Physico – Chemical Characterization of OMWW City Guisser (Region of Settat Morocco) and Monitoring its Color During the (Oxidation by H2O2 Natural Evaporation) and the Study of Its Biological Activity Against the Aspergillus Niger

DOI : 10.17577/IJERTV3IS070776

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Physico – Chemical Characterization of OMWW City Guisser (Region of Settat Morocco) and Monitoring its Color During the (Oxidation by H2O2 Natural Evaporation) and the Study of Its Biological Activity Against the Aspergillus Niger

E. Ouabou1

1: Lab Environment and Applied Chemistry,

Faculty of Science and Technology Settat, Morocco.


1: Lab Environment and Applied Chemistry, Faculty of Science and Technology Settat, Morocco.

S. Hilali2

2: Research Unit Agro-Resources and Environment Moroccan.

Faculty of Science and Technology. Settat, Morocco

Abstract The extraction process of olive oil production generates liquid effluents, or sometimes appointed OMWW or vegetation water. Pressing 1 ton of olives produced on average

    1. tonnes of OMWW with modern modes of production. The variation depends on the extraction process: pre-washing olives or not, humidification pulp during pressing [UNCTAD] The objective of this work is devoted to the first step in the study of the impact of the OMWW (city Guisser) region of Settat Morocco on the environment by studying its physicochemical characteristics and study its biological activity against the Aspergillus Niger, in the second step the monitoring of its natural color during the evaporation and oxidation by


      Keywords OMWW, physico-chemical composition, coloring, biological activity, oxidation.


      The discharge of effluent from industries producing olive oil is a major problem especially in Mediterranean countries. These highly polluted water cause serious environmental damage. The lack of treatment methods adapted push oil mill owners to reject these waters in nature without any control or overload with these toxic substances a sewer system inadequate.

      The vegetable water are poorly degradable because of phytotoxic and antimicrobial substances (phenols, volatile fatty acids, insecticides, etc.) they contain (F DAGGA ABU BÖHMER B). The OMWW are often discharged into natural receivers, rivers without any treatment. The very high organic load prevents these waters to self-purify and pollution can extend over very long distances (MEBIROUK M., 2002). The chemical composition of vegetable water is quite variable. It depends on many factors, particularly the method

      of extraction of the oil, but also of the production period.

      Table 1: General chemical composition of vegetable


      content [%]


      83 88 %

      organic matter

      10.5 15 %


      1.5 2 %

      crude protein

      1.25 2.4 %


      0.03 1 %


      1.0 1.5 %

      Sources : SANSOUCY R. 1984 et MEBIROUK M., 2002

      The scientific work is cited more than 20 processes for the treatment of effluents from olive oil mills technologies. This is in most cases of elementary or combined tested in laboratory scale or in a pilot plant without any real industrial projection. The following techniques have been described as the most used or as potentially applicable. These processing techniques can be classified according to the method used:

      • Thermal: Evaporation (El Alami, 2000 Nucleos Interface, 1992), incineration (Fiestas Ros Ursenos, 1983) or distillation (Ranalli, 1991)

      • Physico-chemical: filtration / ultrafiltration, oxidation, ozonation and coagulation,

      • Biological: aerobic and anaerobic treatment (Hamdi et al 1991; Borja et al 1994 Elalami, 2000 El Hajjouji et al 2007….)

        As part of this work, the focus is on three areas:

        -1-The physico-chemistry of the OMWW Guisser,

        -2- Monitoring its color during the oxidation by H2O2 and natural evaporation.

        -3- The study of the biological activity of the vegetable water against Aspergillus Niger.


        1. – Sampling: The OMWW. Samples of vegetable water were made at a traditional unit of crushing in the region of Settat town Guisser during the olive crop year 2009-2010. The samples were taken from the storage tank and then transported in drums of 20 liters.

          Extract medium potato dextrose. Ingredients: 200g potatoes diced 20g dextrose or plain white sugar cane, 1 liter of water. Aspergillus Niger spores

        2. – Experimental:

The pH was measured using a pH meter. The total dry matter was determined after drying at 105 ° C in an oven.

Volatile material was derived as the difference between the obtained by evaporation at 105 ° C dry matter and ash residue from the calcination at 550 ° C for 2 hours. It is expressed in g / l based on the dry weight.

Phenolic compounds were determined by the colorimetric method using the Folin-Ciocalteu reagent. The results are expressed in equivalent of p-hydroxybenzoic acid (Macheix et al 1990).

The electrical conductivity was measured using a conductivity meter (Rodier, 1984). The chemical oxygen demand (COD) was determined according to the standard method (APHA 1992) by oxidation of the organic matter contained in the sample to 150 ° C by an excess of potassium bichromate in acid medium and in the presence of sulfate money. Excess potassium dichromate was determined by colorimetry at 620 nm. The chemical oxygen demand (COD) was determined by the colorimetric method

dichromate closed reflux.

Suspended solids (TSS) are determined by centrifugation of a 20 ml sample at 3000 g for 28 minutes. The pellet was placed in a porcelain cup previously weighed and then dried in an oven at 105 ° C for 24 hours. The difference between the weight of the dried sample and that of the cup determines the rate of MES. It is expressed in g / l (Rodier, 1984).

Chlorides were determined according to the standard (AFNOR T90-014), by the method of titration of Mohr.

The inorganic material was determined after digestion at 550 ° C in an oven for 24 hours.

The total sugars were determined by the phenol sulfuric calorimetric method in the presence of a standard glucose (Dubois et al 1956) range.

Nitrites, nitrates, ammonium ions, phosphates, calcium and magnesium are calculated by colorimetric assay.

The dissolved oxygen is measured in the sample of vegetable with an oximeter.

Sterilization is a technique for eliminating any microbial organism of a preparation; in this case, the method is used in the processing steam (autoclaving). The percentage of fading is deducted from the concentrations of the mixtures before and after the addition of


%décoloration C0 Ci 100


C0: concentration before adding H2O2 Ci: concentration after the addition of H2O2

So according to the Beer-Lambert law:

A l C

With: A: absorbance or optical density l: Molar extinction coefficient in L mol-1 cm-1 : The thickness of the vessel used (in cm).

The% bleaching is:

%décoloration A0 Ai 100



  1. – physicochemical characterization of vegetable water Guisser

    Table 2: organic characteristics of the vegetable Guisser

    Acidity (pH)


    MES (g/l)


    MS (g/l)


    MV (g/L)


    Ttal sugar (g/l)


    COD (g/l)


    Fat (g/l)


    Phenolics p-hydroxybenzoic acid (mg / l))

    Before extraction


    After extraction


    The concentrated


    The vegetable water are acidic effluent, because of the presence of organic acids (phenolic acids, fatty acids, …). The value recorded in our study is of the order of 5.72 is almost the upper limit of the range reported in the literature (4.5 to 6) In addition, the acidity of the OMWW increases with the duration of their storage station in the natural evaporation due to polymerization reactions which convert the phenolic alcohols to phenolic acids (Hamdy 1991).

    Vegetable waters are rich in suspended solids, their content is about 10g / l, this value is lower than the OMWW Fes 18g / l (Melle Halah aissam 2002). Although in the basin, TSS of vegetable down due to settling. Register value in our study is normal.

    These discharges are also characterized by the volatile matter of the order of 104g / l.

    This percentage is similar to that observed by several authors (Hamdi, 1991a; Garcia Garcia et al, 2000. Assas et al, 2002; Fountoulakis et al, 2002. Fadil et al, 2003.). But similar to that observed by (Ms. Halah aissam 2002) 86 gl-. The average content of MOWW dry matter of about 133g

    / l, it is a higher value than observed by (Halah aissam 2002) 98g / l. This shows that the vegetable water of Guisser is rich

    in organic matter.

    Teneure low in sugars (0.19g / l) shows that the rejection of vegetable microorganisms use these sugars for the


    According to the IR spectrum corresponds to the bands

    degradation of phenolic compounds as an energy source. This concentration of sugars obtained in our study remains well below those reported by other authors (Sayadi and Ellouz, 1993 Martin et al, 1994. Yesilada and Kahraman, 1999).

    (0.28 gl-1) is the value observed by (Halah aissam 2002). This could be explained by the storage of vegetable in the basins, since the presence of sugars indicates the freshness of the vegetable. During storage, the sugars are fermented into organic acids (Hamdy 1992).

    The chemical oxygen demand (COD) is the amount of oxygen consumed by existing in the vegetable materials for their oxidation. The average organic matter content expressed as COD is about 15g / l (The method used is that described by Rodier (1984)) and 14.99 g / l (UV spectroscopic assay). This value is low compared to that observed by (Halah aissam 2002) 154 g of O2. L-1).

    We can say that the shelf life of vegetable water is sufficient for microorganisms degrade biodegradable organic compounds, which explains the low COD value. The fat is a naturally occurring component in many foods and is an essential part of our diet. Oils and fats are also called


    The residual amount of fat present in the vegetation water depends on the olive oil extraction system. The vegetable waters studied in our case have become viscous due to the presence of the oily fraction which is 4g / l. It forms a lipid layer on the surface of vegetable water at basin level, which could limit the natural evaporation. The fraction of vegetable lipid phase of Guisser is large compared to that of basins Fes (1g / l) (Halah aissam 2002), because the process of centrifugation provides low rate compared to the traditional process (Zimbalatti, 1995). These effluents are also characterized by phenolic compounds (which justified the IR spectrum shown below), but a small amount compared to the results listed in the bibliography (Borja et al, 1992. Kissi et al, 2001.;

    Fountoulakis et al. 2002). These toxic compounds give the vegetable OMWW a antimicrobial character.

    Figure 1: Infrared spectrum of vegetable study

    • 1600 cm-1: C-C aromatic

    • 1200 cm-1: O-H phenol

      – 3450-3550cm-1: O-H alcohol

    • 3200-3400cm-1: O-H (polymeric association).

    Spectra shows the existence of phenolic compounds in the vegetable water.

    Table 3: Mineral characteristics of the vegetable water

    Temperature (oC)


    The dissolved oxygen (mg / l)


    Conductivity (ms / cm)




    Moisture (%)


    Chloride (g / l)


    Nitrites (mg / l)


    Nitrates (g / l)


    Ammonium ion (g / l)


    Phosphate (g / l)


    Calcium (g / l)


    Magnesium (g / l)


    The average chloride concentration in the vegetable water is high (7.1 gl-1). It is due to the addition of the salt in a large quantity for the storage of olives. This item has adverse effects on crops and groundwater.

    The teneures of OMWW magnesium and calcium are respectively 3.2 g / l and 4.52 g / l; It is this wealth of minerals that has led many researchers to guide the treatment of vegetable water to their valuation compost or fertilizer for agricultural land. Ille also contains other elements according to several studies such as potassium, sodium…. The electrical conductivity reflects the ability of an aqueous solution to conduct an electric current. This notion is inversely proportional to the electrical resistivity. The conductivity is directly proportional to the amount of solids (minerals) dissolved in water. The higher the concentration of dissolved

    solids, the greater the conductivity high.

    The vegetable is rich in minerals (calcium, magnesium, potassium, sodium ….) which makes its high conductivity, in our case it is about 10.1 ms / cm is 1/3 of the observed value by (Ms. Halah aissam 2002). Conservation of vegetable with good conditions mills use commercial salt, which gives OMWW a high electrical conductivity.

    The olive oil mill effluents are compounds in addition to inorganic and organic compounds of a large fraction of water added during the extraction of olive oil and the olive vegetation. The moisture content of the vegetable water of Guisser is 84.58%, this value belongs to the range observed (Benyahia and all 2003.) (83-88%). Nitrites are obtained by converting the ammonia in the presence of oxygen by bacteria Nitrosomonas. The application of vegetable, rich in nitrogenous elements, still pollutes the water bodies by nitrites that are toxic to fish.


    The content of vegetable water Guisser of nitrite is of the order of 35mg / l. In many countries, water intended for human consumption must comply with the limit values (eg 50 mg / l in France and Europe) to qualify as drinking. WHO recommends not to exceed 25 mg / l. Guisser of the vegetable water contains about 2.74g / L which makes them dangerous discharge for underground water. The results showed that the concentration of vegetable water Guisser of ammonium ion is low (0.066g / l), but this concentration is comparable with the value obtained by (Halah aissam 2002) 0.01g / l. Phosphates are used in agriculture as fertilizer as phosphorus source The OMWW also has a high teneure phosphate ions (PO 3-) (0.09 g / l), this content is almost similar to that of Fes 0.12 g / l (Halah aissam 2002). The dissolved oxygen concentration in a tank varies depending on the temperature of the effluent, elevation, depth of the pool, the time of day, the concentration of organic matter and nutrients basin and the amount of aquatic plants, algae and bacteria in the lake. Guisser of the vegetable water has a dissolved oxygen in the range of rates 2.48mg / l, this

    value is poor by the standards of discharges in Morocco.

  2. – Change the color of the MOWW for natural evaporation and oxidation H2H2

    1. The Natural evaporation

      During natural evaporation, the vegetable water coloring changes to reddish brown to black due to increase of the concentration of phenolic compounds responsible for the color of the OMWW. The following curve shows the evolution of the color and the mass of the vegetable in the natural evaporation of a sample of vegetable water Guisser of mass exposed to sunlight. The coloration of the vegetable is proportional to the concentration of phenolic compounds responsible for the

      color of the vegetable.

      Figure 2: Evolution of the color of the vegetable runs to natural evaporation

      Staining of OMWW its changes during the fifteen days of the reddish brown to black as shown by the curve above, four different areas from each other by color:

      Zone 1: the vegetable to a reddish brown coloration Zone 2: the vegetable has a brown color

      Zone 3: the vegetable to a blackish brown color Zone 4: the vegetable to a black color

      After the vegetable zone 4 does not change color or mass, it becomes a highly viscous paste form resin as several industrial applications (naturally tanned skin, natural pesticide, binder for pelletizing mineral powders, natural dye for the treatment of wood, … (Kitane S. et al.).

    2. oxidation by H2O2

      The UV spectrum of vegetable shows that the vegetable absorbs more at 340nm.

      Figure 3: UV spectrum of vegetable

      The addition of H2O2 allows oxidize organic matter exist in the vegetable water as hydrogen peroxide 30% is too effective. She oxide not only secondary alcohols to ketones, but the oxidation goes further and form ketone peroxides.

      The oxidation of phenolic compounds that are responsible for the color of the vegetable leads to discoloration of the OMWW.

      50% of fading is obtained after adding 12ml of H2O2 (30%), which shows the following figure.

      Fig 4: Percentage of discoloration of vegetable based volume H2O2

  3. – Inhibition of Aspergillus Niger

On increasing the concentration of the vegetable water added to the culture medium, to augment the inhibition to inhibition total.

In our case the vegetable is low in phenolic compounds, which explains the growth of fungus Aspergillus Niger in the first three areas. By increasing the volume of the vegetable water in the medium, this leads to increasing the concentration of polyphenols, thus inhibiting increased.

The development of Aspergillus Niger is almost zero to the culture medium N ° 4.

Table 4: inhibition of Aspergillus Niger by the vegetable





200ml M.C + 400µl

spores dAspergillus Niger + 200µl de la margine


200ml M.C + 400µl

spores dAspergillus Niger + 400µl de la margine


200ml M.C + 400µl

spores dAspergillus Niger + 600µl de la margine


200ml M.C + 400µl

spores dAspergillus Niger + 800µl de la margine


200ml M.C + 400µl

spores dAspergillus Niger

The vegetable water therefore may play an important role due to phenolic compounds which contains, it can replace the pesticides (fungicides) and chemical impact of the curb its compounds on human health.


The OMWW Guisser exhibits physicochemical characteristics that are comparable with the characteristics of other OMWW Fes … except COD and phenolics What very

low values.

This can be explained by the action of microorganisms to the decomposition and degradation of phenolic compounds which are readily biodegradable in the antioxidants of vegetable water storage for a sufficient time so that the COD and polyphenols have a fall important.

We can conclude that the vegetable to antifungal activity against Aspergillus Niger thanks to its chemical composition, in particular phenolic compounds.


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