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Masime Jeremiah O*, Wanjau Ruth, Murungi Jane, Onindo Charles
Department of Chemistry, Faculty of Science, Kenyatta University, Nairobi, Kenya
Key Words: Homemade brews, Nitrate, UV-Visible Spectroscopy
In recent years, an increasing interest in the determination of nitrate levels in food products has been observed, essentially due to the potential reduction of nitrate to nitrite, which is known to cause adverse effects on human and animal health. Nitrate is a naturally occurring compound that is part of the nitrogen cycle, as well as an approved food additive. It plays an important role in the nutrition and function of plants. Nitrate is an important component of vegetables due to its potential for accumulation; this
can be affected by a number of biotic and abiotic
factors [1]. Human exposure to nitrate is mainly exogenous through the consumption of vegetables, and to a lesser extent water and other foods [1]. Nitrate is also formed endogenously. In contrast exposure to its metabolite nitrite is mainly from endogenous nitrate conversion [1].
Nitrates may be found naturally in water or enter the supplies through a number of sources. Sources of nitrate pollution include; use of fertilizers, animal wastes, municipal and industrial waste, lightening among other sources. Nitrates are the products of aerobic stabilization of organic nitrogen [2]. They may
also enter water via fertilizers from agricultural
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runoffs. They can also be formed during thunderstorms and lightening [2]. The concentrations of nitrates in surface and ground water vary within wide limits depending on geochemical conditions, human and animal waste management practices and on industrial discharge of nitrogen compounds [2]. To protect those at risk, the maximum contamination level (MCL) for nitrate in water is 50 mg/l [3].
Intake of large doses of nitrate can be tolerated by adults but not infants and very young children who may suffer from methemoglobinaemia. The primary health hazard from drinking water with nitrate occurs when nitrate is transformed to nitrite in the digestive
system. Bacteria in the infant’s digestive tracts may
genetically impaired enzyme systems for metabolizing methemoglobin [5]. This is because of nitrites, which are produced by reduction of nitrates by microbial action either in the environment or in the body. Children aged between 12 and 14 years who drank water with a nitrate level of 105 mg/l were noted to have a slightly delayed reaction to light stimuli compared to control children drinking water with a nitrate level of 8 mg/l whose methaemoglobin level averaged 0.75 percent [5].
Nitrite is mainly produced as a result of oxygen depletion, anaerobic biological conditions dominate and reduction sets in [5]. Nitrate is reduced to nitrite,
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convert the relatively harmless nitrate to nitrite [4].
The nitrite oxidizes iron in the haemoglobin of the red blood cells to form methemoglobin, which lacks the oxygen – carrying ability of haemoglobin. This creates the condition known as methemoglobinemia (sometimes referred to as “blue baby syndrome”), in which blood lacks the ability to carry sufficient oxygen to the individuals body cells causing the veins and skin to appear blue [5]. Most humans over one year of age have the ability to rapidly convert methemoglobin to haemoglobin within red blood cells so that it remains low in spite of relatively high levels of nitrate/nitrite uptake. However, in infants under six months of age, the enzyme systems for reducing methemoglobin to oxyhaemoglobin are incompletely developed and methaemoglobineamia can occur.
This also may happen in older individual who have
then to nitrogen gas. The conversion of nitrate to
nitrite occurs readily under alkaline conditions. Nitrite is absorbed in the intestine into the blood stream. Concentrations of 45 mg/l produce methaemoglobinemia [5]. The maximum contamination limit for nitrite is 0.03 mg/l [3]. Sources of nitrite pollution are generally the same as those of nitrate pollution.
Sources of phosphorus include; detergents, phosphorus acids, fire works, munitions, rat poisons among others [6]. Presence of these contaminants in high levels in local alcoholic beverages could cause problem to the consumers. Phosphates are one of the most common nutrients to move through the ecosystem in large quantities. Phosphates enter the
waterways through run off from natural sources such
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as phosphate – containing rocks and from human sources such as fertilizer, pesticides, detergents, and industrial wastes.
If a starchy food grain is fermented, it produces enzymes, which start to break down the starch down into sugar. This is how growing plants derive energy; and this is how people release sugar from grains so that they can make alcohol from them, by brewing the grains into beer. During the nineteenth century, in most parts of East Africa, the most used grain was finger millet (elevisine) but some people used
sorghum. Busaa is prepared from cereals, chang’aa
a. Environment: the high concentrations of nitrate in water causes a phenomenon known as "Eutrophication", which means an excessive growth of the algae in water which consumes the oxygen gas dissolved in water causing the death of fishes in that water [14].
b. Animals: especially ruminant animals such as cows, sheep and goats [14]. When ruminants consume feed with high nitrate levels, the nitrate can be converted to nitrite, which causes both nitrate and nitrite accumulation in the rumen [14].
As a result of the accumulation of both nitrate and nitrite in animal rumen, it causes acute and chronic
symptoms which run as reduction in weigh gain,
is a distilled brew coInsumeJd in most paSrts of Kenya ER
[7]. They are made from a variety of grains - malted millet and malted maize being the most common. It has a pleasant sweet flavour and contains at least
50% alcohol [7].; miti is prepared from boiled roots and honey, while muratina is prepared from sugarcane or honey, which is fermented using sausage plant (Kigelia african). The conditions and raw materials used to prepare these home made brews/spirits may introduce toxic materials into the alcohols and hence the need for continuous monitoring of the levels of nutrients in the alcohols to make sure that the population is not exposed to dangerous levels [6].
The high concentration of nitrate has adverse effects
on environment, animals and humans:
reduction in milk production, low appetite, aborted
breathing, blue coloring of mucus membrane, rapid heartbeat, abdominal pain, vomiting, reproductive problems, abortions, and premature death of calves [14]. The acute nitrate poisoning causes death, because the nitrate is reduced to nitrite in the rumen by bacteria. Nitrite is highly toxic because it combines with hemoglobin and form methemoglobin which is enabling to carry oxygen [14]. A death occurs within few hours after the ingestion of a high nitrate feed [14].
c. Human: Nitrate itself is not toxic; however, the conversion of nitrate to nitrite in human and animal bodies is very dangerous if it accumulates in high concentrations. On the other hand, the following could be occurring as a result of the accumulation (or uptake) of large dose of nitrate for human health:
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Methemoglobenemia: drinking water and vegetables are the major sources of nitrate consumed by human stomach [14]. The toxicity of nitrate in humans is due to the body’s reduction of nitrate to nitrite. This reaction takes place in saliva of humans at all ages and in the gastrointestinal tracts of infants during the first three months of life [12]. Nitrate becomes toxic when it is reduced to nitrite, a process that can occur in the stomach as well as in the saliva. Infants are especially susceptible because their stomach juices are less acidic and therefore conducive to the growth of nitrate reducing bacteria [9].
Nitrate is reduced to nitrites which combine with
hemoglobin to form methemoglobin (metHP). The
infected by the methemoglobenemia, this acidity increases the conversion of nitrate to nitrite by providing an appropriate environment for the nitrate reducing bacteria [14]. Other groups that may be risk to form met HP are pregnant women, which may cause birth defects and miscarriages [14]. Also people who have deficiency in glucose-6- phosphate dehydrogenase or metHb reductase may be at risk to form metHP [14]. Adults with lower gastric acidity can be infected with metHP. Fatalities have been reported after single intake of 4-50 g of nitrate (equivalent to
67-833 mg of nitrate per Kg of body weight) [14]. The symptoms related to high levels of metHP in blood
include bluish coloration of skin, headache, dizziness
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best-known effect of nitrite is its ability to react with
haemoglobin (oxyHb) to form methaemoglobin
(metHb) and nitrate:
As a consequence of the formation of metHb the oxygen delivery to tissue is impaired.
Methemoglobin is a compound that cannot combine with oxygen, and that decreases the capacity of the blood to transport oxygen from lungs to body tissues causing a condition known as “Blue Baby Syndromes” or “methemoglobenemia” (Abu-Dayeh,
2006). The normal met HP level in humans is less than 3% in infants under three months of age. However hypotoxic signs may develop at about 20%, while death occurs at 50% metHP or higher [14]. Because of their high stomach acidity, infants less
than one year old (3-6 months old), are highly
difficulty in breathing, in severe cases damage to
brain and death may occur [14].
2. Carcinogenicity: nitrite reacts in stomach with nitrosatable compounds to form N-nitraso compounds [14]. These compounds have been found to be carcinogenic [14]. The US National Research Council found an association between high nitrate intake and gastric and esophageal cancer [14]. High levels of nitrate intake were also linked with the Non-Hodgkin’s lymphoma, bolder cancer, pancreatic cancer and stomach cancer [14]. High levels of nitrate were also linked with the infection with the diabetes, the occurring of some birth defects, and miscarriages [14].
The objective of this research was to;
a) determine the amounts and compare the variability in the nitrate, nitrite and
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phosphate contents in homemade brews and the raw materials used in Nairobi using UV-visible spectroscopy.
b) Evaluate the relative safety of these homemade brews based on the MCL set by the WHO and EPA standards.
The Kenya water quality regulation has adopted the
50.0, 0.03 and 2.2 mg/l standard as the maximum contamination levels (MCL) for nitrate, nitrite and phoshate for regulated public water systems [3].
2.2 Phosphate, sources and health effects Phosphorus is an essential nutrient for plants and animals in the form of ions P04 3- and HPO4 2-. It is a
part of DNA molecules, of molecules that store energy
Phosphate levels greater than 1.0 mg/l may interfere with coagulation in water treatment plants [17]. The recommended maximum for rivers and streams is 2.2 mg/l [3].The maximum contamination level for foods is 5000 mg/kg [16].Total phosphorus concentrations in wade able streams in the U.S ranged from undetectable <1 µg/l) to more than 5000 5g/l. Thresholds for davorable or unfavorable water quality vary from one part of the country to another. The toxic effects of high levels of nitrites and nitrates calls for their levels to be continuously monitored in the environment.
IJSEThis was donRe according to specification in the
(ATP and ADP) and of fats of cell membranes.
Phosphorus is also a building block of certain parts of the human and animal body such as bones and teeth. Natural sources of phosphorus are mainly derived from the weathering of phosphorus bearing rocks and the decomposition of organic matter [17]. Domestic waste water (particularly wastewater containing detergents), industrial effluents, and fertilizer run off, contribute to elevated levels in surface waters, making them major pathways of phosphorus transformation and residue decomposition [17].
In most natural surface waters, phosphorus concentrations range from 0.005 to 0.020 mg/l [17]. To control eutrophication, the EPA has made the following recommendations; total phosphorus should not exceed 0.05 mg/l as phosphorus in streams that do not discharge directly into lakes or reservoirs [17].
Eurasian Journal of Analytical Chemistry 4(2): 204-
214, 2009, Pgs 204-214 [10].
Nitrite, nitrate and phosphorus were analyzed using this method. Ultraviolet and visible (UV-Vis) absorption spectroscopy was the measurement of the attenuation of a beam of light after it passes through a sample or after reflection from a sample surface. Absorption measurements can be at a single wavelength or over an extended spectral range. Ultraviolet and visible light are energetic enough to promote outer electrons to higher energy levels, and UV-Visible spectroscopy was usually applied to molecules or in organic complexes in solution. The UV-Visible spectra have broad features that are of limited use for sample identification but are very useful for quantitative measurements (Brain, 2000).
Determination of the nitrite based on the reactions
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involving sulfanilic acid with methyl anthranilate as the coupling agents followed by reduction using Zn/NaCl and diazotization has been applied successfully to determine trace amounts of nitrite and nitrate in water and pharmaceutical preparations [20].
Orthophosphate and molybdate ions condense in acidic solution to form molybophosphoric acid (phosphomolybdic acid). Upon selective reduction with hydrazinium sulphate, a blue color is produced due to molybdenum blue of uncertain composition. The intensity of the blue colour is proportional to the amount of the heteropoly acid. If the acidity at the
time of reduction is 0.5 M in sulphuric acid and
together with general financial status of people living in those areas. Most of the occupants are known to be living below the poverty level. The sampling stations were majorly the slum areas around Nairobi for example; Gikomba, Githurai, Kibera, Kawangware, Kangemi, Mathare, Runda and Uthiru. This sampling was carried out between December 2003 and August
2004.
All chemicals for nitrite and nitrate analysis were analytical reagent grade. Doubly distilled water was used in the preparation of all solutions in the
IJSEexperiments. RWorking standard solutions were
hydrazinium sulphate is the reductant then the
resulting blue complex exhibits a maximum absorbance at 820-830 nm [21].
Sixteen (16) stations were targeted and ten samples of each brew and water were selected. A total of one hundred and thirty two (132) home-made alcoholic beverages, forty eight (48) water and eighteen (18) raw materials samples were analyzed for nitrate, nitrite and phosphorus. Six different raw materials were selected. Three samples of each were obtained from various places in the sixteen stations. These samples were randomly obtained from various parts of Nairobi and outskirts taking into account the requirements for the preparation of the brews. These stations were chosen on the basis of the home made
alcohol brewing and drinking activities on those areas
prepared by appropriate dilution. Sulfanilic acid (0.5 g in 100ml water) and methyl anthranilate (0.5 ml in 100 ml of alcohol) were used. The following reagents were prepared by dissolving appropriate amountsin water 2
M of HCl and 2 M NaOH [21].
Nitrite stock solution (1000 µg/l) was prepared by dissolving 0.1500g sodium nitrite in water and diluting to 100 ml. Nitrate stock solution (1000 µg/l) was prepared by dissolving 0.7220 g potassium nitrate in water and diluting to 100 ml.
The following reagents were also used for the determination of phosphate ions
(i) 12.5 g of analytical reagent sodium molybdate
(Na2 MoO4 .2H2 O)
(ii) Hydrazinium sulphate solution
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(iii) 1.5 g of analytical reagent hydrazinium sulphate was dissolved in deionized water and diluted to
1000 ml [21]
(iv) Standard phosphate solution (10 ppm P)
(v) 0.04393 g of analytical reagent potassium dihydrogen phosphate was dissolved in de ionized water and was diluted to 1000 ml
(I ml of solution = 0.01 mg P). This solution is to be prepared fresh on monthly basis [21].
(vi) Calibration Solution
The phosphate stock solution containing 10 mg PO4/l was used to prepare standard solutions of between 0-
1 mg PO4 /l.
samples, SDS-270 auto-sampler and the auto-sipper flow through cell can be used [21].
Cleaning of apparatus was adopted from Mendham et al, (2002), and AOAC (2000) [28]. Research apparatus as recommended by Association of Official Analytical Chemists (AOAC) were used. Sampler check blanks were generated in the laboratory or of the equipment cleaning contractor’s facility by processing reagent water through the sampling devices using the same procedure sampling i.e. bottles were cleaned with liquid detergent and thoroughly rinsed with reagent water. The bottles
were then immersed in a hot (50- 60 °C) bath of 1 N
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All the weighing were done using a research
analytical balance (Sartorious research, R 200D, model-40110044, Analos, Belgium). Other apparatus included the following; graduated pipettes (10 and 5 ml), micropipettes (200 ml) and tips, test tubes (13 x
100 ml), small square of parafilm, volumetric flasks
(50 and 100 ml) and computer.
Nitrite and nitrate were analyzed using, a SHADZU (Model No. UV-2550) UV-Visible spectrophotometer with 1 cm matching quartz cell were used for the absorbance measurements. A WTW pH 330 pH meter was used [20].
Phosphate was analyzed using, the UV-visible spectrophotometer (Philips, Scientific equipment, UP-
8700 Series). For the automated analysis of up to 270
trace metal grade HCl for at least 48 hours. The
bottle were then thoroughly rinsed with reagent water and filled with 0.1 % (v/v) ultra pure HCl and double- bagged in new polyethylene zip-type bags until needed [22]. The apparatus were cleansed using detergents, water, rinsed with distilled deionized water and dried overnight in the oven at 100 °C.
A 100 ml samples were collected directly into specially cleaned, pretested, polypropylene bottles using sample handling techniques specially designed for collection of sample for the analysis of metals at trace levels. The samples were then either laboratory preserved by the addition of 5 ml of pretested 10 % HNO3 per litre of sample, depending on the time between sample collection and arrival at the
laboratory.
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The brew sample bottle (acid-washed, 125 ml polyethene bottle) were rinsed 3 times before sampling. Filled to approximately 2/3 full, tighten cap and freeze cruise, cast Niskin bottle number were recorded on the bottle and data sheet. All the brew sample bottles were first rinsed with the alcohol for alcohol samples before the brew samples were collected. The samples were then filtered, the residue discarded and the filtrates from home made brews were decolorized using activated charcoal and re- filtered until the colour disappeared.
In the sample pretreatment, modified procedures for
In the phosphate analysis all samples of the brews, digested raw materials and standards (Including quality control solutions) were processed in the same manner. The pH of a well mixed sample was adjusted from 6.0 to 8.0 using 6 M NaOH and 1M HCL. 10 ml of ammonium persulphate ((NH2 )2 S2 O8 ) solution and
0.25 ml of 5.4 M H2 SO4 were added to each tube and
mixed. The caps were initially tightened, inverted a few times to ensure good mixing, and then the caps were unscrewed until the seals had just become
loose. The tubes in the racks were placed in either
IJSEautoclave or pRressure cooker. For the autoclave, the
washing and drying proposed by Santos et al. (2004)
and Kawashima & Soares (2003) [23]., respectively, were used. First, each raw material samples were rinsed with distilled water to remove dirt and other debris. Then the raw material samples were brushed with polypropylene bristles and washed with deionized water. The raw materials were then grated with a polypropylene grater into porcelain containers. Then the containers with the raw material samples were dried in a laboratory oven at 65 ± 5 ºC for 24 h or until reaching constant weight. Immediately afterwards, the samples were stocked in polypropylene beakers and covered with a PVC film. Finally, they were stored in a desiccators awaiting digestion [23].
manufacturer’s directions were used and the mixture was heated at 120 °C for 30 minutes. The tubes were finally removed from the autoclave or pressure cooker and cooled between 20-30 °C. A volume of 0.40 ml of
6 M NaOH was added followed by 1 drop of phenolphthalein solution to each sample and mixture. Thereafter 6 M NaOH was added into the solution until the solution turned pink and then 5.4 M H2 SO4 until the pink color just cleared.
One (1) gm of the raw materials was weighed and digested using 6 ml of concentrated HNO3, 0.5 ml of concentrated Hydrochloric acid and 1 ml of H2 O2 were added to each one in Teflon vessel in order to dissolve the organic matter. These were placed in the microwave and digested for 30 minutes. A multiwave
300 microwave oven (Perkin-Elmer, Shelton, CT
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USA) was used for the microwave-assisted digestion
[18].
Step | Power | Ramp (min) | Hold (min) | Fan speed |
1 | 750 | 10 | 10 | 1 |
2 | 1200 | 10 | 10 | 1 |
3 | 0 (cool Down) | 0 | 15 | 3 |
To ensure a safe digestion, the multi wave 3000’s IR sensor measures the temperature of each vessel. If a vessel nears its maximum operating temperature of
260 °C then the microwave oven automatically
decreases the applied power. Also, the pressure
Samples were analyzed using Hydride generation atomic absorption spectroscopy and UV-Visible spectroscopy. The maximum holding time for NO3 -N, NO2 -N and PO4 -P was 48 hours. The concentration of the nutrients in solution was determined by measuring the absorbance. Nitrite and nitrate were analyzed at 493 nm and phosphate at 830 nm respectively, and then applying the Beer-Lambert law the concentrations of the solutions were obtained.
A volume 50 ml of alcohol samples and blanks were shaken and filtered sample were put into 125 ml
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sensor sends data to the multi wave oven controller
during the digestion. The multi wave oven automatically reduced power when the maximum pressure of 60 bars was applied [18].
No digestion is performed on unfiltered samples prior to analytical determinations. Portions of 20 ml of the
Erlenmeyer and 1 ml of 40 % concentrated sulphuric acid was added to the mixture followed by 600 ml of distilled water, cooled and diluted to 1 liter. This was evaporated on a hot plate for 30 minutes, cooled, diluted to 50 ml with distilled water and 8 ml of the reducing agent [prepared by mixing in the following
order, 50 ml of 2.5 M H SO , 5 ml of potassium
2 4
neutralized filtered brew samples were evaporated to dryness in separate beakers. The residues were each cooled and extracted with 1 ml phenol disulphonic acid {prepared from 25 g of phenol crystals (BDH Chemicals Ltd, Poole, UK), 150 ml of concentrated H2 SO4 (Fischer Chemicals, UK), 75ml of fuming H2 SO4 (Fischer Chemicals, UK)} and each mixture heated for 2 hours on water bath. All samples (homemade brews, water, raw materials) and blanks (n=3) were digested and diluted using the same
procedure.
antimonyl tartarate (BDA chemicals Ltd, Poole, England) solution, 15 ml ammonium molybdate (Aldrich Chemical Co. Inc., USA) solution and 30 ml of ascorbic acid (Fisher Chemicals, UK) solution] added the solution thoroughly mixed. The standards and the blanks were treated in the same manner. After 30 minutes the absorbances of the test solutions and the standards were measured at 830 nm against the reagents blank as reference using a UV – visible spectrophotometer (Philips, Scientific equipment, UP
– 8700 series).
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Aliquots of stock solution containing 0.2-8.0 µg /l of nitrite were transferred into series of 10 ml calibrated flask. To each flask, 1 ml of 0.5 % sulfanilic acid and 1 ml of 2 mol/l hydrochloric acid solution were added and the solutions were shaken thoroughly for 5 minutes to allow the diazotization reaction to go to completion. Then, 1 ml of 0.5 % methyle anthranilate and 2 ml of 2 M sodium hydroxide solution were added to form an azo dye and the contents were diluted to 10 ml using water. After dilution to 10 ml with water, absorbance of the red colored dye was
measured at 493 nm against the corresponding
added to form an azo dye and the contents were diluted to 10 ml with water. After dilution to 10 ml with water, the absorbance of the red colored dye was measured at 493 nm against the corresponding reagent blank [20].
The levels of nitrate-N in home made brews/spirits and water was determined using UV-visible spectroscopy and the result obtained for various stations are presented in Table 2 and Figure 1.
From the Table 2, the average levels of nitrates were generally high in homemade brews/spirits.
The mean nitrate levels in the brews/spirits were
reagent blank and thIe calibJeration curveSconstructed ER
[20].
In the analysis of nitrate 10 ml sample was pippeted out of the stock solution into a beaker, followed by 5 ml of HCl and 2 ml of Zn/NaCl granular mixture added. This was allowed to stand for 30 minutes with occassional stirring to form a nitrite. The final mixture was filtered into a 100 ml standard flask using what man No. 41 filter pap and diluted up to the mark. Aliquots of stock solution containing 0.26-10.7 µg/l of reduced nitrate were transferred in to series of 10 ml standard flask. 1 ml of 0.5 % sulfanilic acid and 1 ml of 2 mol/l HCl solutions were added, shaken thoroughly for 5 minutes for the diazotization reaction to go to completion. Followed by, 1 ml of 0.5 % methyl
anthranilate and 2 ml of 2 M NaOH solution were
generally lower than the recommended levels of 50
mg/l for water [3]. Nitrite and Phosphate levels were generally higher than the MCL levels of 0.03 and 2.2 mg/l respectively. Kenya Bureau of Standards does not have standards foe nitrates in alcoholic beverages. The mean concentration of nitrate in the home made brews and spirits were calculated and the results were used to plot a graph of concentration against home made brew/spirit as shown in the Figure
1. Karubu had the highest mean concentration of
NO 3 -N at 40.90 ± 1.05 mg/l, followed by Busaa at
39.6 ±1.27 mg/l and Muranina had the lowest mean level at 33.71 ± 11.81 mg/l.
Kangara had the highest mean concentration of nitrite NO2 -N at 6.85 ± 0.40 mg/l, followed by Muratina at NO2 -N at 6.18 ± 5.35 mg/l and Chang’aa had the
lowest mean level at NO2 -N at 0.12 ± 0.22 mg/l.
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Karubu was not analyzed. Busaa had the highest mean concentration of PO4 -P at 3.28 ± 0.37 mg/l, followed by Chang’aa at 3.04 ±0.10 mg/l and Kumikumi had the lowest mean level at 0.13 ± 0.01 mg/l.
Some brews such as Chang’aa, Busaa, Kumi kumi, Miti and Muratina are normally prepared by the river bank to facilitate cooling. Waters from these rivers are also sometimes used in brewing process. Since they are generally polluted with industrial and domestic wastes, the nutrients end up in the brews. The other source of nutrients in the home made brews may
have been due to the use of untreated waters used in
significant differences between the concentrations of nitrates in the brews means as determined by one- way ANOVA (F = 3.098, p = 0.019). All at considered at 95 % confidence interval. In all the homemade brews the value of p < 0.05, implying that there were significant differences in the levels of nitrite in the homemade alcoholic beverages. An ANOVA two way test was also done for nitate and nitrite. Since the calculated Fstatistics (1,5) =383.003, was greater than Fcritical ,= 16.258 we know that there was a statistically significant difference between the nitrate and nitrite concentrations, p < 0.01. Thus, the null hypothesis
was rejected.
the slum areas.
IJSEThe followingRANOVA one way test results were
From ANOVA one way test on nitrate levels in the
homemade brews; the probability of the between- treatments MS being ≥ 0.783 times the within- treatments MS, if the null hypothesis is true, is p = 0.570. The mean was 36.8343, the median 38.3 and the SD was 6.447. We can therefore conclude that there were no statistically significant differences between the concentrations of nitrates in the brews means as determined by one-way ANOVA (F = 0.783, p = 0.570).
The ANOVA one way test on nitrite in the raw materials used gave the following results; the probability of the between-treatments MS being ≥
3.098 times the within-treatments MS. The mean was
4.117, the median 1.41 and the SD was 4.516. We can therefore conclude that statistically there were
found for Phosphate in homemade brews; the probability of the between-treatments MS being ≥
5.122 times the within-treatments MS, if the null hypothesis is true, is p = 0.000. The P-value of 0.000 is less than the significance level (0.01), so we can reject the null hypothesis and safely assume that phosphate concentration affects the health of the users. The mean was 2.945, the median 3.11 and the SD was 0.9494. We can therefore conclude that there were no statistically significant differences between the concentrations of phosphates in the brews means as determined by one-way ANOVA (F = 91.251, p =
0.000). F (91.251) is greater than F crit (2.95), so again, we can reject the null hypothesis. All levels were considered at 95 % confidence interval. In all the homemade brews the value of p < 0.000, implying
that there were significant differences in the levels of
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phosphate in the homemade alcoholic beverages. From these values we can conclude that the levels were also lower than the maximum contamination levels of 2.2 mg/l.
The concentration of nutrients were generally above the maximum allowable limit set by KEBS and WHO [3]. Kenya Bureau of Standards did not have standards for nitrites in alcoholic beverages. Hence they pose health danger to the consumer. Their source could be from the use of nitrogenous fertilizers used for growing raw materials or organic decomposition during the fermentation process, use
of contaminated river water and some additives in
allowable limits of 5 mg/kg of nitrate set by the World Health Organization [11]. An Acceptable Daily Intake (ADI) for nitrate of 3.7 mg/kg b.w./day, equivalent to
222 mg nitrate per day for a 60 kg adult was established by the former Scientific Committee on Food (SCF) and was reconfirmed by the Joint FAO/WHO Expert Committee on Food Additives (JECFA) in 2002. The CONTAM Panel noted that no new data were identified that would require a revision of the ADI [1].
An ANOVA one way test was done on nitrate levels in the homemade brews gave the following results; the
mean was 276.233, the median 276.85 and the SD
some cases.
IJSEwas 37.104. RA P-value of 0.000 is less than the
The levels of nitrates in the raw materials used to make the brews were determined using UV-Visible spectroscopy and the results are represented in Table
3.
Presented here in Table 3 and Figure 2, the mean nutrient levels were generally high in the raw materials used. Nitrate had values ranging from
215.50 ± 33.00 to 326.20 ± 75.00 mg/kg. Millet had the highest concentration at 326.20 ± 75.00 mg/kg followed by sorghum at 298.50 ± 27.50 mg/kg, and Maize had the lowest mean concentration of 215.50 at 281.30 ± 10.44 mg/kg, Maize had the lowest at
215.50 ± 18.31 mg/kg. The levels of nitrate in all
materials were found to be well above the maximum
significance level (0.01) and F (14.535) is greater
than F critical (14.47103), hence the concentrations of the raw materials used did not affect significantly the concentrations of nitrate.
The ANOVA one way test done on the mean levels of phosphate in the homemade raw materials used gave the following results; the mean was 276.233, the median 276.85 and the SD was 37.104. A P-value of
0.000 is less than the significance level (0.01) and F (279.083) is greater than F critical (3.197), hence the concentrations of the raw materials used did not affect significantly the concentrations of nitrate. From this we can deduce that there were significant differences between the phosphates levels in all the raw materials used.
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The ANOVA two way test was also done on nitrate and nitrite. The Fstatistics (1,5) =39.665, was greater than Fcritical ,= 16.258 we know that there was a statistically significant difference between the nitrate and nitrite concentrations, p < 0.01. Thus, the null hypothesis can be rejected.
From table 3, honey had the highest concentration of nitrite ions of 193.00 ± 18.52 mg/kg, followed by sorghum at 180.00 ± 16.02 mg/kg and Millet seeds had the lowest concentration of nitrite at 116.00 ±
41.93 mg/kg. All these means were higher than those observed in the brews, indicating that the raw
materials could be their source. The levels of nitrites
The mean concentrations of phosphate-P in the raw materials were found to be generally low in fact below the maximum contamination level of 5000 mg/kg (Codex, 2001). Concentration ranged from 104 ± 6.08 to 285 ± 23.80 mg/kg.
Jaggery had the highest concentration of 285.00 ±
23.80 mg/kg, followed by sorghum at 190.00 ± 6.45 mg/kg and maize seed had the lowest level of 104.00
± 6.08 mg/kg. These levels are low and therefore do not pose risk to the health of the consumers. This indicates that phosphate pollution is low in the raw material analyzed hence the raw materials could not have contributed in elevating the levels of this nutrient
in the brews and spirits.
in the raw materials Iwere foJund to be higSher than the ER
maximum allowable limits set by the WHO of 2 mg/kg [8]. The high concentration must be due to the use of nitrogenous fertilizers together with the reactions that take place during the fermentation process.
The same test was repeated for nitrite levels in the raw materials and the following results were obtained; the probability of the between-treatments MS being ≥
3.609 times the within-treatments MS. The mean was
148.2, the median 137.5 and the SD was 31.42. We can therefore conclude that statistically there were significant differences between the concentrations of nitrates in the raw materials used means as determined by one-way ANOVA (F = 3.609, p =
0.032). All were considered at 95 % confidence interval. In all the raw materials the value of p < 0.05, implying that there were significant differences in the
levels of nitrite in the raw materials used.
This implies that the raw material whose nutrients
concentrations were generally high for nitrate, nitrite and phosphate contributed positively towards elevating the levels of these nutrients in the brews/spirits. Hence, it is also possible that slum dwellers may be using polluted river waters in brewing.
.
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Millet | 326.20 ± 14.93 | 116 ± 41.93 | 185.00 ± 5.16 |
Sorghum | 298.50 ± 10.44 | 180 ± 16.02 | 190.00 ± 6.45 |
Honey | 263.50 ± 23.34 | 193 ± 18.52 | 118.00 ± 18.17 |
Jaggery | 281.30 ± 14.99 | 145 ± 47.79 | 285.00 ± 23.80 |
Muratina fruit | 272.40 ± 16.38 | 125 ± 14.90 | 165.00 ± 24.68 |
The nitrate-N levels were generally below the maximum contamination levels of 50 mg/l set by the
WHO. In the raw materials the levels were relatively
high. 105 out of 180 samples analyzed for nitrite had levels above the maximum contamination levels of
0.03 mg/l level for water. Phosphate-P were also found to be high in the homemade brews, 102 out of
180 samples analyzed had levels above the maximum contamination levels of 2.2 mg/l for water.
The levels of nitrate levels in raw materials used in the preparation of the homemade alcoholic beverages
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were generally higher than the MCL of 5 mg/kg and the LOD of 8 mg/kg; Nitrite levels in the raw material were higher than the MCL of 2 mg/kg and the LOD of
0.6 mg/kg, but lower than the MCL of 5 000 mg/kg for phosphates.
These levels of nitrate, nitrite and phosphate in the sample areas exceeded the maximum levels recommended by the World Health Organization [11]. Nutrient levels observed in the home made alcoholic beverages most likely reflect the levels of these nutrients in the raw materials used in the brewing processes and the fertilizer used in farming.
Lastly, I thank the Teachers Service Commission for granting me study leave with pay, my lecturers, Dr. Ruth Wanjau, Professor Gerald Muthakia, Dean (SPAS), Dr. Richard Musau, (Chairman, Chemistry Department), Prof.Hudson Nyambaka and Dr. Koga (Academic registrar) all of Kenyatta University, for their support.
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The author wishes to express his sincere gratitude to the Chief chemist, Government chemist and the chief engineer, engineer Maina, ministry of public works, and materials branch for support during the entire research period when there was inadequate funding. The good co-operation of the deputy government chemist, Mrs. Okado and the Head of Departments in both foods and water and the whole government chemist staff is highly acknowledged.
Special thanks go to Professor Jane Murungi and Dr. Charles Onindo of Kenyatta University for their supervision of the thesis and helpful critical comments that resulted in the presentation of the data obtained. Thank you also for your gifted editing, your hard work and your patience. I greatly appreciate the typing
assistance of my loving wife Mrs. Rosalia Masime.
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