastro intestinal irritation and vomiting. Serious toxic effect is observed, if more than one gram of copper is taken at one time or there is continuous intake of 250 mg per day, for a period of time. The toxic effect occurs because of strong affinity of Cu(II) for the –SH group of the different enzyme proteins. The enzyme get deactivated, due to copper binding, and thus specific biochemical activity are inhibited, leading to physical disorders. (TABLE-I)

Chromium- It is involved in the metabolism of glucose in the mammals. Cr (III) and insulin both maintain the correct level of glucose in the blood. People can be exposed to chromium through breathing, eating or drinking and through skin contact with chromium or chromium compounds. The level of chromium in air and water is generally low. In drinking water the level of chromium is usually low as well, but contaminated well water may contain the dangerous chromium(IV); hexavalent chromium. For most people eating food that contains chromium(III) is the main route of chromium uptake, as chromium(III) occurs naturally in many vegetables, fruits, meats, yeasts and grains. Various ways of food preparation and storage may alter the chromium contents of food. When food in stores in steel tanks or cans chromium concentrations may rise. Chromium(III) is an essential nutrient for humans and shortages may cause heart conditions, disruptions of metabolisms and diabetes. But the uptake of too much chromium(III) can cause health effects as well, for instance skin rashes. Chromium(VI) is a danger to human health, mainly for people who work in the steel and textile industry. People who smoke tobacco also have a higher chance of exposure to chromium. Chromium(VI) is known to cause various health effects. When it is a compound in leather products, it can cause allergic reactions, such as skin rash. After breathing it in chromium(VI) can cause nose irritations and nosebleeds. Other health problems that are caused by chromium(VI) are: - Skin rashes - Upset stomachs and ulcers - Respiratory problems - Weakened immune systems - Kidney and liver damage - Alteration of genetic material - Lung cancer - Death

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The health hazards associated with exposure to chromium are dependent on its oxidation state. The metal form (chromium as it exists in this product) is of low toxicity. The hexavalent form is toxic. Adverse effects of the hexavalent form on the skin may include ulcerations, dermatitis, and allergic skin reactions. Inhalation of hexavalent chromium compounds can result in ulceration and perforation of the mucous membranes of the nasal septum, irritation of the pharynx and larynx, asthmatic bronchitis, bronchospasms and edema. Respiratory symptoms may include coughing and wheezing, shortness of breath, and nasal itch.

Carcinogenicity- Chromium and most trivalent chromium compounds have been listed by the National Toxicology Program (NTP) and International Agency for Research on Cancer (IARC) as having inadequate evidence for carcinogenicity in experimental animals. According to NTP, there is sufficient evidence for carcinogenicity in experimental animals for the following hexavalent chromium compounds; calcium chromate, chromium trioxide, lead chromate, strontium chromate,and zinc chromate. (TABLE-2) Iron- Hemoglobin is the oxygen carrying iron protein in mammalian blood. Chromatin material of nucleus having iron is an essential part in metabolic oxidations occurring in nucleus. Though essential, excess iron intake can cause acidity, vomiting and coma conditions. Excess metal gets deposited in different parts of the body, like liver, kidney and brain and can lead to their failure. (TABLE-3) Sodium- People who regularly eat foods high in sodium risk having diseases such as hypertension, Type II diabetes mellitus, respiratory complications, Dislipidemia, Gallbladder disease, osteoarthritis and some cancers (endometrial, breast, colon). Most of the daily sodium intake comes from salt. The DRI Upper Limit (UL) for Sodium in adults is 2300 mg/day. Calcium- The level of calcium in the body is usually controlled by vitamin D and parathyroid hormones. But, if there is a metabolic imbalance of calcium regulation, it gets deposited in the tissues, leading to their calciferation. Formation of stones cataract are due to calcium salt deposition. (TABLE-4) Nickel- it is an essential trace element for several hydrogenases and ureases enzymes. Its deficiency in food slows down the functioning of the liver in chicks. It is highly toxic to plants and moderately toxic to mammals. It is carcinogenic if present in higher concentrations in biological systems.

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It causes skin and respiratory disorders. It can produce bronchial cancer. It deactivates cytochrome C oxidase and also the enzymes, assisting dehydrogenation process, and thus inhibits biochemical processes. (TABLE-5) Lead- It has no known biological function. It is highly toxic to plants and is a cumulative poison for mammals. It inhibits the synthesis of hemoglobin in mammals and is highly toxic for central nervous system. Lead tertraethyl used in gasoline as an antiknock and lead pigments are serious health hazard. Lead gets deposited in the softer tissues. From there, the reversibly fixed lead passes to the blood stream. Like transition metals, lead has strong affinity for the –SH group of the enzymes and hence it gets bound to the enzymes strongly and deactivates them. In the blood stream, lead is known to inhibit the activity of several enzymes, involved in the synthesis of heme. Excess lead lowers the formation of delta amino levulinic acid, its conversion to porpho-bilinogen and also the conversion of protoporphyniogen to protoporphyrin IX. Thus the biosynthesis of heme is inhibited, leading to anemia. Lead also affects the biosynthesis of bones, because, divalent lead replaces calcium in bone. Deposition of lead in brain results in its reduced activity, leading to depression, nervousness and lack of concentration. Excess lead leads to damage of kidney, liver and intestinal track, with consequent loss of appetite, muscle and joint pain, weakness and tremors. Excess lead also causes dental carries and abnormalities in female reproductive system. (TABLE-6)

5. Metal concentrations and limits The concentrations of metals in alcoholic beverages can vary widely, metals are sometimes present in alcoholic beverages at rather high concentrations. In fact, extreme metal ion concentrations in home-produced beer and spirits from different parts of Africa, India, Europe, and Canada revealed concentrations of up to 58, 68, and 245 mg/L of Cu, Zn, and Fe, respectively (Reilly, 1973). Homemade—but commercially available—alcoholic beverages in Tanzania can have up to twice the World Health Organization recommended maximum for Zn in drinking water (i.e., 5 mg/L), and one brewed beverage was found to contain toxic amounts of Mn (12.8 mg/L) (Nikander et al., 1991). These instances dramatically demonstrate that the establishment of limits is indispensable. Table IX shows examples of metal concentration limits established by various regulatory agencies for selected beverages.

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Note that limits imposed for alcoholic beverages are higher than those established for water utilized in human consumption. This reflects the lower intake of the former (Green et al., 1997). For example, alcoholic drinks usually contribute only marginally to typical dietary Cu intakes (0.05 mg/day) (Sadhra et al., 2007). Regrettably, marked absences in legislation do exist (Baldo and Daniele, 2005). Metal ion concentrations in wastewaters of breweries and alcoholic beverage factories are typically below the limits established for the introduction of such concentrations into municipal treatment systems. Metals that are sometimes present in concentrations near the upper regulation limits include Hg and Cu (Koller and Sahlmann, 1987).

6. Metal removal Methods for metal removal from alcoholic beverages can be exemplified by the following cases: An alternative metal removal scheme consists in raising the pH (e.g., in wine and brandy) with NaHCO3 or CaCO3 to ca. 4.5–5, then adding tannins or tannic acids and allowing the mixed substances to react for several days. Finally, gelatin and bentonite are added to react with the metal tannates and the mixture is stirred, decanted, and filtered. Large reduction factors in Cu, Fe, and Zn concentrations are thus obtained. Ion exchange resins decrease the metal content of certain distillates to the allowable limits (Hodejeu et al., 1972). (As a case in point, we have obtained up to 99.6% removal of Cu from tequila.) MgCO3 and CaCO3 can act as cationic exchangers to remove Cu(II) from sugar cane spirits. Cu is removed from spirituous beverages by precipitation with rubeanic acid. Some metal ions (e.g., Cu and Fe) are removed from alcoholic beverages by adding polymers that contain metal-binding groups (Detering et al., 1991; Kern, 1987). Certain chelating agents (e.g., the trisodium salt of nitrilo tris-methylenephosphonic acid) in wine and cognac can precipitate Fe, Al, and Cu within 1 week (Romantseva et al., 1982). The same compound can be used in brandy to precipitate Fe and Al (Cu does not precipitate here) (Mekhuzla et al., 1978).

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Potassium hexacyanoferrate (II) binds some metals and sulfides present in alcoholic beverages, although it cannot be used in applications where the Cu content exceeds that of Fe; besides, it may release poisonous HCN.

7. Speciation The chemical form in which an element, ion or compound is found in a given medium depends on the physicochemical conditions of the system. Thus, the distribution of species in an aqueous medium (e.g., an alcoholic beverage) is contingent on pH, composition, temperature, and the oxidation-reduction potential of the solution. These variables define precipitation, dissolution, redox, and complexation reactions. Biological phenomena (e.g., bioaccumulation) frequently depend on the chemical form or speciation of a metallic ion. The redox environment can determine some properties of metallic and non-metallic species. For example, when arsenic is present in oxidizing environments as As(V), its toxicity is very low; however, its reduced form, As(III) is highly poisonous. The opposite occurs with Cr(VI), which is much more toxic than its reduced counterpart, Cr(III). Solubility can also be severely affected by the redox environment. Examples of this are Fe(II) and Mn(II) species, typically soluble in aqueous solutions deficient in dioxygen, whose oxidized forms precipitate quite easily. Similarly, ligand speciation may drastically affect the nature and physicochemical properties of the metallic complexes they form. Bioavailability and toxicity of metal ions in aqueous systems are often proportional to the concentration of the free metal ion and thus decrease upon complexation. However, some metal compounds are more dangerous than the metallic element itself (e.g., methyl mercury vs. mercury). Because of the aforementioned reasons, determination of the total concentration of a metal in a given matrix often does not adequately or effectively characterize it; as a result, speciation has gained considerable ground. Speciation is the process that yields evidence of the atomic or molecular form of an analyte. It can be defined either functionally (e.g., the determination of species that have certain specific functions), or operationally (e.g., the determination of the extractable forms of an element from a given matrix) (Ibanez et al., 2007). Examples of the importance of speciation in alcoholic beverages include the following:

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Metals can be either in a state ‘‘bound’’ to the wine matrix, or ‘‘unbound’’ in solution. Some analytical methods (e.g., anodic stripping voltammetry, ASV) may not detect the total amount of the metal in the sample, but merely identify the unbound fraction. This problem has been circumvented by means of the standard addition method (SAM) used for their quantification (Akkermans et al., 1998). The concentration of metals in distillates may be affected by complex formation in the stills, given that such complexes may be less volatile than the metal ions themselves and thus their final concentration in the distillate would be smaller than what could be predicted (Adam et al., 2002). Bioavailability is usually more important than the total concentration of a metal. Labile metal complexes (ML) are normally more toxic because they are more easily absorbed by organisms (Arcos et al., 1993). Organic complexes of Cu are present in residues of the distillation process of Scotch whiskies (Adam et al., 2002). Cu and Zn form soluble complexes in the spent wash of whiskey distilleries. Cu is believed to be bound to an organic fraction containing carbohydrate and ninhydrin, whereas Zn is complexed by a lower molecular weight fraction containing hexose and phenolic moieties (Quinn et al., 1982). Fe can complex certain triketones, the main bittering compounds in beer. In doing so, it may induce their deprotonation and thus contribute to changes in beer quality during ageing (Blanco et al., 2003).

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8. Determination of metals in alcoholic beverages Analytical methods frequently require sample pre-concentration and/or pretreatment for the destruction of the organic matrix such as wet digestion, dry ashing, and microwave oven dissolution (Salvo et al., 2003; Dugo et al., 2004). Common analytical methods include atomic absorption spectrometry (AAS), atomic emission spectrometry (AES) and inductively coupled plasma–optical emission spectrometry (Camean et al., 2001). Ion chromatography is also used for the analysis of metals, for example in vodka (Obrezkov et al., 2000). The SAM can be used with some of these schemes; for example, Cu is determined in cachaza with SAM–AAS (Farias-Almeida et al., 2003). Another alternative involves a previous fractionation step of the metals from the beverage matrix using either ionic resins or adsorbents before analyzing food or wine samples (Pohl, 2007a). Unfortunately, most of these methods involve rather expensive instrumentation (with its concomitant high-cost maintenance) that precludes their widespread use among alcoholic beverage producers. Electrochemical methods have gained considerable ground in the analysis of alcoholic beverages because of their simplicity, rapidity, and relative low cost but in the present study FAAS and Flame Photometer were used because of their exact accuracy and less time consuming property.

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EXPERIMENTAL

Sampling The samples of alcoholic beverages analysed in the present study (n = 6) were purchased from the locality of Agra, Uttar Pradesh. Distilled alcoholic beverages (n = 6) were selected from the most popular brands consumed by the local inhabitants taking into consideration their food habits. The samples were brought to the laboratory of the Department of Chemistry of St. John’s College, Agra where they were stored at 18 0C until analysed. Materials and methods Apparatus A Perkin-Elmer AAnalyst100 double beam atomic absorption spectrophotometer (Perkin-Elmer corp., CT) was used at a slit width of 0.7 nm, with hollow cathode lamps for mineral measurements by FAAS. Samples were atomized for Cr, Cu, Fe, Pb, and Ni. All analyses were performed in peak height mode to calculate absorbance values. SYSTRONICS Flame photometer 130 was used for the estimation of Ca and Na. Sample preparation All solutions were prepared from analytical reagent grade reagents, for e.g., Commercially available 1,000 μg/mL Cu [prepared from Cu(NO3)2.3H2O in 0.5 M HNO3] were used. The water employed for preparing the standards for calibration and dilutions was ultra pure water with a specific resistivity of 18 m_ cm-1 obtained by filtering double-distilled water through a Milli-Q purifier (Millipore, Waters, Mildford, MA) immediately before use. Sample treatment and analysis Cu, Fe and other elements in spirits, gin, vodka, whisky, rum and similar beverages can be analysed by atomic absorption. It is accurate, fast and needs no special sample preparation. The samples are aspirated directly and standards are made up in alcohol to match the content of the particular sample. Calcium and sodium can be easily analysed by Flame Photometer. Standards can be prepared as follows-

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Calcium – 1000 ppm Dissolved 2.497 g CaCO3 in approx 300 ml glass distilled water and added 10 ml conc. HCl diluted to 1 litre. For calibration 20, 40, 60, 80 and 100 ppm solutions were prepared from the stock solution.

Sodium- 1000 ppm Dissolved 2.5416 g NaCl in one litre of glass distilled water. For calibration 20, 40, 60, 80 and 100 ppm solutions were prepared from the stock solution.

ATOMIC ABSORPTION SPECTROSCOPY (AAS) INTRODUCTION/ BASIC PRINCIPLE Spectroscopy is the measure and interpretation of electromagnetic radiation absorbed, scattered or emitted by atoms, molecules or other chemical species. When the electromagnetic radiation absorbed by atoms is studied, it is called atomic absorption spectroscopy. This absorbance is associated with changes in the energy state of the interacting chemical species since each species has characteristics energy states. Atomic absorption spectroscopy (AAS) or atomic absorption (AA) or atomic absorption spectrometry (AAS) uses the absorption of light to measure the concentration of gas-phase atoms. Since samples are usually liquids or solids, the analyte atoms or ions must be vaporized in a flame (such as air-acytelene flame) or graphite furnace that contains the free atoms become a sample cell. The free atoms absorb incident radiation focused on the from a source external to a flame and reminder is transmitted to a detector where it is changed into an electrical signal and displayed, usually after amplification, on a meter chart recorder or some other type of read-out device. The sample solution is introduced as an aerosol into the flame and atomized. A light beam from the source lamp (hollow cathode lamp, HCL) composed of that element (intense electromagnetic radiation with the wavelength exactly the same as that is absorbed maximum by the atoms) is directed through the flame, into a monochromator and onto a detector that measures the amount of the light absorbed by the atomized element in the flame (Fig. 1). Because each metal has its own characteristic absorption wavelength, the amount of energy at the characteristics wavelength absorbed in the flame is proportional to the concentration of the element in the sample over a limit concentration range. The atoms absorb ultraviolet or visible light and make transitions to higher electronic energy levels. The analyte concentration is determined from the amount of absorption.

24

Applying the Beer-Lambert law directly in AAS is difficult due to the variations in the atomization efficiency from the sample matrix, and non uniformity of concentration and path length of analyte atoms (in graphite furnace AA). Concentration measurements are usually determined from a working curve after calibrating the instrument with standard of known solution.

ATOMIC TRANSITION THEORY The probability that an atomic spectroscopic transition will occur is called the transition probability or transition strength. This probability is determine the extent to which an atom is absorb light at a resonance frequency, and the intensity of the emission lines from an atomic excited state. The spectral width of a spectroscopic transition depends on the widths of the initial and final states. The width of the ground state is essentially a delta function and the width of an excited state depends on its lifetime.

INSTRUMENTATION

Light source- The light source is usually a hollow cathode lamp of the element that is being measured. Lasers are also used in research instruments. Since laser are intense enough excite atoms to higher energy levels, they allow AA and atomic fluorescence measurements in a single instrument. This disadvantage of these narrow-band light sources is that only one element is measurable at a time.

Atomizer- AA spectroscopy requires that the analyte atoms be in the gas phase. Ions or atoms in a sample must undergo desolvation and vaporization in a high temperature source such as a flame or graphite furnace. Flame AA can only analyze solutions, while graphite furnace AA can accept solutions, slurries or solid samples. Flame AA uses a slot type burner to increase the path length, and therefore to increase the total absorbance (see Beer-Lambert law). Sample solutions are usually aspirated with the gas flow into a nebulizing/mixing chamber to form small droplets before entering the flame. The graphite furnace has several advantages over a flame. It is much more efficient atomizer than a flame and it can directly accept very small absolute quantities of sample.

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Samples are placed directly in the graphite furnace and the furnace is electrically heated in several steps to dry the sample, ash organic matter, and vaporize the analyte atoms.

Light separation and detection- AA spectrometers use monochromators and detectors for UV and visible light. The main purpose of the monochromator is to isolate the absorption line from background light due to interferences. Simple dedicated AA instruments often replace the monochromator with a band pass interference filter. Photomultiplier tubes (PMT) are the most common detectors for AA spectroscopy.

26

AAS AT A GLANCE

Principle- It measures the decrease in light intensity from a source (HCL) when it passes through a vapour layer of the atoms of an analyte element. The hollow cathode lamp produces intense electromagnetic radiation with a wavelength, exactly the same as that absorbed by the atoms, leading to high sensitivity. Construction- It consists of a light source emitting the line spectrum of the element (HCL), a device for the vaporizing the sample (usually a flame), a means of isolating an absorption line (monochromator) and a photoelectric detector with its associated electronic amplifying equipment. Operating Procedure- HCL for the desired elements is installed in instrument and wavelength dial is set according to the table and also slit width is set according to the manual. Instrument is turned on for about 20 min to warm up. Air flow rate and acetylene current are adjusted according to the manual. Standard solution is aspirated to obtain maximum sensitivity for the element is adjusting nebulizer. Absorbance of this standard is recorded. Subsequent determinations are made to check the consistency of the instrument and finally the flame is extinguished by turning off first acetylene flame and then air. Lamps- Separate lamp (HCL) is used for each element since multi element hollow cathode lamps generally provide lower sensitivity. Vent- A vent is paced about 15-30 cm above the burner to remove the fumes and vapours from the flame.

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Determination of Heavy Metals- Reagents- 1. Air- cleaned and dried through a filter air. 2. Acetylene- standard, commercial grade 3. Metal free water- all the reagents and dilutions were made in metal free water 4. Methyl isobutyl ketone (MIBK)- Reagent grade MIBK is purified by re-distillation before use. 5. Ammonium pyrrolidine dithiocarbamate (APDC) solution- 4 g APDC is dissolved in 100 ml water. 6. Conc. HNO3 7. Standard metal solutions: Five standard solutions of 0.01, 0.1, 1, 10 and 100 mg/L concentrations of metals such as Cr, Mn, Fe, Ni, Cu, Zn, Cd and Pb for instrument calibration and sorption study are prepared by diluting their stock solution of 1 g/l, i.e., 1 ml = 1 mg metal. Procedure- a. Instrument operation- same as above. Solution is aspirated into flame after adjusting the final burner position until flame is similar to that before aspiration of solvent. b. Standardization- five standard metal solutions in metal free water are selected for the standardization of the instrument. Transfer standard metal solutions and blank to a separatory funnel and added 1 ml APDC, 10 ml MIBK and was shaken vigorously. Aqueous layer is drained off and organic extract was directly aspirated into the flame. c. Sample analysis- Atomizer (nebulizer) is rinsed by aspirating water saturated MIBK and organic extracts obtained by above the method were directly aspirated into the flame. d. Calculation- concentration of each metal ion in milligrams per litre is recorded directly from the instrumentation readout.

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EXPERIMENTAL

29

FLAME PHOTOMETER Flame photometry is an atomic emission method for the routine detection of metal salts, principally Na, K, Li, Ca and Ba. Quantitative analysis of these species is performed by measuring the flame emission of solution containing the metal salts. Solutions are aspirated into the flame. The hot flame evaporates the solvent, atomizes the metal, and excites a valence electron to an upper state. Light is emitted at characteristic wavelengths for each metal as the electron returns to the ground state. Optical filters are used to select the emission wavelength monitored for the analyte species. Comparison of emission intensities of unknown to either that of standard solution, or to those of an internal standard, allows quantitative analysis of the analyte metal in the sample solution. Introduction- SYSTRONICS flame photometer 130 is an instrument with which it is possible to estimate, with speed and accuracy, minute quantities of sodium (Na), Potassium (K), Calcium (Ca) and Lithium (Li). The principle of operation is simple. The fluid under analysis is sprayed as a fine mist into a non-luminous (oxidizing or colorless) flame which becomes colored according to the characteristic emission of the metal. A very narrow band of wavelength corresponding to the element (Na: 589 nm, K: 768 nm, Ca: 622nm, Li: 671 nm) being analysed is selected by a light filter and allowed to fall on a photo-detector whose output is measure of concentration of the element. The output of photo-detector is connected to an electronic metering unit which provides digital readouts. Before analyzing the unknown fluids, the system is standardized with solutions of known concentrations of the element of interest. The total system consists of two units- 1- Main unit, 2- Compressor unit. The main unit consists of an atomizer (for aspiration of solutions), mixing chamber, burner, optical lens, light filters, photodetectors, control valves and electronic circuit. Compressed air (oil free) from the compressor unit is supplied to the atomizer. Due to a draught of air at the tip of the atomizer, the sample solution is sucked in and enters in the mixing chamber as a fine atomized jet. Liquefied petroleum gas (LPG) or laboratory gas from a suitable source is also injected into mixing chamber at a controlled rate. The mixture of gas and atomized sample is passed on to the burner and is ignited. The emitted light from the flame is collected by a lens and is passed through an appropriate filter (Selectable for different element). The filtered light is then passed on to energize a sensitive photo-detector, the output of which is applied to the electronic circuit for readout.

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OPERATING PROCEDURE AND SAMPLE ESTIMATION Once the burner is ignited and set, followed the steps described below- Put on the mains supply to the unit. Digital display turned on. Turned the SET F.S. COARSE and FINE controls in maximum clockwise position. Select appropriate filter with the help of Filter Selector wheel (Na on the left side and K on the right side). Feed distilled water to the atomizer and wait atleast for 30 seconds. Adjust the SET REF. COARSE and FINE controls for a zero readout as nothing aspirated, for K only. Aspirate 1 mEq/L of Na solution (or the standard 1.0 / 0.01 mEq/L of Na/K solution). Wait atleast 30 s and then adjust the SET REF. COARSE and FINE controls for a readout of 100 for, Na only. Aspirate the standard mixed 1.7/0.85 mEq/L of Na/K solution and wait atleast for 30 s. Adjust SET F.S. control of the Na side for a readout of 170 and that of the K side for a readout of 80. The unit stands calibrated. For a recheck, aspirate the standard mixed solution of 1.0/0.01 mEq/L of Na/K. the readout for Na and K should be close to 100 and 10 respectively. Then feed sample solution to the atomizer to get the relative concentration. Wait atleast for 30 s before taking the reading.

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9. Results and discussion Levels of measured metals (Ca, Cr, Cu, Fe, Pb, Ni and Na) in alcoholic beverages from Agra are shown in TABLE - VIII. A variability in the concentrations of the seven elements analysed can be noted. This finding can be related to the drink type, manufacturing and bottling process. Drinks of high alcoholic content Ca, Cr, Cu, Fe, Pb, Ni and Na levels in the whisky, vodka, rum, spirit, deshi-liquor, and brandy samples analysed are summarized in TABLE-VIII, where a variation can be observed among the different samples. The variability is even more pronounced in the case of Cu and Fe. In drinks of high alcoholic content, Cu and Fe concentrations determined in the present study are considerably higher in Brandi and whisky samples (TABLE-VIII). In liquors, although Ca contents measured by us are higher than those shown in TABLE-IX. It is interesting to note that many of the food composition and nutrition tables included in TABLE-IX contain measurements of mineral content for spirits, although they only collect trace or 0.00 μg L-1 levels for Ca. Due to the different manufacturing processes used to produce the liquors and complex brandies we found that Cu level measured in liquor and vodka was lower than those found in the group of complex brandies (TABLE-VIII). This result could be related to the use of copper stills for complex brandies (whiskies, rums and brandies) which increases Cu concentrations. Since Cr and Pb both are very hazardous to health and their concentration must be very low, it was found to be also. Table-VIII shows the very less concentration of both the elements in all the beverages studied. A little bit amounts of these metals would have been added due to processing steps like bottling, aging / storage and adulteration etc. All the metal concentrations were under the DL, PL and MAL limits. Even Brandi was found to be a good source of Fe and Cu.

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10. Conclusions Levels of iron, copper, chromium, nickel, lead, calcium and sodium were measured in alcoholic beverages (Whisky, vodka, rum, brandy, spirit and deshi liquors) using flame atomic absorption spectrometry (FAAS) and flame photometer. A critical review is offered concerning the different sources, effects, concentrations, removal methods, speciation, and analysis of metals (e.g., Ca, Cr, Cu, Fe, Pb, Ni and Na) present in a variety of alcoholic beverages. Mineral concentrations were found to be significantly different between the six alcoholic products studied. In distilled alcoholic beverages, Cu measured concentrations were statistically different for each of the 6 groups of alcoholic beverages studied. Contrarily, Cu concentrations were statistically lower. Remarkably, for Cu, the concentration determined in brandy was statistically higher. From all studied elements, Cu was the one for which alcoholic beverages constitute a significant source (more than 10% of recommended daily intake). These findings are of potential use to food composition tables. Metals find their way into alcoholic beverages through many possible sources discussed here. Their effects on the beverages as well as on humans after consuming such beverages are quite varied. This then necessitates that metals be subject to regulations. FAAS offers distinct advantages for their analysis. Caution: Consumption of liquor is injurious to health

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TABLE-I

Nutrient Life Stage Group

RDA/AI (μg/d)

UL (μg/d)

Copper

Males 14-18 y 19-50 y Females 14-18 y 19-50 y Pregnancy 19-30 y 31-50 y Lactation 19-30 y 31-50 y

890 900 890 900 1000 1000 1300 1300

8,000 10,000 8,000 10,000 10,000 10,000 10,000 10,000

34

TABLE-II


RDA/AI (μg/d)

UL (μg/d)

Chromium


35 35 24 25 30 30 45 45

ND ND ND ND ND ND ND ND

35

TABLE-III


RDA/AI (mg/d)

UL (mg/d)

Iron


11 8 15 18 27 27 9 9

45 45 45 45 45 45 45 45

36

TABLE-IV


RDA/AI (mg/d)

UL (mg/d)

Calcium


1,300 1,000 1,300 1,000 1,000 1,000 1,000 1,000

2,500 2,500 2,500 2,500 2,500 2,500 2,500 2,500

37

TABLE-V


RDA/AI (mg/d)

UL (mg/d)

Nickel


ND ND ND ND ND ND ND ND

1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0

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TABLE-VI

LEAD

For Whom Amount Known To Cause Health Problems (μg/d)

FDA’s Recommended Safe Daily Diet Lead Intakes (μg/d)

For children under age 6 60 6

For children 7 and up 150 15

For Adults 750 75

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TABLE-VII Concentrations of Metals in mg/L Found in Various Alcoholic Beverages during This Project-

Element Beverage

Calcium (mg/L)

Chromium (mg/L)

Copper (mg/L)

Iron (mg/L)

Nickel (mg/L)

Sodium (mg/L)

Brandi 12 0.118 0.056 2.727 -1.135 29

Rum 37 -0.011 0.009 0.474 -1.069 ND

Spirit 16 0.177 0.045 1.371 -0.468 ND Whisky 20 -0.144 0.024 0.791 -1.169 18

Vodka 02 -0.078 0.016 -0.105 -0.797 ND

Deshi liquor

34 -0.144 0.025 -0.229 -1.383 ND

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TABLE-VIII

Examples of metal concentration limits for selected beverages

Metal Beverage Concentration limit, mg/L Obligatorietya Reference Cu Wine 0.5 Recommended Green et al. (1997) Pb Wine 0.2 Recommended Green et al. (1997) Cu Brandi 0.05 Recommender Mayer et al. (2003) Cu Rum 0.1 Recommended Richter et al. (2001) Cu Vodka 0.02 Recommended Green et al. (1997) Cu Whisky 0.04 Recommended Salvo et al. (2003), Dugo et al. (2005) Pb Brandi 0.01 MAL Dugo et al. (2005) Pb Rum 0.01 MAL Dugo et al. (2005) Pb Vodka 0.01 MAL Dugo et al. (2005) Pb Whisky 0.01 MAL Dugo et al. (2005) Cr Brandi 0.02 MAL Soufleros et al. (2004) Cr Rum 0.01 MAL Salvo et al. (2003), Dugo et al. (2005) Cr Vodka 0.01 MAL Soufleros et al. (2004) Cr Whisky 0.01 MAL Richter et al. (2001) Fe Brandi 3.0 Recommended (Almeida Neves et al., 2007) Fe Rum 0.8 Recommended (Almeida Neves et al., 2007) Fe Vodka 0.1 Recommended (Almeida Neves et al., 2007) Fe Whisky 1.0 Recommended (Almeida Neves et al., 2007) Cu Beer 0.05 Recommended Mayer et al. (2003) Cd Wine 0.1 MAL Salvo et al. (2003), Dugo et al. (2005) Cu Wine 1.0 MAL Richter et al. (2001) Pb Wine 0.2 MAL Commission of the European Communities (2006), Dugo et al. (2005) Cu Brandi 0.8 MAL Richter et al. (2001) Zn Wine 0.5–5 MAL Salvo et al. (2003), Dugo et al. (2005) Cu Cachaza 5 MAL Richter et al. (2001) Cu Fruit distillate 5 MAL Soufleros et al. (2004)

MAL = maximum acceptable limit.

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Table IX. Ca content in alcoholic beverages in the most

frequently used food composition and nutrition tables in different

countries

Reference (country)

Drinks of high

alcoholic content

Ca (μg L-1)

Souci et al. (1989,

Germany)

Whisky 15.0

Favier et al. (1995,

France)

Liquor 60.0

Mataix Verdu and

Carazo Martin (1995,

Spain)

Liquor 0.000

Muñoz et al. (1999,

Spain

Liquor 0.000

Holland et al. (2001,

United Kingdom)

Spirit Trace

USDA (2005, USA)

Liquor

Spirit

26.2

0.000

Farran et al. (2004,

Spain)

Liquor

Spirit

60.0

Trace

Spanish Ministry of

Agriculture (2004,

Spain)

Spirit

0.000

Food Institute

Informatics (2005,

Denmark)

Whisky

Gin

Rum

0.000

0.000

0.000

Ministry of Health

(2005, Canada)

Liquor

Spirit

0.000

0.000

42

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<http://archive.food.gov.uk/maff/archive/food/infsheet/

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<http://www.nix.oit.umass.edu/_fdsci150/pdfs/food_tox_pdfs/

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28. Varju, M., 1972. Determination of iron, copper, calcium, magnesium, and

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30. www.nap.edu

31. www.wikipedia.com

32. [email protected] (Analytical Research Laboratory)

33. http://mc.manuscriptcentral.com/tfac

http://www.nap.edu/

http://www.wikipedia.com/

mailto:[email protected]

http://mc.manuscriptcentral.com/tfac

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A project report on alcohol 2

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