inhibition effect of barley and oat aqueous extract on the crystal...

Journal of Pharmacy Research Vol.8 Issue 8. August 2014

Refaat A. Saber / Journal of Pharmacy Research 2014,8(8),1162-1170

1162-1170

Research ArticleISSN: 0974-6943

Available online throughhttp://jprsolutions.info

*Corresponding author.Refaat A. SaberSoil and Water Department, Faculty of Technology and Development,Zagazig University, Zagazig. Egypt .

Inhibition Effect of Barley and Oat Aqueous Extract on the crystal growth rate of Calcium Oxalate Monohydrate Crystals

Refaat A. SaberSoil and Water Department, Faculty of Technology and Development, Zagazig University, Zagazig. Egypt .

Received on:15-06-2014; Revised on: 22-07-2014; Accepted on:11-08-2014

ABSTRACTThe inhibiting effect of Barley (Ba) and Oat (Oa), aqueous extract on the precipitation rate of calcium oxalate crystals has been studied usingconstant composition technique in which the super-saturation and ionic strength were maintained constant at 37oC. The presence of aqueousextracts of Barley(Ba) and Oat (Oa), even at relatively low concentration 10-6 M, markedly reduced the rates of precipitation. The suggestionof predominately surface Controlled mechanism is also supported by the observed low value of the activation energy Ea = 4.0 kcal/mol. Theeffective order of crystal growth reaction is determined in absence and presence of aqueous extract of additive where parabolic rate low withn – 2 The action of additives can be interpreted in terms of adsorption, following the Langmuir isotherm, of additives at the active crystalgrowth sites. Dependence of the degree of inhibition with the change in driving force has been observed. The order of the degree of inhibitionon the rate of crystal growth of calcium oxalate monohydrate for various additives studied is as follows Ba> Oa .These results suggest thatthe use of natural plants ,weeds and its extracts rather than medicine prepared chemically which have vast side effects, may provide a usefuldrug manufacturing and therapeutic approach to urinary stones and many disease.

KEYWORDS: Crystal growth, Constant composition method, Seed crystals, Growth from solution, calcium oxalate monohydrate(COM).

1. INTRODUCTIONUrinary stones formation is a consequence of increased urinary su-persaturation with subsequent formation of crystalline particles. Crys-tal growth and aggregation of calcium oxalate depend not only on theexcess of calcium and oxalate concentration but also on the presenceof various foreign substances. Calcium oxalates were the main inor-ganic components in pathological deposits and play a key role in theformation of ordinary stones. There are at least four theoretical causesof the formation of large crystals and aggregation of calcium oxalatein the urine,1-A high level of supersaturation with respect to the salt,2-A high calcium / oxalate ratio (at agiven level of supersaturation asin 3-A low level of inhibitors of its crystal growth,4-An increasedlevel of some promoter of crystallization (either heterogeneous nuclea-tor or glutting agent. Several factors affect the growth of urinarycalculi. Different mineral metabolisms are important in the formationof urinary stones 1. At the same time, hypercalciuria has receivedmore attention from researchers of urolithiasis. Menon et al.2 havediscussed the pathophysiology of calcium oxalate stone formation.The urinary calculi are composed of mainly crystalline components.

Multiple steps are involved in the formation of the crystals, which arenucleation, growth and aggregation. The stone formation begins fromthe occurrence of nuclei and the formation of these nuclei is fromsupersaturated urine. Supersaturation also depends on urinary pH,ionic strength, and solute concentration of certain glycoproteins,complexations and the pathogenic factors, which are quite complexand well explained by Menon et al. 2.

These crystals are generally in the form of calcium oxalate monohy-drate COM. Humans normally have biological control mechanisms toprevent COM crystallization in the urine by inducing inhibitors thatdecrease nucleation, growth, and aggregation of COM crystals 3,4. Inparticular, inhibitors in urine will transform COM to calcium oxalatedihydrate (COD) 5,6. However, common aqueous solutions are muchdifferent from those in biological systems. Urinary stones are usuallyformed within membrane-bound micro space, and the nucleation,growth of urinary stones are regulated by organic matrix 7, 8.

Although, technological progress of medicine There is no satisfac-tory drug available for use in clinical therapy then there is a need toexplore more safe and cheap drugs from natural resources. Plantshave played important role as sources of medicine for human beingsince ancient times. More recently several studies have been madeon the crystal growth and dissolution of calcium oxalate monohy-



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Fig.(1), X-ray difractogram of calcium oxalate monohydrate crystal

drate (COM) such as citrate 9, macromolecules and, such as poly passing aliquots through ion exchange resin (Dewix-50) in the hydro-gen form and titrating the eluted acids with standardized sodiumhydroxide solution of suitable concentration using phenolphthaleinas indicator. All solutions prepared and stored in Pyrex vessels.

2.2. Aqueous extracts preparationBarley fresh seeds and Barley(Ba) and Oat (Oa) were collected, andcrushed by using of (Micro-Feinula-Culotti) FMC. Aqueous extractsof seeds of Barley (Ba) and Oat (Oa), was prepared by boiling 100gm. of powder seeds in 400 ml distilled water for 1hour and thenfiltering it with a Whatman filter paper twice.

2.3. Preparation of seedsCalcium oxalate seeds were prepared by adding one liter of 0.1 Mcalcium chloride solutions to one liter of sodium oxalate solution (0.1M) at 298 K at a rate of 250 ml per half an hour. The sodium oxalatesolution was constantly stirred throughout the addition. The seedsuspension was allowed to age with stirring for one day and was thenfiltered and the seed crystals were washed with deionized distilledwater to remove surface contamination essentially chloride and ox-alate ions. The seed crystals were aged for one month, then were re-filtered and washed further with deionized distilled water. The laterprocess was repeated several times. The seeds were then filtered anddried. The seed material was then subject to x-ray powder diffractionstudies, scanning electron microscope, TGA analysis and determina-tion of specific surface area (SSA).

2.4. Calcium oxalate characterization:X-ray powder diffraction studies were made using a Philips X-raygenerator and powder diffractometer (model XRG-300, Philips Elec-tronic Instrument), using CU-K radiation. The solid sample was wellground and mixed with internal standard, potassium bromide the ratioof about 4:1 by weight. The sample and standard were filled in arectangular cavity (1.5 cm × 1.0 cm × 0.05 cm) of a 3.8 cm × 3.8 cm × 0.2cm aluminum solid holder and were slowly scanned at a speed of 10 /4 20 = 10° to 90°.

2. MATERIALS AND METHODS

2.1. ChemicalsSolutions of calcium chloride, sodium oxalate, Sodium chloride andhydrochloric acid, were prepared using Reagent Grade chemicals (ElNasr– Pharmaceutical Chemical Company) with triply distilled deion-ized water.

Calcium chloride and sodium oxalate solutions were analyzed by

The common oat (Avena sativa) is species of cereal grain mainlygrown for its utilization for human consumption as oatmeal as well asfor livestock feed Oat has always been regarded as a health promot-ing food without clear knowledge of its specific health related effects.However, today it is known for its effects on satiety and retardedabsorption of nutrients as well as a deterrent of various disorders ofthe gastrointestinal tract. These beneficial effects are chiefly due tothe soluble fiber content of oats. Today oats is among the richest andmost economical sources of soluble dietary fiber The present interestin soluble oat fiber originated from reports that showed that dietaryoats can help in lowering cholesterol27-29, postprandial blood glucoselevel 30- 32 as well as modifying immune response and reducing risk ofcolon cancer 33,34.

Barley (Hordeum vulgare L.) was considered as antipyretic, sedative

In Egypt, medicinal plants were and still are widespread among theEgyptians for the treatment of several ailments. Scientists have at-tempted over the years to record and study the old recipes. Further-more, there is an increasing trend toward the use of natural remedies,hence the role of naturally occurring compounds and herbal teas areproviding to be of increasing interest in the field of alternative medi-cine. Barley and Oat are still used today in folklore medicine. In thepresent study, we report a systematic investigation on the inhibitioncrystal growth of calcium oxalate by barley (Ba) and Oat (Oa) whichhave multifunctional properties.

amino acids(poly-L-Asp and poly-L-Glu) and poly(acrylic)acid 10. aminoacid sequence 11 and length 12, metallic ions13 and their complexes,sodium dodecyl sulphate 14, α-keto glutaric acid 15 (a normal physi-ological constituent of urine), maleic acid copolymers16 ,protein fromhuman kidney17 plant extracts 18-20on inhibition of calcium oxalatecrystallization.

infusion recommended to children to keep their vitality, depurative,laxative and diuretic21. Barley ß-glucan is effective ashypocholesterolaemic ingredient in foods22. The results suggestthat barley ß-glucan concentrate has potential as ingredient in reduc-ing plasma LDL Cholesterol in human 23.antioxidant 24in Bread Mak-ing and their effects on Human Glycemic Response25 and functionalfood ingredients26.



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The scanning electron microscope was studied to insure that theprepared seed is COM, Scanning Electron microscopic micrograph ofCOM seeds is shown in Fig. (2) by using optical microscopy and sizedetermined with filer eye piece at a magnification of X 10.000. Ca C2O4. H2O (bi axial positive) is strong. Thermal gravimetric analysis (TGA)study was done on prepared seed is shown in (Fig. 3).

Fig. (2). SEM Micrographs of COM seeds

Fig(3):TGA thermogram of COM crystals

0.00 10.00 20.00 30.00 40.00 50.00 60.00Time [min]

20.00

40.00

60.00

80.00

100.00

%TGA

2.5. Measurements of surface area:The specific surface area (SSA) was determined by the BET methodapplying equation (1)

RTPAN

. Vc . )/S(S . )P/P(1 1/W.SAA 0cs0

gcg0−= ..........(1)

Where:W: Weight of solid,Sg : Desorption single area,

Sgc: Single area of calibration,Vc : Volume of calibration,N° : Avogadro’s number,Acs: Cross-sectional area of adsorbateMolecule,Po : Ambient pressure.

2.6. Crystal growth measurementsCrystallization experiments were made in a Pyrex glass vessel of ap-proximately 300 ml capacity. The solutions were maintained at therequired temperature (37°C) by circulating thermostated water throughthe outer jackets. Working solutions subjected to continuous mag-netic stirring (and pre-saturated nitrogen gas bubbled through thesolutions during the experiments to exclude carbon dioxide). Crystal-lization experiments were followed through potentiometric measure-ments. The potentiometric measurements were performed by meansof a Metrohm combititrator: consisted of dosimate model 665,Impulsomate model 614, pH mater model-632 stirrer model E649, wasused to control the addition of titrant solution consisting of 0.15 Msodium, chloride, into the reaction vessel since the Impulsomate pro-vides proportional system was able to respond to a change of Emf of< 0.002 mv on the addition of reagents. Using a calcium specific ionelectrode (Metrohm, Model 9050), coupled with a calomel referenceelectrode (Model 90.02 Orion Research incorporated laboratory prod-ucts group).

3. RESULTS AND DISCUSSIONThe effect of aqueous extracts of Ba and Oa on the mechanism ofcrystal growth of COM crystals are studied at 37oC, I = 0.15 mol.dm-3,pH = 6.5±0.05 using constant composition method. Crystallization isubiquitous in biological systems where interactions between inor-ganic (salt, ions, etc.) and organic components (proteins, lipids, etc.)often mediate physiological processes in the human body, such asbone and teeth formation 35, 36. In spite of the importance of studyingthe dissolution and crystallization of calcium oxalate, it is poorly de-veloped, particularly from a kinetic point of view and still containsmany unanswered questions. The simplest models of calcium oxalatestone formation describe it as taking place in for stages:

1. There must be a period of high super saturation during whichcrystal nucleation is “triggered off either homogeneouslyor; heterogeneously,

2. This is followed by a period of rapid crystal growth and / oraggregation when the primary particles accrete in size,

3. If this occurs sufficiently rapid, a particle may be generatedwhich is just large enough to be trapped at some narrowpoint in the urinary system (the so-called free particle theory),

4. Alternatively it has been suggested that a primary particlemay become attached to the walls of the renal tubules orcollecting system through the participation of some gluingmaterial (the so-called field particle theory). In both cases a



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nucleus is formed which constitutes a center around whicha stone will form by the continued processes of crystal growthand adhesion.

The concentrations of ionic species in the solutions were calculatedfrom mass-balance and electro-neutrality expressions as describedpreviously 37, by successive approximation for, I, the ionic strength,using the thermodynamic equilibrium constant, K, for the variousassociated species. Activity coefficients were calculated from theextended form of the Debye-Huckel equation proposed by Davies 38.

s -log s Rate /10-9 mol min-1 m2 -log R

0.25 0.602 1.0938 7.9650.30 0.523 1.6032 7.7950.35 0.456 2.3988 7.6200.40 0.398 3.1989 7.4950.45 0.347 4.3652 7.3600.50 0.301 5.8210 7.2350.55 0.260 7.3961 7.1310.60 0.222 9.5499 7.020

The order of crystal growth of COM crystals in the presence of 10-7

mol dm-3 was found ̃ 2 which indicates surface controlled mechanismfig(4).

7.0 7.2 7.4 7.6 7.8 8.0

0.20

0.25

0.30

0.35

0.40

0.45

0.50

0.55

0.60

0.65

- Lo

g R

- Log σ

Fig (4). Plots of –log R against –log s for crystal growth of calcium

The the effects of change of temperature on the rate of crystal growth

7.4 7.6 7.8 8.0 8.2 8.43.20

3.25

3.30

3.35

3.40

3.45

3.50

3.55

-Lo

g R

10 3 1/T

Fig. (5). Plots of –log R against 1/T for crystal growth of calciumoxalate monohydrate crystals in the presence (Ba) at s = 0.4 usingEMF.

S is proportional to the number of growth sites available on the seedcrystals surface and n is the effective order of reaction. In general, therates of crystal growth and dissolution of alkaline-earth metal saltsare markedly inhibited by the addition of certain additives.

In the present study, the rates of crystal growth of COM crystalswere studied at 37°C, I = 0.15 (NaCl) mol/dm-3, pH= 6.5 and at σ = 0.4in the presence of aqueous extracts of Ba and Oa using constantcomposition method. The effect of degree of supersaturation (σ =0.25– 0.60) on the rate of crystal growth of COM crystals were studied.The effect of change of σ on the rates of crystal growth of COM thepresence of 10-7 mol dm-3 Ba was studied as show in table (2).

Table 1: Effect of degree supersaturation, s on the rate of crystalgrowth calcium oxalate at t = 370C.

The relative super-saturation, s , may be expressed by:

σ = (π0 1/2 – π1/2) / π0

1/2 ......................eq. (2)

In which π0 1/2 is the molar concentration product of calcium oxalate,[Ca2+] [C2O4

2-], in the solution, and π1/2 is the solubility value at equi-librium at the same ionic strength (0.15 mol dm 3- ). For many sparinglysoluble salts, Ma Ab the rate of precipitation, normalized for seedsurface area, can be explained by equation. 2, in which k is the growthrate constant,R = d [ma Ab] /dt = k s σn ......................eq.(3)

- log f2 = A Z2 {I1/2 / (1 + I1/2) – 0.3 I} ......................eq. (1)

oxalate monohydrate crystals at 37 °C in the presence of Ba

of COM crystals in the presence of 10-7 mol dm-3 Ba were studied.Calcium oxalate crystal growth experiments at various temperaturesfrom 10 to 39oC in the presence of Ba at relative supersaturation (s =0.4) are plotted as show in fig (5). A plot of log R versus l/T was linearwith a slope corresponding to an activation energy for crystal growth,Ea=5.33 k cal .The small value of activation energy and independenceof the of Ea of the of the crystal growth rates on the stirring rate andthe negligible change in rate, rules out bulk diffusion of electrolyte tothe crystal surface as the rate controlling step and suggests an inter-face mechanism for the growth of calcium oxalate monohydrate.



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The effect of change of ionic strength on the rates of crystal growthof COM crystals in the presence of 10-7 mol dm-3 Ba at the sameexperimental conditions was studied as show in fig (6). It was foundthat increasing ionic strength of the medium leads to increasing therate of crystal growth of COM crystals i.e. decrease the degree ofinhibition of Ba. The changing of rates of crystal growth of COMwith changing ionic strength of the medium indicates that the inhibi-tion is electrically in its nature.

10 12 14 16 18 20

2.0

2.5

3.0

3.5

4.0

4.5

5.0

5.5

Rat

e / 1

0-8 m

ol m

in-1 m

-2

I x10-2 mol dm-3

Fig. (6). Effect of change of ionic strength on the rate of crystalgrowth of COM in presence of (Ba) additive at s = 0.4 using EMF.

The different rates of crystal growth of COM crystal in presence ofaqueous extracts of Ba and Oa were measured and the results aresummarized in Table (2), shows that concentration as low as 10-6 moll-1 for each additive reduced the crystal growth rates by at least 72.99and 61.73 %, Ba and Oa respectively. Compared to that in puresolution at the same relative super-saturation σ = 0.4.

Table (2): Crystal growth of calcium oxalate crystals Tca+2: Tox-2 = 1:

1 at σ = 0.4, t =37oC, I = 0.15 mol dm-3, pH= 6.5 in the presence of Baand Oa using EMF.

Tca+2/10-4 s Additives/10-6 [inhibitor] Rate/10-8 R0/R0 -RI R0 -R I / R0Mol dm-3 Mol dm-3 -1 mol dm-1

m-2

2.791 0.40 3.19892.791 0.40 1.00 Ba 10 2.518 4.7 21.282.791 0.40 2.00 Ba 5 2.076 2.850 35.092.791 0.40 3.00 Ba 3.33 1.795 2.279 43.872.791 0.40 4.00 Ba 2.5 1.558 1.950 51.282.791 0.40 5.00 Ba 2 1.392 1.770 56.502.791 0.40 6.00 Ba 1.67 1.248 1.640 60.972.791 0.40 7.00 Ba 1.43 1.167 1.574 63.532.791 0.40 8.00 Ba 1.25 1.003 1.455 68.722.791 0.40 9.00 Ba 1.11 0.952 1.424 70.222.791 0.40 10.00 Ba 1 0.864 1.370 72.992.791 0.40 3.19982.791 0.40 1.00 Oa 10 2.7577 7.25 13.792.791 0.40 2.00 Oa 5 2.4843 4.13 24.212.791 0.40 3.00 Oa 3.33 2.1827 3.08 32.472.791 0.40 4.00 Oa 2.5 1.9590 2.85 38.762.791 0.40 5.00 Oa 2 1.7896 2.27 44.052.791 0.40 6.00 Oa 1.67 1.6609 2.08 48.082.791 0.40 7.00 Oa 1.43 1.4974 1.88 53.192.791 0.40 8.00 Oa 1.25 1.4018 1.78 56.182.791 0.40 9.00 Oa 1.11 1.3172 1.7 58.822.791 0.40 10.00 Oa 1 1.2243 1.62 61.73

Typical plots of the rate of crystal growth of calcium oxalate monohy-drate in the presence of inhibitors are shown in Fig (7). The crystalgrowth rate of calcium oxalate monohydrate in the presence of addi-tives molecules decreases with the successive additions of inhibitor.

0 2 4 6 8 100.0

0.5

1.0

1.5

2.0

2.5

3.0

3.5

B a O a

Rat

e/ 1

0-8 m

ol m

in-1 m

-2

C x 10 -6 m o l dm -3

Fig. (7). Plot of the rate of crystal growth of COM crystal against Baand Oa at s = 0.4, t = 37 °C and I = 0.15 mol dm-3.

The inhibitory activity on the COM crystal growth in the presence ofthe various additives tested is expressed as

Inhibition percentage= 100 (Ro - Ri) / Ro (8)

It can be seen that, the percentage inhibition of the crystal growth asa function of the additive concentration is shown in table (3), fig (8).The effectiveness of the inhibitors tested was of the order: Ba > Oa atthe same relative super-saturation

0 2 4 6 8 1 00

1 0

2 0

3 0

4 0

5 0

6 0

7 0

8 0

B a O a

% In

hib

itio

n

C X 10 -7 m o l d m -3

Fig (8):- Degree of inhibition of COM in the presence of Ba andOa ats =0.4, t=37oC, s = 0.4, t = 37 °Cand I = 0.15 mol dm-3

Crystal growth inhibitors are able to retard or blocked the crystalliza-tion process even if added in trace amounts. The effectiveness ofinhibitors can explained either by complexation of the inhibitor, usu-



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ally a chelating or sequestering agent, with the lattice cation, or byadsorption of the additives at active sites on the crystal surfaces andsometimes may be both mechanisms can exist at the same time. In-hibitors which are anionic are very good inhibitors for dissolutionand crystallization of COM crystals especially those containing -OH,-COO, -OCH3, -NH2, -SO3 and P-O-P (39).

Barley (Hordeum vulgare L) grain, like other cereals, contains differ-ent chemical compounds–carbohydrates , proteins, lipids, minerals,and small quantities of B-group vitamins, including thiamine(B1),riboflavin (B2), nicotinic acid, and pyridoxine (B6), as well aspanthothenic acid, biotion, folic acid, and vitamin E40, the ß-glucansoluble fibre of cereals, which consists of linear unbranched polysac-charides of linked ß-(1 g3)-(1 g4)-D glucopyranose units41. essen-tial amino acids Thr ,Val, Meth, Iso- Leu, Tyr, Phe, Lys,Cys,Tyro andhis and non-essential amino acid Asp, Ser ,Glu ,Pro ,Gly, Ala andArg42-44. Barley contains all the known tocols (tocopferols andtocotrienols)45 antioxidant and phenolic compound 46. The phenoliccompounds in barley include phenolic acids (benzoic and cinnamicacid derivatives), flavonoids, proanthocyanidins, tannins, and aminophenolic compounds 47.

Oats (Avena sativa) are cereals rich in dietary fibers; grain containsprotein with beneficial amino acid composition, advantageous profileof fatty acids, with high quantity of water-soluble ß-glucans andantioxidants 48-51. There are many types of dietary fibres present inoats; cellulose, arabinoxylans and beta-glucans. Cellulose is a (1g4)-ß-D-glucan, The mixed linked (1g3),(1g4)-ß-D-glucan are composedof ß(1g4)-linked glucose units with a single ß(1g3)-linked glucoseevery two or three units. The (1g3) linkages make beta-glucanssoluble52. The beta-glucans are high molecular weight polysaccha-rides that form highly viscous solution53, antioxidant and unsatur-ated fat54.

The aqueous extract of Ba and Oa containing amino acids, proto-nated –COOH group, negatively charged groups or ions as NH2 –,SO32- and OH- that adsorbed effectively on cationic sites, and reduc-ing the rate of crystal growth of calcium oxalate monohydrate crys-tals. As the negativity of the additive increases on the surface ofCOM crystal, Ca2+ active sites nearly completely blocked and thecrystal growth rate of COM crystal decrease. From previous studieson the effect of amino acids on the dissolution and crystallization ofCOM, it was found that the effective part in anionic part as NH2 –,SO3

2- and OH- or any basis group. Anions of the additive moleculeslow down the rate of crystal growth of COM crystals by adsorptionon mineral surface in competition with oxalate ions thus reducing the

number of empty sites to receive dissolving lattice oxalate anions.The same case occurs in case of cationic part of the additive that willadsorb at Ca2+ active sites in competition, with calcium ions. In thiscase, of inhibition, it might be expected to be highly sensitive to thenature (morphology) of the exposed crystal surface, which decreasedthe rate constant. .In general, the electrostatic potential near the surface of the particlesplay an important role not only for static properties but also for thekinetics behavior55. The strong binding affinity of di- and tri-carboxy-lic acids for calcium stone minerals, particularly CaOx, would indicatethat proteins rich in these amino acids are more likely to play a func-tional role in stone pathogenesis than those possessing only fewsuch residues. The binding affinity of these acids is thought to bedue to the ability of their zwitterions to adopt favorable conforma-tions in which two carboxyl groups and the amine group can interactwith the mineral surface, without further rotation56.

The marked effect of aqueous extracts (Inhibitors), on the crystalliza-tions of calcium oxalate from Supersaturated solutions have beenexplained in terms of the following three factors: (a) inhibitors, ifionic, will influence on the ionic strength of the solution; (b) inhibi-tors may form stable complexes with calcium ion; (c) Adsorption ofthe aqueous extracts on the crystal surfaces at specific growth sitesand prevent further crystallization. Since the amounts of additive insolution are small, the growth inhibition is most likely caused byadsorption of the inhibitors on the active growth sites on crystalsurfaces rather than binding to solution Ca2+ ions.

Aqueous extracts of Ba and Oa contains (1>3), (1>4)-ß-D-glucan,and phenolic compound. The presence of –OH and -OCH3 groups invery much in the component of aqueous extract of Ba and Oa make agood inhibition for crystallization of calcium oxalate monohydratecrystals. Additives of either organic or inorganic nature play an im-portant role in crystallization processes. It is important to know howthe additives influence the crystallization process as well as the typeand number of polar functional groups contained in additives mol-ecule. Hydrophobic and hydrophilic regions, the molecular weightand concentration of additives and a close match between the spac-ing of acid groups and the spacing of cations of the crystal surfaceare considered among the factors that influence crystallization. It isproposed that the additives have two functions:

a- They could inhibit crystal growth by binding to the growth sites ofthe crystals,b- They could act as a heterogeneous nucleator,



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Generally, the change in the rates of crystal growth produced by theaddition of foreign substances may result either from complexation ofthe inhibitor, usually a chelating or sequestering agent, with the lat-tice cation and by adsorption of the molecules at active sites at thecrystal surfaces. The latter of “threshold effect” may be inducedthrough adsorption at much lower concentrations of the additive

0 2 4 6 8 100.0

0.5

1.0

1.5

2.0

2.5

3.0

3.5

4.0

4.5

5.0

5.5

6.0

6.5

7.0

7.5

Ba Oa

R0/(

R0 -

RI)

1/C 106 mol-1 dm3

Fig. (9). Plot.of R0/(R0 – RI) against [additives]-1 of crystal growth ofCOM in presence of Ba at s =0.4.

4. CONCLUSIONS :The crystal growth of calcium oxalate crystals was investigated in theabsence and presence of Egyptian of herbal medicinal plants Barley

(Hordeum vulgare L.) and oat (Avena sativa). Measurements showthat, a highly significant effect of aqueous extract (Ba) and (Oa), ininhibitions of calcium oxalate crystal growth. The aqueous extract of(Ba) produced maximum inhibition of COM crystal growth followedby the aqueous extract of (Oa), in vitro conditions. This study isuseful to formulate the necessary dosages to inhibit and preventcalcium oxalate calculi.

5. ACKNOWLEDGEMENTS:We express our gratitude to Prof.Dr/Gehan Alaemary Assist Prof. ofBio-Chemistry ,faculty of Technology and Development ,Zagaziguniversity for their advice and reviewing the manuscript.

6. REFRENCES:1. Coe FL , Favus MJ, (Eds.). Disorders of Bone and Mineral

Metabolism, Raven Press, New York, 1992.2. Menon MD, Parulkar BG , Drach GW. Campbell’s Urology,

seventh ed., W.B. Saunders Company, New York, 1988.3. Qiu SR , Wierzbicki A, Orme CA., Cody AM, Hoyer, JR,

Nanocollas GH, Zepeda S, De Yoreo JJ. Molecular modula-tion of calcium oxalate crystallization by osteopontin andcitrate. Proc Natl Acad Sci USA ., 2003,101, 1811.

4. Oehlschlager S, Fuessel S, Meye A, Herrmann J, FroehnerM, Albrecht S, Wirth MP .Role of cellular oxalate in oxalateclearance of patients with calcium oxalate monohydrate stoneformation and normal controls. Urology., 2009,73, 480–3.

5. Thongboonkerd V, Samangoen T, Chutipongtanate S. Fac-tors determining types and morphologies of calcium oxalatecrystals: Molar concentrations, buffering, pH, stirring andtemperature. Clin Chim Acta., 2005, 367,120–31.

6. Jung T, Kim WS, Choi CK. Biomineralization of calciumoxalate for controlling crystal structure and morphology.Mater Sci Eng C., 2004,24, 31–3.

7. Ouyang JM, Dua L, He JH, Tieke B. chem.Lett., 2003,32,268.8. Ouyang JM, Dua L, He J.H . ActaChim.Sinica., 2003,61,159.9. Wang LG, Zahang W, QIU S , Zachowice WJ, Guan XY,

Tang RK, Hoyer JR , De Yoreo JJ, Nancollas GH. Inhibitionof calcium oxalate monohydrate crystallization by the com-bination of citrate and osteopontin . Journal of crystal growth., 2006,29(1):160 – 165

10. Jung T, Sheng X X , Choi CK, Kim WS , Wesson JA , WardMD, Probing crystallization of calcium oxalate monohydrateand the role of macromolecule additives with in situ atomicforce microscopy, Langmuir ., 2004,20, 8587–8596.

11. Silverman LD, Saadia M, Ishal JS, Tishbi N, Leiderman E,Kuyunov I, Recca B, Reitblat C, Viswanathan R. Hydroxya-patite growth inhibition by osteopontin hexapeptide se-quences, Langmuir ., 2010, 26(12):9899–9904.

R0 / (R0 – Ri ) = 1+ 1 / KC ......................eq. (4)

Where R0 and Ri are the rate of crystal growth in the absence andpresence of inhibitor, respectively. The applicability of the Langmuirmodel is demonstrated by the linearity of the plots in Fig.9. The ad-sorption, affinity constant, KL, given by the inverse slop of the linesin Fig.9 are 2.73 and 1.60 x105 l mol-1 for Ba and Oa, respectively.These values reflect the high adsorption affinity at the same relativesupersaturation s = 0.40 in order: Ba > Oa, respectively. The highvalue of the affinity constant for Ba may reflect stronger equilibriumadsorption of Ba on the crystal surface, compared to that of Oa.

The latter effect may be interpreted in terms of the Langmuir adsorp-tion isotherm 58 .This requires a linear relationship between the in-verse of the relative reduction in rate, R0 / (R0 - Ri), and the reciprocalof the inhibitor concentration, according to the relationship:

molecules. The influence of the inhibitors on crystal growth must bestudied under highly reproducible conditions by the constant com-position method described by Nancollas et al. 57and Grases et al. 39.



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