Claims:CLAIMS
I/We claim,
1. An efficient process for the preparation of water-soluble iron carbohydrate complex having a weight average molecular weight of 80 kDa to 400 kDa obtainable from starch.
2. An efficient process for the preparation of water-soluble iron carbohydrate complex as claimed in claim 1, wherein an iron product complex comprising starch.
3. An efficient process for the preparation of water-soluble iron carbohydrate complex as claimed in claim 1, wherein the process comprises of a) Hydrolysis of starch to give hydrolysed sugar derivative, b) Oxidizing one or more dextrins with hypochlorite, oxone, hydrogen peroxide, etc. c) Reacting the oxidized sugar derivatives with iron (III) salt to provide iron (III) carbohydrate.
4. An efficient process for the preparation of water-soluble iron carbohydrate complex as claimed in claim 1, wherein the process is by the preparation of ferric carbohydrate complex which comprises hydrolysis of solution of starch solution wherein hydrolysis was performed using hydrochloric acid, sulfuric acid, acetic acid, resin.
5. An efficient process for the preparation of water-soluble iron carbohydrate complex as claimed in claim 1 to 3, wherein the hydrolyzed starch with consistent molecular weight will result in iron carbohydrate complex with consistent average molecular weight.
6. An efficient process for the preparation of water-soluble iron carbohydrate complex as claimed in claim 1, wherein the preparation of ferric carbohydrate complex which comprises reacting an aqueous solution of ferric hydroxide and aqueous solution of oxidized sugar derivative wherein the oxidation is carried out using a non- hypohalite oxidizing agent.
7. An efficient process for the preparation of water-soluble iron carbohydrate complex as claimed in claim 1 to 3, wherein the hydrolysis is carried out at a pH of 1 to 3, at temperatures in the range of 90 to 120 °C.
8. An efficient process for the preparation of water-soluble iron carbohydrate complex as claimed in claim 1 to 3, wherein the oxidation is carried out at a pH of 4 to 9, at temperatures in the range of 15 to 45 °C.
9. An efficient process for the preparation of water-soluble iron carbohydrate complex as claimed in claim 1 to 3, wherein the oxidation is carried out using oxidizing agents such as sodium hypochlorite, hydrogen peroxide, Oxone at various pH.
10. An efficient process for the preparation of water-soluble iron carbohydrate complex as claimed in claim 1 to 3 wherein the oxidation is carried out using non hypohalite oxidizing agents at acidic pH
11. An efficient process for the preparation of water-soluble iron carbohydrate complex as claimed in claim 7 and 8, treated with iron (III) salt in water at 25 oC to provide ferric carbohydrate complex
12. An efficient process for the preparation of water-soluble iron carbohydrate complex as claimed in claim 1 and 9, wherein comprising dia-filtering the iron (III) carbohydrate.
13. An efficient process for the preparation of water-soluble iron carbohydrate complex as claimed in claim 1 and 9, wherein, comprising isolating the ferric carbohydrate complex by precipitation using ethanol, methanol, isopropanol.
14. An efficient process for the preparation of water-soluble iron carbohydrate complex as claimed in claim 1 and 9, wherein comprising activation of the iron (III) carbohydrate using citric acid.
15. An efficient process for the preparation of water-soluble iron carbohydrate complex as claimed in claim 1 to 14, wherein Iron carbohydrate complex being an oxidation product of starch and being consistent molecular weight of between 80-400 kDa.
, Description:TITLE
EFFICIENT PROCESS FOR THE SYNTHESIS OF IRON (III) CARBOHYDRATE COMPLEXES
FIELD OF THE INVENTION
The present invention relates to efficient processes for the synthesis of iron (III) 5 carbohydrate complex with consistent average molecular weight and iron content.
BACKGROUND OF THE INVENTION Iron carbohydrate complexes are intravenously administered for the treatment of iron deficiency anaemia as iron replacement therapies for the patients with serious clinical consequences. Iron colloids are composed of an iron oxyhydroxide or iron oxide core and 10 a complex carbohydrate coat with an average particle size ranging from 10 to 40 nm. The role of carbohydrate as shell is to stabilize and protect the iron core from hydrolysis, precipitation, and polymerization until the delivery of iron colloids to the mononuclear phagocyte system (MPS). In addition, the carbohydrate shell allows to release iron slowly into the bloodstream and, hence, it prevents the formation of non transferrin bound iron 15 (NTBI) and oxidative toxicity. Hence, iron sugar complexes with various kinds of sugar coating will demonstrate differences in uptake, in vivo stability, iron release profiles, and the content of labile and bound iron species. US FDA has approved several iron-carbohydrate complexes for treatment of iron deficiency that are used intravenously in serious clinical conditions. For example, Iron-20 gluconate (Ferrlecit), Iron-sucrose (Venofer), low molecular weight Iron-dextran (Cosmofer/inferD), high molecular weight Iron-dextran (Dexferrum), generic Iron-dextran (Ironate), Iron- isomaltoside (Monfer), Ferumoxytol (Feraheme), Ferric carboxymaltose (Ferinject/Injectafer). WO2003098564A1 describes the preparation of sodium ferric gluconate by coupling the 25 ferric oxyhydroxide with sodium gluconate. US7179939 discloses a manufacturing
3
process for the sodium ferric gluconate complex, substantially free of excipients for providing the ferric gluconate complex co-precipitated with sucrose. EP1733058B1 discloses the process of preparing iron sucrose complex, containing no more than 15% (wt./wt.) of excipients. Iron sucrose complex in sucrose generally contains 30 contaminants including excipients, free sucrose and by products of the synthesis of the complex which are readily detected by techniques such as gel permeation chromatography (GPC). GB 1076219 describes a method for the manufacture of iron complex with low molecular weight dextrin or dextran and sorbitol for the treatment of iron deficiency anaemia. 35 US4927756 demonstrated a procedure for the manufacture of an iron dextran compound with dextran’s molecular weight in the range of 2000-4000. It is known that iron dextran and saccharides with molecular weight below 1000 Daltons in the reaction conditions results in the formation of toxic products. US6291440B1 discloses iron dextrans consisting of dextrans having a weight average 40 molecular weight of 700 to 1400 and in stable association with ferric oxyhydroxide. This iron dextran’s complexes demonstrated that the number of incidences of anaphylactic side effects were reduced. US3076798 discloses a process for the preparation ferric hydroxide polymaltose complex which comprises of a water-soluble non-retrograding dextrin having an average intrinsic 45 viscosity of 0.025 to 0.075 at 25 oC with an aqueous solution containing ferric ions. US7612109, US20120214986A1, WO2016151367A, US20210155651A1 disclosed a process for water-soluble ferric carbohydrate complexes obtained by oxidation of dextrins followed by coupling with iron (III) chloride. Dextrin contained in the mixture lies between 2 and 40 dextrose equivalents which resulted in water soluble iron carbohydrate 50 complex having an average molecular weight of 80,000 to 400,000 Daltons.
It is evident from published methods that the ferric carbohydrate complexes prepared from dextrins having a varying dextrose equivalents unit. Broadly, it involved two steps or in situ one step processes where both require the oxidation of dextrin and coupling with
4
iron salts. The oxidation methods include sodium hypochlorite with without sodium 55 bromide, hydrogen peroxide, sodium hypochlorite + hydrogen peroxide, oxone + hydrogen peroxide, sodium tungstate etc. In the second stage, iron salts were used for coupling to yield ferric carboxymaltose. The maltodextrins were used with varying dextrose units ranging from 5 to 20 DE which gave varying molecular weights of ferric carboxymaltose. All the processes in the previous art suffer from the following 60 drawbacks, formation of iron carbohydrate complexes with inconsistent average molecular weight and iron content, formation of undesired chlorinated by-products such as chlorides, metal bromides, and carbonates. Thus, generating a large quantity of chemical waste which is difficult to treat and hence, unsuitable for the commercial scale.
The dextrin with varying dextrose equivalents used in the reported methods can be 65 obtained according to the literature methods. However, the methods used various combinations of dextrin’s from 5 to 20 DE which significantly affect the average molecular weight of the ferric carboxymaltose. In order to avoid the variations and to maintain the consistency of the product chemical hydrolysis of starch was used for producing the required ferric carbohydrate complex. This method is robust, novel and 70 gives consistency in the product molecular weight through all the batches during product manufacture.
OBJECTIVES OF THE INVENTION 75 The objective of present invention is to provide a facile and robust process for the preparation of iron carbohydrate complex having average molecular weight of 80 kDa to 400 kDa obtained from starch. The starch after acid hydrolysis was oxidized using sodium hypochlorite and coupled with ferric hydroxide to produce iron carbohydrate complex. 80 Another object of the present invention is to provide an efficient process for the preparation of ferric carbohydrate complex obtainable from oxidation of hydrolyzed starch with sodium hypochlorite, hydrogen peroxide followed by coupling with ferric chloride.
85
5
SUMMARY OF THE INVENTION
Accordingly, the present invention is to provide a facile and robust process for the preparation of iron carbohydrate complex having average molecular weight of 80 kDa to 400 kDa is obtainable from starch. The starch was hydrolyzed, oxidized, and coupled with ferric hydroxide to produce iron oxyhydroxide carbohydrate complex. 90
In addition, the present invention provides a facile and efficient process for the preparation of ferric carbohydrate which comprises reacting an aqueous solution of iron (III) complex and aqueous solution of oxidized product wherein the oxidation is carried out using hypohalite or hydrogen peroxide as oxidizing agents.
DETAILED DESCRIPTION OF PRESENT INVENTION 95 The present invention is to provide a facile and robust process for the preparation of iron carbohydrate complex having average molecular weight of 80 kDa to 400 kDa is obtainable from starch. The starch was hydrolyzed by acid hydrolysis to obtain dextrin of constant molecular weight through all the batches which was oxidized using sodium hypochlorite and coupled with ferric hydroxide to obtain the iron carbohydrate complex. 100
In the present invention, the soluble starch was hydrolysed with hydrochloric acid at pH 1.5 at a temperature of 100 °C. When the molecular weight of hydrolysed product reached desirable value, the acid hydrolysis was terminated by cooling and neutralisation. The neutralised carbohydrate solution was precipitated with isopropyl alcohol, filtered, and dried. The reducing capability of hydrolysed carbohydrate was decreased by the selective 105 oxidation with aqueous sodium hypochlorite (4-6% active chlorine) solution at basic pH in which the terminal aldehyde groups oxidated to carboxylic acid groups.
In another embodiment, the oxidised carbohydrate solution produced as above was mixed with ferric chloride hexahydrate solution. To the above agitated mixture, saturated sodium carbonate solution is added and thereafter the pH is raised to 11.5 with 27% 110 aqueous NaOH. The mixture thus obtained was heated above 100 °C until it turns to brown or dark brown colloidal solution. The solution was cooled, neutralised to pH 6.0 and filtered. The solution thus obtained was purified by membrane process. After
6
purification, citric acid was added, and pH adjusted to 8.0 with sodium hydroxide and the solution was stabilized by raising the temperature to above 100 °C for 60 minutes. 115
In another embodiment, the present invention provides an environmentally friendly process for the preparation of ferric carbohydrate complex which comprises reacting an aqueous solution of iron (III) complex and aqueous solution of oxidized carbohydrate wherein the oxidation is carried out using non-hypohalite oxidizing agent, hydrogen peroxide at various pH and temperatures. 120
In a preferred embodiment, the process for the preparation of ferric carbohydrate complex according to the present invention comprises treating starch with aqueous hydrochloric acid or sulfuric acid to hydrolyzed product with molecular weight 1200 Daltons, reacting the oxidized starch with sodium hypochlorite and non-hypohalite oxidizing agents to give oxidized product. 125
In yet another preferred embodiment, the process for the preparation of ferric carbohydrate complex according to the present invention comprises treating ferric chloride with aqueous sodium hydroxide and hydrolyzed starch to prepare ferric hydroxide sugar complex.
In yet another preferred embodiment the present invention provides an efficient process 130 for the preparation of ferric carbohydrate complex which comprises: oxidizing hydrolyzed starch in an aqueous solution at different pH from 4 to 7 and a temperature 25 to 50 °C, with aqueous sodium hypochlorite solution to form an oxidized dextrin solution, the oxidized sugar derivative reacted with solution with an aqueous iron (III) chloride solution and pH of the oxidized sugar derivative solution and iron (III) 135 salt was maintained between 8 to 12, isolating ferric carbohydrate complex by adding alcohol to the aqueous complex solution. To prepare the iron sugar complex, the obtained oxidized dextrin was reacted with ferric hydroxide. To do so, the oxidized carbohydrate derivatives can be obtained from starch and re-dissolved. Alternatively, the obtained aqueous solutions of the oxidized dextrins are directly utilized for further 140 reaction with ferric hydroxide. For instance, the aqueous solution of the oxidized dextrin can be mixed with ferric chloride and performed the reaction.
7
The oxidation was carried out in an acidic solution, for example at a pH of 1 to 3. The oxidation may be carried out at temperatures in the range of 15 to 50 °C, preferably 25 °C. The reaction may be carried out for a period of 30 minutes to 4 hours, e.g., 1 145 to 3 hours. The aqueous solution of the iron (III) salt and oxidized sugar derivative can be mixed and carried out the reaction. It is preferred to proceed in a manner so that during and immediately after mixing of the oxidized dextrin and the iron (III) salt, the pH is moderately acidic and adjusted to an alkaline pH to a value in the range of 8 to 12, preferably 10 to 11, and maintaining the reaction at a temperature of 25 to 60 °C, 150 preferably 50 to 55 °C. During the oxidation pH is initially maintained at 1 to 3 and at a temperature of 40 to 100 °C, followed by adjusting the pH between 8 to 12 with an aqueous alkali hydroxide, preferably sodium hydroxide and maintaining the reaction at a temperature of 35 to 55 °C, preferably 50 to 55 °C for iron complex formation. Later, the pH can be lowered to 5 to 6, preferably 5.5, by the addition of an acid and 155 maintaining the reaction at a temperature of 35 to 125 °C, preferably 50 to 100 °C. It is preferable to use inorganic or organic acids or a mixture thereof, specifically hydrohalic acids such as aqueous hydrochloric acid. The following examples describes the nature of the invention and are given only for the purpose of illustrating the present invention in more detail and are not limitative and relate to solutions which have been 160 particularly effective on a bench scale.
Comparative Analysis
Gel Permeation Chromatography (GPC)
The GPC measurements were performed on a Thermo Fisher Ultimate 3000 HPLC system coupled with an online refractive index detector (Refractomax 520). The 165 formulations were eluted through a column which was calibrated using commercial Pullulan polysaccharide molecular weight standards (Sigma Aldrich, St. Louis, MO, USA). The standards and samples were analysed using a refractive index detector. The samples were analysed using two different methods described below.
Method 1: Samples were diluted 20 times with deionized water (500 uL sample diluted 170 into a 10 mL volumetric flask) for GPC analysis. Chromatography was performed on a 300x8 mm Shodex OH pak SB-G6B + SB-804 HQ + SB802.5 HQ column (polyhydroxy
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methacrylate with a particle size of 13 nm and a pore size of 1500 nm). Eluent consisted of 7.16 g/L Na2HPO4.12H2O + 3.59 g/L Na2HPO4.2H2O in water at a flow rate of 1.0 mL/min. 0.4% (w/v). Pullulan polysaccharide standards (Shodex standard p-82), Dextran 175 standards (PSS) were used for generating the calibration curve.
Method 2: Chromatography was performed on a 300 x7.8 mm Toso Haas TSK Gel G40000SWXL column (spherical silica with a particle size of 8 um and a pore size of 25 nm). Aqueous solution of sodium azide (0.1 %) and 0.3 % samples at pH 7.0 was used as the mobile phase at an isocratic flow of 0.5 mL/min. Samples were 1% w/v with mobile 180 phase solution. Pullulan polysaccharide standards from 25 kDa to 800 kDa were used for generating the calibration curve.
Dynamic Light Scattering (DLS) and Zeta Potential
The particle size and zetapotential were measured using Horiba SZ100 (HORIBA India 185 Private Limited). The DLS uses a high energy laser light source and a sensitive detector with scattering information at the particles size measurements at both 90° and 173°. HORIBA has included the most sensitive PMT detector available.
In the following the characteristics of the iron carbohydrate complexes are compared with a known methods. It was reported that the iron carbohydrate complex prepared from 190 maltodextrins with varying dextrose equivalents (DE) values gave product with varying molecular weights and iron contents (see the table 1). For example, maltodextrin source with 14.2 DE gave ferric carbohydrate complex with 300 kDa molecular weight, with 6.0 DE gave 258 kDa product. When both sources with 6.0 and 14.2 DE were mixed, it gave 176 kDa product. Also, the ratio of the mixture was not mentioned in the reported methods 195 (WO2011055374A2, US20120214986A1, US7612109). To obtain the required molecular weight of the iron carbohydrate complex, different dextrins with DE values were combined in different ratios which will be always a difficult task to maintain. Therefore, our invention starting from starch followed by chemical hydrolysis and coupling with iron salts to obtain ferric carbohydrate complex is robust, reliable that 200 results in product with consistent molecular weights useful for clinical applications (Table 1).
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Table 1. Comparison of Molecular weights of the iron carbohydrate complexes obtained from different sources (reported) with the product obtained from starch hydrolysis. 205
Source (DE)
Iron carbohydrate complex
Avg. MW. (kDa)
Iron %
Reference
14.2
300
21 w/w
WO2011055374A2
US20120214986A1
6.0
258
25 mg/g
6.0 + 14.2
176
50 mg/mL
16.0
258
24 mg/g
9.6
271 or 143 or 140
26-30.0 w/w
US7612109
6.6
189 or 170
26.5 w/w
14.0
118
29.9 w/w
5.4
178
27.9 w/w
Starch hydrolysis
140-160
24-26%
This patent
In addition, the comparative analysis of our product with the product obtained from maltodextrin showed full agreement of average molecular weights including the iron content and particle size distribution. The core size was found to be 26.8 nm whereas product obtained from maltodextrin showed 25 nm (see table2). The detailed size 210 distributions of the innovation product and maltodextrin was shown in figure 2. Molecular wights obtained from starch were consistent and shown in the range of 140-160 kDa which was similar to the product obtained from maltodextrin.
215
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Table 2. comparison of iron carbohydrate product obtained from starch with maltodextrin
Parameter
From Starch
From Maltodextrin
Iron
5.0
5.0
pH
6-7
6-7
MW [Kda]a
140-160
130-150
MW [Kda]b
250-380
250-350
PDI
2.05
2.1
Thermal treatment
120 oC/15ʹ
120 oC/15ʹ
Core size
26.8 nm
25 nm
Zeta potential
-78.6mV
-47.2mV
a) = Method 1, b) = Method 2
EXAMPLES 220
Example 1
Starch (100 g) was dissolved in distilled water, added 10N HCl. The pH was adjusted to 1.5 and heated at 95 °C for 3 h. Hydrolyzed starch solution was precipitated with isopropyl alcohol, filtered, and dried in oven at 60 °C, to obtain the product as dried powder is 64 g. The hydrolysed starch (60 g) was dissolved in water (60 mL) at 25-30 °C 225 and stirred for 45 minutes at the same temperature. Sodium bromide (0.17 g) was added to the reaction mixture at 25-30 °C. Aqueous sodium hydroxide solution was added to the reaction mixture at 25-30 °C. Aqueous Sodium hypochlorite solution (4% Chlorine, 60 mL) was added to the reaction mixture at 25-30 °C and stirred for 60 minutes at the same temperature. Methanol (150 mL) was slowly added to the reaction mixture at 25-30 ° C 230 and stirred for 45 minutes at the same temperature. Filtered the precipitated solid which was dissolved in water (60.0 mL) was added and stirred for 45 minutes at the same temperature. Ferric chloride (90 g) was slowly added to a pre-cooled water (50.0 mL) at 10-15 ºC and stirred for 15 minutes at the same temperature. Slowly added the ferric chloride solution to the reaction mixture at 25-30 °C. To the reaction mixture, aqueous 235 sodium carbonate solution was added to the reaction mixture at 25-30 °C and stirred for
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30 minutes at the same temperature. The pH was adjusted to 11.0 with aqueous sodium hydroxide solution at 25-30 °C. Heated the reaction mixture to 95-100 °C for 3 hours with stirring. Filtered the reaction mixture through hyflow bed. The obtained filtrate was purified by dia-filtration technique. Ethanol (300 mL) was added to precipitate the 240 compound which was filtered, washed with acetone and dried to get the ferric carbohydrate complex. Weight: 75.5 g. Molecular Wt.: 151 kDa; Iron Content: 19.11 %; Polydispersity index (PDI): 1.70.
Example 2
Starch (200 g) was dissolved in distilled water (2000 mL), added conc. HCl and adjusted 245 the pH to 2.0 and heated at 105 °C for 3 h. Hydrolyzed starch solution was precipitated with isopropyl alcohol, filtered, and dried in oven at 60 °C, to obtain the product as dried powder is 104 g. To the hydrolysed starch (100 g), water (200 mL) was added at 25-30 °C followed by sodium bromide (0.17 g) and the reaction mixture stirred at 25-30 °C for 20 min. pH was adjusted to 9.0 by addition of aqueous sodium hydroxide solution. 250 Sodium hypochlorite solution (4% chloride, 200 mL) was added at 25-30 °C and stirred for 90 minutes at the same temperature. Methanol (400 mL) was slowly added to the reaction mixture at 25-30 °C and stirred for 45 minutes at the same temperature and filtered the precipitated solid (90 g). To the oxidized product (90 g) was added water (180 mL) and stirred for 45 minutes at the same temperature. Ferric chloride (135 g) was 255 slowly added to a pre-cooled water (250 mL) at 10-15 ºC and stirred for 15 minutes at the same temperature. Slowly added the ferric chloride solution to the reaction mixture at 25-30 °C. To the reaction mixture, aqueous sodium carbonate solution was added at 25-30 ° C and stirred for 30 minutes at the same temperature. The pH was adjusted to 11.0 with aqueous sodium hydroxide solution at 25-30 °C. Heated the reaction mixture to 95-100 260 °C and stirred for 5 hours at the same temperature. Filtered the reaction mixture through hyflow bed. The obtained filtrate was purified by dia-filtration technique. Ethanol (260 mL) was added to the obtained compound at 25-30 °C. and stirred for 1 hour at the same temperature. Filtered the precipitated solid, washed with acetone and dried to get the title compound. Yield: 130.5 g. Molecular Wt.: 250 kDa; Iron Content: 22.11 %; 265 Polydispersity index (PDI): 1.20.
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Example 3
Starch (200 g) was dissolved in distilled water (2000 mL), added conc. HCl and adjusted the pH to 3.0 and heated at 105 °C for 3 h. Hydrolyzed starch solution was precipitated 270 with isopropyl alcohol, filtered, and dried in oven at 60 °C, to obtain the product as dried powder is 84 g. To the hydrolysed starch (80 g), water (160 mL) was added at 25-30 °C followed by sodium bromide (0.17 g) and the reaction mixture stirred at 25-30 °C for 20 min. pH was adjusted to 9.0 by addition of aqueous sodium hydroxide solution and added aqueous Sodium hypochlorite solution (4% chloride, 160 mL) at 25-30 °C and stirred for 275 80 minutes at the same temperature. Methanol (400 mL) was slowly added to the reaction mixture at 25-30 °C. and stirred for 45 minutes at the same temperature. Filtered the precipitated solid which was dissolved in water (160 mL) and stirred for 45 minutes at the same temperature. Ferric chloride (120 g) was slowly added to a pre-cooled water (200 mL) at 10-15º C and stirred for 15 minutes at the same temperature. Slowly added 280 the ferric chloride solution to the reaction mixture at 25-30 °C. To the reaction mixture, aqueous sodium carbonate solution was added at 25-30 °C and stirred for 30 minutes at the same temperature. The pH was adjusted to 11.0 with aqueous sodium hydroxide solution at 25-30 °C. Heated the reaction mixture to 95-100 °C and stirred for 5 hours at the same temperature. Filtered the reaction mixture through hyflow bed. The obtained 285 filtrate was purified by dia-filtration technique. Ethanol (260 mL) was added to the obtained compound at 25-30 °C. and stirred for 1 hour at the same temperature. Filtered the precipitated solid, washed with acetone and dried to get the title compound. Yield: 73.0 g. Molecular Wt.: 151 kDa; Iron Content: 16.5 %; Polydispersity index (PDI): 1.30.
Example 4 290
Starch (100 g) was dissolved in distilled water (1000 mL), added Conc. H2SO4 and adjusted the pH to 1.5 and heated at 100 °C for 3 h. Hydrolyzed starch solution was precipitated with isopropyl alcohol, filtered, and dried in oven at 60 °C, to obtain the product as dried powder is 68 g. To the hydrolysed starch (65 g), water (130 mL) was added at 25-30 ° C followed by sodium bromide (0.17 g) and the reaction mixture stirred 295 at 25-30 ° C for 20 min. pH was adjusted to 9.0 by addition of aqueous sodium hydroxide solution and added aqueous Sodium hypochlorite solution (4% chloride, 100 mL) at 25-30 °C and stirred for 80 minutes at the same temperature. Methanol (400 mL) was slowly
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added to the reaction mixture at 25-30 °C. and stirred for 45 minutes at the same temperature. Filtered the precipitated solid which was dissolved in water (270 mL) and 300 stirred for 45 minutes at the same temperature. Ferric chloride (98 g) was slowly added to a pre-cooled water (250 mL) at 10-15 ºC and stirred for 15 minutes at the same temperature. Slowly added the ferric chloride solution to the reaction mixture at 25-30 °C. To the reaction mixture, aqueous sodium carbonate solution was added at 25-30 °C and stirred for 30 minutes at the same temperature. The pH was adjusted to 11.0 with 305 aqueous sodium hydroxide solution at 25-30 °C. Heated the reaction mixture to 95-100 °C and stirred for 5 hours at the same temperature. Filtered the reaction mixture through hyflow bed. The obtained filtrate was purified by dia-filtration technique. Ethanol (260 mL) was added to the obtained compound at 25-30 °C. and stirred for 1 hour at the same temperature. Filtered the precipitated solid, washed with acetone and dried to get the title 310 compound. Yield: 145.5 g. Molecular Wt.: 251 kDa; Iron Content: 19.11 %; Polydispersity index (PDI): 2.30.
Example 5
Starch (200 g) was dissolved in distilled water (200 mL), added 10N HCl. The pH was adjusted to 1.5 and heated at 100 °C for 2.5 h. Hydrolyzed starch solution was 315 precipitated with isopropyl alcohol, filtered, and dried in oven at 60 °C, to obtain the product as dried powder is 135 g. The dextrin obtained was subjected to the oxidation using sodium hypochlorite (4% chlorine, 120 mL) at 25-30 °C for 2 h. To the oxidized product was diluted with water (200 mL) and stirred for at the same temperature. Ferric chloride (205 g) was slowly added to a pre-cooled water (165 mL) at 10-15º C and stirred 320 for 15 minutes at the same temperature. Slowly added the ferric chloride solution to the reaction mixture at 25-30 °C. To the reaction mixture, aqueous sodium carbonate solution was added and stirred for 30 minutes at the same temperature. The pH was adjusted to 11.0 with aqueous sodium hydroxide solution at 25-30 °C. Heated the reaction mixture to 100 °C and stirred for 3 hours and allowed to cool to room temperature. Ethanol (260 325 mL) was added to the reaction mixture to precipitate the solid which was filtered and dried under vacuum. Yield: 205 g. Molecular Wt.: 290 kDa; Iron Content: 19.11 %; Polydispersity index (PDI): 1.70.
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Example 6 Starch (100 g) was dissolved in distilled water, added 10N HCl. The pH was 330 adjusted to 1.5 and heated at 95°C for 3 h. Hydrolyzed starch solution was precipitated with isopropyl alcohol, filtered, and dried in oven at 60 °C, to obtain the product as dried powder is 55 g. The hydrolysed starch (50 g) was dissolved in water (50 mL) at 25-30 °C and stirred for 45 minutes and aqueous sodium hydroxide solution was added to the reaction mixture at 25-30 °C. Aqueous Sodium hypochlorite solution was added to the 335 reaction mixture at 25-30 °C. and stirred for 60 minutes at the same temperature. Ferric chloride (100.0 g) was slowly added to a pre-cooled water (50.0 mL) at 10-15 ºC and stirred for 15 minutes at the same temperature. Slowly added the ferric chloride solution to the reaction mixture at 25-30 °C. To the reaction mixture, aqueous sodium carbonate solution was added and stirred for 30 minutes at the same temperature. The pH was 340 adjusted to 11.0 with aqueous sodium hydroxide solution at 25-30 °C. Heated the reaction mixture to 95-100 °C for 3 hours with stirring. Filtered the reaction mixture through hyflow bed. The obtained filtrate was purified by dia-filtration technique. Ethanol (300 mL) was added to the obtained compound at 25-30 °C. and stirred for 1 hour at the same temperature. Filtered the precipitated solid, washed with acetone and dried to get the title 345 compound. Yield: 73.0 g. Molecular Wt.: 350 kDa; Iron Content: 19.11 %; Polydispersity index (PDI): 1.90. Example 7 Starch (100 g) was dissolved in distilled water, added 10N HCl. The pH was adjusted to 1.5 and heated at 95 °C for 3 h. Hydrolyzed starch solution was precipitated with isopropyl alcohol, filtered, and dried in oven at 60 °C, to obtain the product as dried powder is 55 g. The hydrolysed starch (50 g) was dissolved in water (50 350 mL) at 25-30 °C and stirred for 45 minutes and aqueous sodium hydroxide solution was added to the reaction mixture at 25-30 °C. Aqueous Sodium hypochlorite solution was added to the reaction mixture at 25-30 °C. and stirred for 60 minutes at the same temperature. Ferric chloride (200 g) was slowly added to a pre-cooled water (200 mL) at 10-15 ºC and stirred for 15 minutes at the same temperature. Slowly added the ferric 355 chloride solution to the reaction mixture at 25-30 °C. To the reaction mixture, aqueous sodium carbonate solution was added and stirred for 30 minutes at the same temperature. The pH was adjusted to 11.0 with aqueous sodium hydroxide solution at 25-30 °C. Heated the reaction mixture to 95-100 °C for 3 hours with stirring. Filtered the reaction mixture through hyflow bed. The obtained filtrate was purified by dia-filtration technique. Ethanol 360
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(300 mL) was added to the obtained compound at 25-30 °C. and stirred for 1 hour at the same temperature. Filtered the precipitated solid, washed with acetone and dried to get the title compound. Color of the solid was dark black. Yield: 69.0 g. Molecular Wt.: 145 kDa; Iron Content: 29.11 %; Polydispersity index (PDI): 1.30.
Example 8 365
Starch (200 g) was dissolved in distilled water (200 mL), added 10N HCl. The pH was adjusted to 1.5 and heated at 100 °C for 2.5 h. Hydrolyzed starch solution was precipitated with isopropyl alcohol, filtered, and dried in oven at 60 °C, to obtain the product as dried powder is 135 g. The hydrolyzed sugar derivative was subjected to the oxidation using sodium hypochlorite (4% chlorine, 120 mL) at 25-30 °C for 2 h. To the 370 oxidized product was diluted with water (200 mL) and stirred for at the same temperature. Ferric chloride (205 g) was slowly added to a pre-cooled water (165 mL) at 10-15º C and stirred for 15 minutes at the same temperature. Slowly added the ferric chloride solution to the reaction mixture at 25-30 °C. To the reaction mixture, aqueous sodium carbonate solution was added and stirred for 30 minutes at the same temperature. The pH was 375 adjusted to 11.0 with aqueous sodium hydroxide solution at 25-30 °C. Heated the reaction mixture to 100 °C and stirred for 3 hours and allowed to cool to room temperature. To the reaction mixture citric acid (0.75%, based on volume) was added and heated to 100 oC for 2 h. Ethanol (1000 mL) was added to the reaction mixture to precipitate the solid which was filtered and dried under vacuum. Yield: 245 g. Molecular Wt.: 350 kDa; Iron 380 Content: 19.11 %; Polydispersity index (PDI): 2.05.
Example 8
Stage I: Hydrolysis of starch
Starch (200 g) was dissolved in distilled water (2000 mL), added conc. HCl and adjusted the pH to 1.5 and heated at 105°C for 3 h. Hydrolyzed starch solution was precipitated 385 with isopropyl alcohol, filtered, and dried in oven at 60 °C, to obtain the product as dried powder is 124 g.
Stage II: Oxidation of hydrolyzed starch
To the hydrolysed starch (100 g), water (200 mL) was added at 25-30 °C followed by sodium bromide (0.17 g) and the reaction mixture stirred at 25-30 °C for 20 min. pH was 390
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adjusted to 9.0 by addition of aqueous sodium hydroxide solution and added aqueous Sodium hypochlorite solution (4% chloride, 100 mL) at 25-30 °C and stirred for 80 minutes at the same temperature. Methanol (400 mL) was slowly added to the reaction mixture at 25-30 °C. and stirred for 45 minutes at the same temperature. Filtered the precipitated solid and dried to get the title compound. Yield: 90.0 g. 395
Step III: Complexation with Iron To the oxidized product (90 g) was added water (270 mL) and stirred for 45 minutes at the same temperature. Ferric chloride (113 g) was slowly added to a pre-cooled water (260 mL) at 10-15º C and stirred for 15 minutes at the same temperature. Slowly added the ferric chloride solution to the reaction mixture at 25-30 °C. To the reaction mixture, 400 aqueous sodium carbonate solution was added at 25-30 °C and stirred for 30 minutes at the same temperature. The pH was adjusted to 11.0 with aqueous sodium hydroxide solution at 25-30 ° C. Heated the reaction mixture to 95-100 °C and stirred for 5 hours at the same temperature. Filtered the reaction mixture through hyflow bed. The obtained filtrate was purified by dia-filtration technique. Ethanol (260 mL) was added to the 405 obtained compound at 25-30 °C. and stirred for 1 hour at the same temperature. Filtered the precipitated solid, washed with acetone and dried to get the title compound. Yield: 62.5 g. Molecular Wt.: 160 kDa; Iron Content: 24.11 %; Polydispersity index (PDI): 1.90. Example 9 20 g of hydrolyzed starch were dissolved in purified water (50 mL) and was added 410 sodium hypochlorite (40 mL, 4% chlorine), at room temperature over a period of 15 minutes and stirred for 3 h at room temperature. Ferric chloride (30 g) was slowly added to a pre-cooled water (60 mL) at 10-15º C and stirred for 15 minutes at the same temperature. Slowly added the ferric chloride solution to the reaction mixture at 25-30 °C. To the reaction mixture, aqueous sodium carbonate solution was added at 25-30 °C 415 and stirred for 30 minutes at the same temperature. The pH was adjusted to 11.0 with aqueous sodium hydroxide solution at 25-30 °C. Heated the reaction mixture to 100 °C and stirred for 3 hours at the same temperature. Filtered the reaction mixture through hyflow bed. Ethanol (250 mL) was added to the obtained compound at 25-30 °C. and stirred for 1 hour at the same temperature. The obtained brown solid was filtered, dried 420 in vacuum at 50 °C for 1-2 hours. The obtained brown amorphous solid was dried
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under vacuum at 50°C for 2-3 hours. Molecular weight = 325 kDa. Iron content = 23.0 % w/w. Example 10 20 g of hydrolyzed starch were dissolved in purified water (50 mL) and was added H2O2 425 (5 mL), at room temperature over a period of 15 minutes and stirred for 3 h at room temperature. Ferric chloride (30 g) was slowly added to a pre-cooled water (60 mL) at 10-15º C and stirred for 15 minutes at the same temperature. Slowly added the ferric chloride solution to the reaction mixture at 25-30 °C. To the reaction mixture, aqueous sodium carbonate solution was added at 25-30 °C and stirred for 30 minutes at the same 430 temperature. The pH was adjusted to 11.0 with aqueous sodium hydroxide solution at 25-30 °C. Heated the reaction mixture to 100 °C and stirred for 3 hours at the same temperature. Filtered the reaction mixture through hyflow bed. Ethanol (250 mL) was added to the obtained compound at 25-30 °C. and stirred for 1 hour at the same temperature. The obtained brown solid was filtered, dried in vacuum at 50 °C for 1-2 435 hours. Molecular weight = 340 kDa. Iron content = 21.6 % w/w. Example 11 20 g of hydrolyzed starch were dissolved in purified water (50 mL) and was added sodium hypochlorite (40 mL, 4% chlorine), at room temperature over a period of 15 minutes and stirred for 3 h at room temperature. Ferric chloride (30 g) was slowly added 440 to a pre-cooled water (60 mL) at 10-15º C and stirred for 15 minutes at the same temperature. Slowly added the ferric chloride solution to the reaction mixture at 25-30 °C. To the reaction mixture, aqueous sodium carbonate solution was added at 25-30 °C and stirred for 30 minutes at the same temperature. The pH was adjusted to 11.0 with aqueous sodium hydroxide solution at 25-30 °C. Heated the reaction mixture to 120 °C 445 and stirred for 3 hours at the same temperature. The color of the solution became very dark black. Filtered the reaction mixture through hyflow bed. Ethanol (250 mL) was added to the obtained compound at 25-30 °C. and stirred for 1 hour at the same temperature. The obtained black solid was filtered, dried in vacuum at 50 °C for 1-2 hours. The obtained brown amorphous solid was dried under vacuum at 50 °C for 2-450 3 hours. Iron content = 19.0 % w/w. GPC was not clear, and the product was decomposed.
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Example 12 20 g of hydrolyzed starch were dissolved in purified water (50 mL) and was added sodium hypochlorite (40 mL, 4% chlorine), at room temperature over a period of 15 455 minutes and stirred for 3 h at room temperature. Ferric chloride (30 g) was slowly added to a pre-cooled water (60 mL) at 10-15 ºC and stirred for 15 minutes at the same temperature. Slowly added the ferric chloride solution to the reaction mixture at 25-30 °C. To the reaction mixture, aqueous sodium carbonate solution was added at 25-30 °C and stirred for 30 minutes at the same temperature. The pH was adjusted to 11.0 with 460 aqueous sodium hydroxide solution at 25-30 °C. Heated the reaction mixture to 75 °C and stirred for 3 hours at the same temperature. Filtered the reaction mixture through hyflow bed. Ethanol (250 mL) was added to the obtained compound at 25-30 °C. and stirred for 1 hour at the same temperature. The obtained brown solid was filtered, dried in vacuum at 50 °C for 1-2 hours. The obtained brown amorphous solid was dried 465 under vacuum at 50°C for 2-3 hours. Iron content = 12.0 % w/w. MW. = 75 Kda Example 13 20 g of hydrolyzed starch were dissolved in purified water (50 mL) and was added sodium hypochlorite (40 mL, 4% chlorine), at room temperature over a period of 15 470 minutes and stirred for 3 h at room temperature. Ferric chloride (30 g) was slowly added to a pre-cooled water (60 mL) at 10-15 ºC and stirred for 15 minutes at the same temperature. Slowly added the ferric chloride solution to the reaction mixture at 25-30 °C. To the reaction mixture, aqueous sodium carbonate solution was added at 25-30 °C and stirred for 30 minutes at the same temperature. The pH was adjusted to 11.0 with 475 aqueous sodium hydroxide solution at 25-30 °C. Heated the reaction mixture to 50 °C and stirred for 3 hours at the same temperature. Filtered the reaction mixture through hyflow bed. Ethanol (250 mL) was added to the obtained compound at 25-30 °C. and stirred for 1 hour at the same temperature. The obtained brown solid was filtered, dried in vacuum at 50 °C for 1-2 hours. The obtained brown amorphous solid was dried 480 under vacuum at 50°C for 2-3 hours. Iron content = 6.0 % w/w. MW. = 5 Kda, Reference Example
Maltodextrin (100 g, 12.0 dextrose equivalent) was dissolved in 200 mL water by stirring at 25 °C. Sodium hypochlorite solution (4 % active chlorine) and 0.7 g sodium bromide was added at pH 10. After the oxidation of maltodextrin solution, iron (III) chloride solution (125 g) was added, and sodium carbonate solution (154 g in 500 mL water). Then the pH is adjusted to 10.5 by addition of 27% aqueous sodium hydroxide solution and heated to 100° C for 5 h. After cooling the solution to room temperature, the pH is adjusted to 6-7 by the addition of sodium hydroxide. Filtered the reaction mixture through hyflow bed and washed the bed with water. The complex is isolated by precipitation with ethanol in a range of 1:1 and washed with acetone and then dried under vacuum at 50 °C to obtain the ferric carboxymaltose as a brown amorphic powder. The yield is 139 g (88%) having an iron content of 22.8% weight/weight measured complex metric titration. Molecular weight mw 350 kDa, Polydispersity: 1.47.

DESCRIPTION OF THE DRAWINGS:

Figure 1. Gel permeation chromatography spectra of the iron carbohydrate complexes obtained from starch and product obtained from maltodextrin.
Figure 2. Particle size distribution of the iron carbohydrate complex produced from starch in comparison with that obtained from maltodextrin.