spray drying based technologies for the double ......spray drying based technologies for the double...

Spray Drying Based Technologies for the Double Fortification of Salt with Iron and Iodine

by

Dan Romita

A thesis submitted in conformity with the requirements for the degree of Master of Applied Science

Department of Chemical Engineering and Applied Chemistry University of Toronto

© Copyright by Dan Romita, 2011

ii

Spray Drying Based Technologies for the Double Fortification of

Salt with Iron and Iodine

Dan Romita

Master of Applied Science

Department of Chemical Engineering and Applied Chemistry University of Toronto

2011

Abstract

The fortification of salt with iron may reduce the prevalence of iron deficiency globally, but

fortification is complicated by iron-iodiate interactions. To minimize this interaction, a spray dry

microencapsulation system was developed. This study evaluated the creation and use of this

system, and produced engineered iron premixes for integration into coarse iodized salt.

Bioavailable ferrous fumarate powders were encapsulated to produce small particles (<20µm).

Feed systems containing both suspended and dissolved ferrous fumarate were compared to find

optimal conditions. The premixes were blended into iodized salt at 1000ppmiron and stored at

40OC, ~60%RH. The salt was sampled periodically for 6 months to evaluate iodine stability. All

encapsulated samples showed increased stability. The capsules ability to adhere to the salt as

well as its colour and apparent bioavailability were evaluated. The evaluated samples indicate

that stable double-fortified salt based on inexpensive, coarse, unrefined salt may be obtained

economically by this approach.

iii

Acknowledgments

I would like to express my sincerest gratitude to my supervisors, Professor L.L. Diosady and

Professor Y.-L. Cheng, for giving me the opportunity to advance my knowledge-base and

engineering skills on such a significant and interesting project. Without their invaluable support,

guidance and encouragement, this experience would not have been as wonderfully fruitful as it

was. I greatly appreciate their continuous trust which has allowed me to develop a number of

skills and significantly expand my abilities.

I would like to acknowledge the financial support from all funding agencies, without which this

project would not have been possible. The technical guidance and consultations from Dr. Annie

Wesley of the Micronutrient Initiative were valuable and insightful.

I would also like to thank the members of the Food Engineering Group, in particular Dr. Olive

Li, Lana Kwan and Bih-King Chen, who were outstanding co-workers and friends throughout

this experience.

iv

Table of Contents

Acknowledgments.......................................................................................................................... iii

List of Tables ................................................................................................................................ vii

List of Figures .............................................................................................................................. viii

List of Appendices .......................................................................................................................... x

1 INTRODUCTION...................................................................................................................... 1

2 BACKGROUND........................................................................................................................ 3

2.1 Health and Iron Deficiency................................................................................................. 3

2.1.1 Iron Requirements and Iron Loss............................................................................ 3

2.1.2 Iron Storage............................................................................................................. 3

2.1.3 Iron Deficiency ....................................................................................................... 4

2.1.4 Symptoms of Iron Deficiency................................................................................. 4

2.1.5 The Economic Impact of Iron Deficiency .............................................................. 5

2.1.6 Prevalence of Iron Deficiency and Public Health Classifications .......................... 6

2.1.7 Strategies to Reduce Deficiency Prevalence........................................................... 7

2.2 Iron Sources ........................................................................................................................ 8

2.2.1 Absorption into the body ........................................................................................ 8

2.2.2 Properties of Various Iron Salts .............................................................................. 9

2.2.3 Characteristics of Ferrous fumarate ...................................................................... 10

2.3 Iron-Iodate Interactions..................................................................................................... 10

2.4 Microencapsulation........................................................................................................... 11

2.4.1 Overview of Microencapsulation Processes ......................................................... 11

2.4.2 Spray-drying Encapsulation.................................................................................. 14

2.4.3 Heat and Mass Transfer in Spray drying .............................................................. 16

2.5 Particle Engineering.......................................................................................................... 19

v

2.5.1 Common Particle Formation Processes ................................................................ 20

2.5.2 Particle Formation Processes ................................................................................ 20

2.5.3 Properties Affecting Particle Formation Pathway ................................................ 22

2.6 Encapsulating materials .................................................................................................... 24

2.7 Effect of Storage Conditions............................................................................................. 26

2.8 Spray Drying Based Technologies for the Double Fortification of Salt........................... 26

3 MATERIALS AND METHODS............................................................................................. 27

3.1 Materials ........................................................................................................................... 27

3.2 Experimental Overview .................................................................................................... 28

3.3 Determination of Iron Content.......................................................................................... 28

3.4 Spray drying Conditions ................................................................................................... 28

3.5 Particle Imaging and Surface Analysis ............................................................................. 29

3.6 Experimental Set-up.......................................................................................................... 30

3.7 Double Fortified Salt Preparation and Stability Test ........................................................ 32

3.8 Iodine Analysis ................................................................................................................. 32

3.9 Adhesion Testing .............................................................................................................. 32

3.10 In-vitro Bioavailability...................................................................................................... 33

3.11 Colour evaluation.............................................................................................................. 33

4 RESULTS AND DISCUSSION .............................................................................................. 34

4.1 Process Considerations ..................................................................................................... 34

4.1.1 Iron Oxidation....................................................................................................... 34

4.1.2 Iron Throughput .................................................................................................... 35

4.1.3 Yield Optimization................................................................................................ 36

4.2 Capsule Evaluation ........................................................................................................... 38

4.2.1 Particle Imaging .................................................................................................... 38

4.2.2 EDTA Leaching .................................................................................................... 39

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4.2.3 TOF-SIMS ............................................................................................................ 40

4.2.4 Iodine Stability...................................................................................................... 42

4.3 Formula Evaluation........................................................................................................... 43

4.3.1 Capsule Size and Shape ........................................................................................ 43

4.3.2 The Effect of Excipient on Particle Shape and Size ............................................. 44

4.3.3 Iron Loading.......................................................................................................... 48

4.3.4 Encapsulant Chemistry ......................................................................................... 49

4.3.5 Capsule Colour...................................................................................................... 51

4.4 Adhesion ........................................................................................................................... 52

4.5 Bioavailability................................................................................................................... 56

5 CONCLUSION ........................................................................................................................ 59

6 RECOMMENDATIONS ......................................................................................................... 61

7 REFERENCES......................................................................................................................... 62

vii

List of Tables

Table 2.1: Average values for iron losses in adult humans ............................................................ 3

Table 2.2: Symptoms of Iron Deficiency........................................................................................ 5

Table 2.3: Public Health Category of Numerous Countries based on Anemia .............................. 6

Table 2.4: Health Classifications for Anemia from the World Health Organization ..................... 7

Table 2.5: Summary of Common Iron Salts ................................................................................... 9

Table 2.6: Summary of Encapsulation Processes ......................................................................... 12

Table 2.7: Summary of Spray-dry encapsulation wall materials used in food products .............. 24

Table93.1: Microencapsulated ferrous fumarate formulation design ........................................... 30

Table104.1: Ferrous fumarate solubility in acetic acid .................................................................. 34

Table114.2: Suspended feed versus dissolved feed ....................................................................... 36

Table124.3: Yield as a function of Temperature............................................................................ 36

Table134.4: Yield as a function of Feed Rate ................................................................................ 37

Table144.5: Colour Evaluation of Various Capsules..................................................................... 52

Table154.6: Iron content in sieves after 5 min. of sifting .............................................................. 55

viii

List of Figures

Figure 2.1: Structural Formula of Ferrous fumarate ..................................................................... 10

Figure 2.2: Process Flow Diagram of Spray Drying Process ....................................................... 15

Figure 2.3: Pure Liquid Droplet Drying Balance.......................................................................... 16

Figure 2.4: Mass Balance on a Drying Droplet ............................................................................ 17

Figure 2.5: Summary of Drying Regimes c/o Reference 42......................................................... 20

Figure63.1: Experimental Overview............................................................................................. 28

Figure74.1: SEM image of particles formed from a feed stream containing 30% w/v solids ...... 38

Figure84.2: SEM images of various particles............................................................................... 39

Figure94.3: ToF-SIMS images of various capsules...................................................................... 41

Figure104.4: Iodine stability in iodized salt................................................................................... 44

Figure114.5: SEM micrographs of capsules. ................................................................................. 45

Figure124.6: Iodine stability in iodized salt blended with ferrous fumarate capsules of various

excipient levels.............................................................................................................................. 47

Figure134.7: Iodine stability in iodized salt blended with ferrous fumarate capsules with low

molecular weight excipients. ........................................................................................................ 47


nominal ferrous fumarate loadings ............................................................................................... 49


common encapsulating materials .................................................................................................. 50

Figure164.10: SEM Image of capsules. ......................................................................................... 51

Figure174.11: Microcapsules adhering to salt ............................................................................... 53

ix

Figure184.12: Sieve Fraction Evaluations ..................................................................................... 56

Figure194.13: Relative apparent bioavailablity plot ...................................................................... 57

x

List of Appendices

8.1 Iron Test Calibration Curve .............................................................................................. 67

8.2 Ferrous fumarate Solubility in Acetic acid Solution......................................................... 67

8.3 Ferrous fumarate Particle Size .......................................................................................... 70

8.4 Process Oxidation ............................................................................................................. 70

8.5 Sample Dependant Process Conditions ............................................................................ 71

8.6 Sample Yields and Selectivity .......................................................................................... 74

8.7 Iodine Stability Data: ........................................................................................................ 76

8.8 EDTA Leaching ................................................................................................................ 83

8.8.1 EDTA leaching iron quantification via Spectrophotometry ................................. 83

8.8.2 EDTA leaching iron quantification via AAS........................................................ 84

8.8.3 Leaching iron quantification via ICP.................................................................... 84

8.9 Salt Moisture Content ....................................................................................................... 91

8.10 Tamil Salt Particle Distribution prior to Blending............................................................ 92

8.11 Iron Distribution for all Samples Evaluated for Adhesion ............................................... 93

8.12 Pictures of Salt Analyzed for Colour ................................................................................ 94

8.13 In vitro Bioavailability Data ............................................................................................. 95

8.14 Viscosity Data ................................................................................................................... 97

1

1 INTRODUCTION

Humans require micronutrients in amounts of micrograms to milligrams per day. Many

micronutrients are vital for healthy development, making them especially important for pregnant

women and children [1]. Deficiencies of vitamin A, iron and iodine are the most prevalent, and

impact more than two billion people [2]. Each year, 115,000 pregnancy related fatalities are

associated with iron deficiency [1]. These deficiencies negatively impact the mental and

physical development of billions, the majority of whom reside in developing countries.

Humans require iodine, on a regular basis, in order to produce key hormones necessary for many

developmental and metabolic processes [3]. Like iron, human bodies have limited storage for

inactive iodine [4]. Salt iodization has been successful in reducing the prevalence of iodine

deficiency, and is widely accepted globally. Iodized salt is available to ~70% of the world, and

is mandated by numerous governments [3].

The success of salt iodization programs can be attributed to the vehicle. Salt is inexpensive, and

consumed universally in uniform quantities; this results in the regulated dosage of added

nutrients [5] [6]. Since salt fortification has been effective with iodine, it would be sensible to

use the iodization infrastructure to incorporate other nutrients into salt.

The addition of iron to iodized salt is complicated by iron-iodate interactions, resulting in the

sublimation of the iodine and the oxidation of the iron. The resulting iron(III) is less

bioavailable, and must be enzymatically reduced prior to absorption into the body [4]; the

sublimed iodine is lost to the atmosphere. To prevent this interaction, a physical barrier between

the iron salt and the iodine source is useful.

2

Research related to the double fortification of salt with iron and iodine has been ongoing at the

University of Toronto for numerous years with principle investigator Professor Diosady in

collaboration with the Micronutrient Initiative. A technique to manufacture salt sized, coated

iron particles has been developed and successfully field-tested. Ferrous fumarate was extruded

with flour into pellets that were later coated with white pigment and polymeric material using a

fluidized bed. This system was found to create an effective barrier and prevent iron-iodine

interactions [8]. Despite the success of these coated particles, there are still a number of

challenges.

The iron particles manufactured by extrusion- then fluidized-bed-coating were noticeable within

the salt, and the particle sizes achievable through this process are limited; the smallest particles

that can be produced effectively by extrusion are ~0.2 mm [9] [10]. The iron particles will

segregate if they are not the same size as the salt grains; therefore, this technology is not

adaptable to fortify salts of all sizes. Sufficiently small capsules can adhere to salt grains of any

size, and would prevent iron segregation.

The objective of this study was to investigate the use of a single-step spray drying encapsulation

process with the ultimate goal of producing iron capsules that reduce the iron-iodate interaction,

and adhere to commercial salt. Capsules must also have suitable organoleptic properties, and

maintain a high iron bioavailability. This was achieved in this study through the evaluation of

encapsulating polymers (dextrin, hydroxypropyl methylcellulose, Arabic gum, and sodium

carboxymethyl-cellulose); encapsulant molecular weight (ranging from 15cps to 3000cps in 2%

aqueous solution); active material loadings (varying from 40% to 80%); and use of excipients

(type and loading), as well as the evaluation of feed conditions including: feed rate, atomizing

energy, drying gas flow rate, and inlet temperature.

3

2 BACKGROUND

2.1 Health and Iron Deficiency

2.1.1 Iron Requirements and Iron Loss

Iron is required for oxygen transport in humans. It is also necessary for DNA synthesis and many

cell regulation processes [11]. The recommended dietary allowances (RDA) set by Health

Canada (2010) range from 7 to 27 mg/day with dependence on age and condition [12]. The

recommended dietary allowance is determined such that the iron absorbed into the body balances

the iron losses, which are estimated to be between 0.5 and 2 mg/day, depending on gender, age

and amount of physical activity [11] [13]. Iron loss within clinical test subjects were shown as

follows [14]:

Table 2.1: Average values for iron losses in adult humans

Type of Loss (where applicable):

Amount lost (mgiron/day):

Urinary losses <0.1

Exfoliating skin cell losses 0.2 to 0.3

Exfoliating intestinal cell losses 0.1

Bowel movement blood losses 0.4

Menstrual losses 0.5

2.1.2 Iron Storage

The two major iron storage compounds in the body are haemosiderin and ferritin [11].

Haemosiderin is composed of colloidal ferric oxides which are found in many tissues [14].

Ferritin is a readily mobilized, iron containing protein which is found in many cells including

erythroblasts and RES liver cells [11]. Healthy human bodies contain 3000 – 4000 mgiron where

4

only about 270 - 770 mgiron is stored as inactive iron [4]. Haemoglobin (Hb) is an active form of

iron.

2.1.3 Iron Deficiency

Iron deficiency is described by 3 severity levels [14]:

Level 1) “Pre-latent iron deficiency” can be characterized by a lack of haemosiderin in the

bone marrow [14]. During this phase, iron absorption is increased, and the

symptoms of iron deficiency are not easily observed [14].

Level 2) “Latent iron deficiency” can be characterized by serum ferritin levels below

15µg/Lblood and Hb levels above 12g/100mLblood [11]. During this phase iron

absorption increases further and symptoms are still not easily observed [14].

Level 3) “Iron deficiency anaemia” or “manifest iron deficiency” involves complete

depletion of the bone marrow iron and substantial reduction of the serum iron [11]

[14]. This can be characterized by serum ferritin levels below 15µg/Lblood and Hb

levels below 12g/100mLblood [11] [14]. The further quantification is also

dependant on age, gender and conditions, ranging from Hb threshold values of 11-

13g/100mLblood for individuals living at sea level [15].

Functional iron deficiency can also occur when disorders or diseases restrict the use of ingested

iron [11].

2.1.4 Symptoms of Iron Deficiency

Iron deficiency can result in a number of impairments and dysfunctions. These include the

impairment of motor and mental development, and the increased risk of neonatal mortality [11].

5

The following are also considered clinical symptoms of iron deficiency [11]:

Table 2.2: Symptoms of Iron Deficiency

Angular stomatitis

Aphasia Appetite disorders

Burning tongue

Concentration Disorders

Constipation Depressive moods

Emotional lability Exhaustion Friable nails and hair

Forgetfulness Headaches Loss of hair Nervousness Plummer-Vinson syndrome

Reduced productivity

Sensitivity to cold

Susceptibility of inflection

Tiredness

It has also been shown that iron deficiency can exacerbate the symptoms of iodine deficiency

and reduce the effectiveness of iodine supplementation [16].

2.1.5 The Economic Impact of Iron Deficiency

The prevalence of iron deficiency negatively impacts the economies of many countries. The

previously stated symptoms make it impossible for iron deficient individuals to perform

strenuous tasks for prolonged periods. Even non-strenuous physical activities are difficult for

iron deficient individuals. The developmental effects of iron deficiencies reduce the productivity

of future workers [17].

Ross and Horton [17] have quantified and combined both the current loss and predicted future

loss of productivity caused by iron deficiency. They found Bangladesh loses 1.9% of their GDP

due to iron deficiency; India, Malawi, and Oman also had costs of greater than 1% of their GDP

associated with iron deficiency. Iron deficiency is also thought to increase health care costs as

iron deficient people are more susceptible to environmental contaminants and infections [17].

6

2.1.6 Prevalence of Iron Deficiency and Public Health Classifications

Using Hb concentration in blood as an indicator, the World Health Organization estimated that

1.50–1.74 billion people were affected by anaemia from 1993 to 2005; this represents 22.9–

26.7% of the global population [15]. The highest prevalence was found among pre-school-aged-

children with an estimated 45.7–49.1% being affected [15]. The geographic region with the

highest prevalence is Africa, with an estimated 47.5–67.6% of the population affected [15].

A classification system was used with country-level anaemia estimates for 3 population groups:

pre-school-aged-children, pregnant women and non-pregnant women [15]. The results are

summarized below [15]:

Table 2.3: Public Health Category of Numerous Countries based on Anemia

Number of countries classified in each category Public health category with regards to anaemia

Pre-school-aged-children

Pregnant women

Non-pregnant women

No public health problem 2 0 1

Mild public health problem 40 33 59

Moderate public health problem 81 91 78

Severe public health problem 69 68 54

Total countries included: 192

7

The health categories are defined as follows [15]:

Table 2.4: Health Classifications for Anemia from the World Health Organization

Prevalence of Anaemia (% of People)

No public health problem Less than 4.9

Mild public health problem 5.0–19.9

Moderate public health problem 20.0–39.9

Severe public health problem Greater than 40.0

Iron deficiency is spread among the world and strategies to reduce its prevalence must also be

global.

2.1.7 Strategies to Reduce Deficiency Prevalence

One approach to reducing the prevalence of nutrient deficiencies is the education of people on

the composition of foods, and the requirements of the human body. This is difficult to

implement in developing countries where a majority of people do not have access to education or

a variety of foods.

Another strategy is supplementation, where nutrients are provided on a regular basis. Success of

the “Sprinkles” supplementation program proves this approach can be effective in certain areas

[18]. Due to the non-trivial cost associated with monitoring compliance in remote areas, this

approach can be expensive, even when consolidating multiple nutrients into a single delivery

system. An effective approach to reducing the prevalence of micronutrient deficiencies is food

fortification.

8

There have been many fortification strategies implemented and attempted [5]. These include the

enrichment and fortification of wheat, cereals and rice as well as a number of infant targeted

products, beverages, salt and condiments [5]. The advantages to fortification are: regulated

dosage as a result of regular consumption [5], and increased absorption with the simultaneous

digestion of other foods [4].

2.2 Iron Sources

2.2.1 Absorption into the body

Iron is absorbed in the proximal small intestine, mostly in the duodenum and to a lesser extent in

the upper ileum [4]. The body absorbs iron in the ferrous form [4]. A membrane bound enzyme

present in proximal small intestine reduces iron to the required state [4]. Ferrous iron, in general,

is more soluble, and does not require enzymatic reduction for absorption [4].

Iron absorption is generally increased by organic acids in the stomach, which chelate the iron,

allowing it to remain soluble in the proximal small intestine [4]. Notable acid systems in which

this effect has been observed include: ascorbic, citric, malic, lactic and tartaric acids, as well as

the associated salts [4].

9

2.2.2 Properties of Various Iron Salts

Many iron compounds have been evaluated for the purpose of fortification [19] [20]:

Table 2.5: Summary of Common Iron Salts

Iron Salt Iron Content (w/w %)

Average Relative Bioavailability in Humans

Approximate Relative Cost (% of Ferrous sulphate ●7H2O)

Water Soluble Salts Ferrous sulphate ●7H2O 20 100 100 Anhydrous ferrous sulphate 33 100 70 Ferrous gluconate 12 89 510 Ferrous lactate 19 106 410 Ferric ammonium citrate 18 No Data 210 Ferrous ammonium sulphate 14 No Data 210

Dilute Acid Soluble Salts Ferrous fumarate 33 100 130 Ferrous succinate 35 92 410 Ferric saccharate 10 74 520 Ferric glycerophosphate 15 No Data 1050 Ferrous citrate 24 74 390 Ferrous tartrate 22 62 390

Sparingly Soluble Salts Ferric pyrophosphate 25 21-74 230 Ferric orthophosphate 28 25-32 410

Elemental Iron Electrolytic 97 75 No Data H-reduced 97 13-148 No Data Carbonyl 99 5-20 No Data

Ferrous fumarate, ferrous sulphate and ferrous lactate all are readily available biologically [19].

Less of these iron salts are required to achieve the same level of absorption compared to the

others. Ferrous lacate has significantly less iron content than ferrous sulphate and ferrous

fumarate [19]. Both have Ferrous fumarate and ferrous sulphate are good compromises between

bioavailability, cost and iron density. Ferrous fumarate has an advantage of being almost

tasteless when compared to other potential fortification materials [21].

10

2.2.3 Characteristics of Ferrous fumarate

The structural formula of ferrous fumarate is shown below [21]:

Figure 2.1: Structural Formula of Ferrous fumarate

The stability of this compound is attributed to fumarate, a bidendate ligand, which gives ferrous

fumarate an overall polymeric structure [21]. Ferrous fumarate has no known toxic affects and

has a LD50 of 3,850 mg/kgrat [21]. It is available in granular powders of reddish-orange or

reddish-brown colour [21]. Ferrous fumarate is practically insoluble in water and common

organic solvents [21].

2.3 Iron-Iodate Interactions

Developing countries, India, China and Australia all use potassium iodate for salt iodization [22].

The iodine in postassium iodate can be reduced as shown in the following half cell reactions

[23]:

IO3− + 5H+ + 4e− ⇄ HIO(aq) + 2H2O EO = 1.13V (2.1)

2HIO(aq) + 2H+ + 2e− ⇄ I2(s) + 2H2O EO = 1.44V (2.2)

The electrons required for these half cell reactions are supplied from the oxidation of ferrous

iron:

Fe2+ ⇄ Fe3+ + e− EO = -0.77V (2.3)

11

Resulting in this overall reaction:

10Fe2+ + 2IO3− + 12H+ ⇄ 10Fe3+ + I2(s) + 6H2O EO = 0.42V (2.4)

Diatomic iodine has a vapour pressure of 0.3 mmHg at room temperature, and is driven to the

atmosphere [24]. This reduction, then sublimation, has been demonstrated by various studies

[7]. Encapsulating the iron source has been shown to prevent this reaction [8] [20].

2.4 Microencapsulation

2.4.1 Overview of Microencapsulation Processes

Microencapsulation is required to prevent the iron-iodate interaction and to produce capsules that

are small enough to adhere to salt. Various microencapsulation techniques have been accepted

in the chemical, pharmaceutical, cosmetic, printing and food industries [10]:

12

Table 2.6: Summary of Encapsulation Processes

Obtainable Particle Sizes (µm):

Category Method Min. Max.

Maximum Active Material Loading

(w/w %)*

Simple Coacervation 20 200 60

Complex Coacervation 5 200 90 Chemical Techniques

Molecular Inclusion 5 50 10

Spray Drying 1 50 40

Spray Chilling 20 200 20

Extrusion 200 2000 20

Mechanical Techniques

Fluidized Bed 100 90

*Active Material Loading Based on Volatile Flavours

Particles of less than 0.1mm in effective diameter can only be produced by coacervation,

molecular inclusion, spray drying and spray chilling. The energy required for capsules to adhere

to a surface increases with capsule size [25]. To ensure the electrostatic forces present between

the salt and the capsules are sufficient, the particle size should be minimized. Smaller particles

are also more difficult to detect visually or by tongue-feel.

2.4.1.1 Coacervation

Coacervation is the oldest microcapsulation process [26]. Within the food industry, coacervation

is typically used to produce pastes or powders for use in chewing gums, toothpastes and baked

goods [10].

13

Coacervation processes are divided into 2 categories: simple and complex. Both processes

require a non-continuous phase, which contains the active material, to be dispersed in a liquid.

In both cases, the encapsulating materials in the continuous phase migrate to the non-continuous

phase interface, and are cured or set by a chemical process [10]. In complex coacervation, two

or more encapsulating materials form a complex with low solubility; this reduces the amount of

material retained by the continuous phase [27].

Without substantial work on formulating a stabilized dispersion, the non-continuous phase will

coagulate and produce large capsules [28]. This coagulation is expected to increase the particle

size beyond that which will readily adhere to salt.

2.4.1.2 Molecular Inclusion

Molecular inclusion is applied to some confectionery, instant drink and extruded snack products

[10] [28]. It involves the migration of a single molecule of active material into the centre of a

single β-cyclodextrin molecule. This is achieved by having the active material in either gaseous

or dissolved form [28]. Ferrous fumarate cannot be vaporized and is difficult to solublize.

Ferrous fumarate has an increased solubility under acidic conditions; these conditions would

hydrolyze the β-cyclodextrin ring, and result in polysaccharide fragments which are unable to

entrap the active material.

Molecular inclusion can also be achieved through the use of macromolecules such as liposomes.

Liposome-encapsulated iron sources have been shown to perform inadequately in iodized salt

[29].

2.4.1.3 Spray chilling encapsulation

Spray chilling encapsulation is achieved by dispersing or dissolving the active material in a hot,

molten encapsulant [10] [28]. The hot solution or mixture is then sprayed into a cooled chamber

14

where the molten coating material is brought below its melting point, and solidified [10] [28]. In

this process, hydrogenated oils are preferred as encapsulants as their melting points range from

32O - 70OC [10]. The hydrophobic nature of oil capsules impedes adhesion to the salt, and

causes segregation; thus capsules produced by this technique are not preferred for salt

fortification.

2.4.2 Spray-drying Encapsulation

Spray drying is widely used, and produces particles which are collected using simple, low cost

separation processes [10] [30]. Spray drying equipment is readily available and is easily scaled

for production rates up to and in excess of 100 tonnes per hour [10] [30] [31].

Spray drying encapsulation involves a feed stream which consists of wall and active materials,

dissolved or suspended, within a solvent or carrier liquid. This feed is atomized, and the solvent

or carrier liquid vaporizes as the droplets are entrained within a warm air stream. Once the

drying is complete, solids are separated from the air and collected. A diagram of a spray drying

process is shown below [32]:

15

Figure 2.2: Process Flow Diagram of Spray Drying Process

Spray chambers are typically cylindrical and sized such that the height is 3 or 4 times that of the

diameter [31]. Three types of atomizing nozzles are used: rotary atomizers, pressure nozzles and

two-fluid nozzles [33]. Spray drying typically produces matrix-type microcapsules where the

active material is evenly distributed throughout the particle [34].

There are a number of parameters which effect capsule yield, quality, shape and size. These

parameters include: atomizer type, inlet temperature, drying air flow rate, feed rate, material

solubility, material diffusivity and drying time [32] [34].

Spray-drying has reliably produced capsules less than 20 µm in size for the encapsulation of

volatile flavours [10], oils [35], micro organisms [36], vascular drugs [34], nasal drugs [37] and

sparingly soluble inorganic nutrients [38], using generally-recognized-as-safe materials.

16

2.4.3 Heat and Mass Transfer in Spray drying

The droplet drying process is a combination of heat and mass transfer processes. Once droplets

are formed, they are entrained in warm air. Heat from this warm air is transferred to the droplets

while water vapour from the surface of the droplets transfers, along a concentration gradient, to

the bulk air. As this occurs, the droplets shrink, and the material in the droplets precipitate

and/or coalesce, affecting the transfer rates.

2.4.3.1 Gas Phase

Prior to precipitation, the drying of the droplet is relatively straight forward [33] [34]:

Figure 2.3: Pure Liquid Droplet Drying Balance

The convective heat transfer constant is typically between 2-25 W/((m2)(K)) [39]. In the case of

spray drying, where droplets are moving at the same speed as the surrounding gas, both the

17

Nusselt and the Sherwood number are equal to 2, and heat and mass transfer constants to be

estimated by using the following formulae [33]:

and (2.1 & 2.2)

Where r is the radius of the droplet, α is the thermal diffusivity through the film layer and Dv is

the diffusivity of water vapour in air (~0.305 cm2/s at 60OC) [33]. Concentration and

temperature differences are calculated using the log means as the gradients are non-linear [33].

A mass balance on a single droplet yields the following [33]

[40]:

Figure 2.4: Mass Balance on a Drying Droplet

18

This can be expressed in terms of change in radius:

, (2.3)

and can be integrated (given the initial condition of r = rO) [41]:

(2.4)

This equation can be used to determine the drying rate prior to shell formation. This rate of

vaporization is important to estimate when precipitation first occurs [34].

To incorporate the effects of Stefan Flow, the equation could be changed to the following [41]:

(2.5)

Empirical formulae also exist to estimate drying rate [34].

2.4.3.2 Liquid phase and Droplet Surface

Once the surfaces of the droplets are saturated; crusts, skins or shells are formed [34] [42]. The

crust, skin or shell is thickened through Oswald ripening as more material precipitates, and the

droplets continue to dry [34]. This layer affects mass and heat transfer, impacting the way

droplets dry and how particles are formed.

19

Two groups of models are used to explain the factors which affect droplet drying and particle

formation after the initial precipitation [42] [43]. The semi-empirical approach involves the

construction and use of Characteristic Drying Curves (CDCs) [42] [43]. Alternatively, without

the use of empirical data, the relevant factors can be determined by a Reaction Engineering

Approach (REA) which accounts for crust formation and droplet shrinkage [42].

An understanding of the factors affecting drying and particle formation are required for the

engineering of suitable particles.

2.5 Particle Engineering

Parameters such as bulk temperature, material concentration, material molecular weight, initial

droplet surface area and rheology of the droplet have all been shown to affect the capsule

properties by influencing the particle formation process [34] [42] [31].

20

2.5.1 Common Particle Formation Processes

The following is a summary of particle formation pathways [42]:

Figure 2.5: Summary of Drying Regimes c/o Reference 42

(Permission of Use issued January 10, 2011 by author M. Kraft)

Many particle formation pathways lead to less effective capsules. Shattered, blistered and

shrivelled particles have the active material exposed and may not prevent the iron-iodate

interaction. The particle formation pathway must be controlled to prevent the production of

these types of particles.

2.5.2 Particle Formation Processes

21

Handscomb and Kraft have produced 4 studies between 2008 and 2010 that employed the REA

to relate a number of properties to the major particle formation pathways. The 3 particle

formation pathways are [42]:

1) Effective Diffusion Phase: This process is described by a Fickian-type model and describes the

phase of droplet drying prior to skin formation [42].

2a) Shrinking Core or Dry Shell Phase: After crust formation, the particles could undergo drying

characterized by a shrinking core [42]. Once a crust is formed, the particle size is

constant and the liquid core shrinks as solvent vapour diffuses through the pores

in dried layer [42]. This regime results in solid, homogenous particles [42].

Systems where the wall materials are suspended and/or highly concentrated have

been observed to undergo dry shell particle formation [40].

Alternatively:

2b) Bubble Formation or Wet Shell Phase: This is an alternate process to the dry shell particle

formation described previously [42]. In this regime, bubbles are formed within

the centres of the droplets [42]. This typically results in the formation of hollow

spheroid particles [42]. The start of the wet shell phase involves a sub-phase

known as Shell Thickening or Crust Formation. This describes a process where

shell layers thickens as the core continues to recedes [42]. This drying regime is

typically observed when the materials precipitate before reaching saturation in the

bulk of the droplet [34].

2.5.2.1 Shell Thickening

Shell thickening occurs when the pressure difference across the shell causes buckling, creating

new growth sites for the wall material [42]. Therefore shell thickening stops when the shell has

reached sufficient thickness to overcome the pressure difference [42].

22

Two material properties were shown to affect shell thickening: elastic modulus and Poisson’s

ratio [42]. Elastic modulus and Poisson’s ratio are related to buckling pressure [42]:

(2.6)

Where T is the shell thickness, R is the droplet radius, ν is Poisson’s ratio and E is the elastic

modulus [42]. This indicates that shells formed from weaker (low elastic modulus) and more

rigid (low Poisson’s ratio) materials are thicker as they will continue to fracture, generating new

growth sites, until a greater thickness is achieved [42].

The pressure difference is not related to the properties of the wall material, rather to the

permeability, which in this case, is related to the space fraction void of wall material [42].

Once shell thickening ends, if the shell does not prevent drying entirely, the remaining materials

are deposited unevenly within the shell [42].

2.5.3 Properties Affecting Particle Formation Pathway

The particle formation pathway is determined by the materials ability to redistribute itself within

the droplet prior to skin formation [34]. In addition to drying rate, the diffusivity and solubility

of the dissolved materials impact how the particles are formed [34].

As the droplet shrinks, materials become more concentrated within the droplet. In wet shell

particle formation, dissolved materials are not given sufficient time to redistribute within the

droplet [34]. The rate of drying is so fast that materials are being concentrated at the interface

[34]. Precipitation conditions will be met locally on the surface, and a shell will start to form.

23

This shell hinders the escape of solvent vapours, causing an internal vapour bubble to form [42].

The shell thickening stage is entered and hollow spheroids capsules are created [34] [42] [43].

Dry shell particle formation occurs when the dissolved materials are given enough time to

redistribute themselves within the droplet [34] [42]. When this occurs, initial precipitation

happens when the bulk is near saturation. Since the bulk is concentrated, the vapour occupies

pores within the precipitating material [42]. This produces rigid, solid, uniform particles.

Feeding concentrated solutions or suspensions could also initiate dry shell particle formation; in

this case, the bulk is sufficiently concentrated to undergo dry shell particle formation, even prior

to drying [42].

To ensure particles are not blistered or shrivelled, the material must be highly mobile and/or

soluble to encourage dry shell particle formation. The diffusivity of a dissolved material is a

measure of its ability to redistribute itself within a droplet. Diffusivity is related to molecular

weight and the medium (Einstien-Stokes equation) [44].

The molecular weight of chitosan has been shown to influence the particle size of spray dried

powders [37]. Higher molecular weight materials have lower diffusivities; as seen with chitosan,

this results in earlier shell formation, and the creation of larger particles [34] [37]. This indicates

that the molecular weight differences are large enough to influence precipitation timing, and thus

particle formation.

The molecular weight of the encapsulant is one factor affecting particle formation. The type of

encapsulant must also be selected such that an effective physical barrier can be formed.

24

2.6 Encapsulating materials

Various materials have successfully been used for spray dry encapsulation in food products since

the 1950s [35] [45] [46] [47] [48] [49] [50] [51] [52] [53]:

Table 2.7: Summary of Spray-dry encapsulation wall materials used in food products

Category Wall Material

Properties Examples Current Applications

Hydrolyzed Starches

Very good oxygen-barrier, low cost, low viscosity at high concentrations, poor emulsion stabilization

Corn syrup solids, maltodextrins, dextran, dextrin

Citral and linalyl acetate, ethyl caprylate, cheese aroma, linoleic acid, orange peel oil, lemon oil, Arachidonyl L-ascorbate, Bixin, Cumin oleoresin, Short chain fatty acids

Modified Starches

Very good emulsion stabilization, inexpensive, regulatory limits in some countries

Capsul, N-lok, Hi-Cap

Meat flavor, fish oil, orange oil, d-limonene, l-menthol, butter oil, cream, black pepper oleoresin, vitamin E, Cardamom oleoresin, L-Menthol

Gums Good emulsion stabilization, very good retention of volatiles

Arabic, mesquite, alginate, acacia, agar, carrageenan

Essential oils, monoterpens, orange peel oil, vegetable oils, cardamom oleoresin, linoleic acid, bixin, short-chain fatty acids, lipids, acetyl pyrroline, soy oil, d-limonene, ethyl butyrate, d-limonene and ethyl n-hexanoate, Arachidonyl L-ascorbate, Cardamom essential oil, Bixin, L-Menthol, Black pepper oleoresin, Cumin oleoresin

Car

bohy

drat

es

Cyclo-dextrin

Molecular inclusion of volatile molecules, good oxygen barrier, costly

β – cyclodextrin

Pine flavour, shiitake flavour, d-limonene, ethyl hexanoate, caraway fruit oil, lemon oil

25

Table 2.7 Continued

Cellulosic Material

Good oxidative and thermal stability, good emulsify properties, effective at low concentrations, high solubility at lower temperatures, viscous, available in a variety of molecular weights

Carboxymethyl-cellulose, methyl cellulose, ethylcellulose, celluloseacetate-phthalate, celluloseacetatebutylate-phthalate Hydroxylpropyl methyl cellulose

Calcium citrate, calcium Lactate, fish oil, protiens

Car

bohy

drat

es C

ontin

ued

Hydro-gelling

materials

Sustained and controlled release, pH swelling, cross linkable, costly

Cross linked chitosan, cross linked alginate, Ethylene-co-vinyl acetate, polyethylene oxide, other synthetics

Vitamin C, proteins, pharmaceuticals, insulin, riboflavin

Milk Proteins

Caseinates, skim milk powders, whey proteins

Milk fat, linoleic acid, soy oil, ethyl butyrate, ethyl caprylate, orange oil, oregano flavour, citronella flavours, marjoram flavours, Caraway essential oil

Pro

tein

s

Other Protein

products

Very good emulsifying properties, costly, pH dependant, allergenic potential Soy proteins, soluble

soy, gluten, gelatin,

albumin, peptides

Fish oil, soy oil, oregano flavour, marjoram flavour, caraway essential oil, Lycopene, Arachidonyl L-ascorbate

Maillard products Fish oils

Polyvinyl pyrrolidone Enterococcus fæcium

polymethacrylic acid Calcium citrate, calcium

lactate

Ethylcellulose Ibuprofen, pharmaceuticals

Other Various

Poly(lactic acid) and related polymers Enzymes

26

In addition to the properties listed, it is important to consider the lipophilic characteristics of the

material. The produced capsules must behave like salt to prevent detection when added into

liquid foods. Modified celluloses, gums and starches are available in varieties which are either

hydroscopic, hydrophilic and/or water soluble, resulting in particles with densities in excess of 1

g/mL, preventing the capsules from floating in water, beverages or soups.

2.7 Effect of Storage Conditions

It has been shown that both temperature and humidity are key factors in the acceleration of the

iodate-iron interaction [7] [8] [20] [54]. Higher temperatures result in increased kinetics and

vapour pressures. Greater humidity increases the water content of the salt allowing aqueous

material to be more mobile, thus increasing the likelihood of interactions. Capsules must be

effective in hot, humid environments as these conditions are typical in many developing

countries.

2.8 Spray Drying Based Technologies for the Double Fortification of Salt

In order to prevent the symptoms of iron deficiency, and reduce its prevalence through salt

fortification, microcapsules need to be developed that can: reduce the iron-iodate interaction

under the appropriate conditions; adhere to unrefined, coarse salt; and behave like salt in liquid

foods. Spray drying, compatible with a variety of wall materials, has the potential to create these

capsules. In order to ensure adequate capsule performance, the variables affecting capsule

formation must be manipulated.

Given that a number of carbohydrates have the necessary properties to form adequate iodate-iron

barriers and would be undetectable in liquid foods, the low cost, widely available, GRAS

carbohydrates: dextrin, hydroxypropyl methylcellulose, Arabic gum, and sodium carboxymethyl-

cellulose were selected as encapsulating material candidates. As presented, molecular weight

affects particle formation; the range of molecular weights (measured by solution viscosity)

27

evaluated were 15 cps – 3000 cps in 2% aqueous solution. Low molecular weight excipients

were also used to manipulate the particle formation.

3 MATERIALS AND METHODS

3.1 Materials

Food-grade ferrous fumarate (mean diameter ~10 µm) was generously provided by Dr. Paul

Lohmann Chemicals, Germany. White TiO2 pigment (Ti-Pure R931; Dupont Titanium

Technologies, Delaware) was added for colour improvement. Several food grade polymers were

tested as encapsulating agents. Hydroxypropyl methylcellulose (HPMC E15) was provided by

Dow Chemicals Co., USA. Four molecular weights of sodium carboxymethylcellulose

(NaCMC) were assessed. Lower molecular weight NaCMC (Type: 7M8SF PH ~250 cps in 2%

solution) was provided by Hercules Inc. (Wilmington, DE). Higher molecular weight NaCMC

grades (~3000 cps in 2% solution and ~1000 cps in 2% solution) were obtained from Zhejiang

Sanh Food Science and Technology, Zhejiang, China. Gum Arabica (FT) and another NaCMC

grade (trade name: CMC 15; viscosity~15 cps in 2% solution) were provided by TIC Gums,

Maryland. All NaCMC grades have reported degrees of substitution of 0.7 to 0.95.

Maltodextrin (C*Dry MD DE=7) was given by Cerestar, Indianapolis IN.

Sodium fumarate, glacial acetic acid and all analytical grade reagents used in analyses were

purchased from Sigma–Aldrich, Toronto, Canada. Two food-grade salt products, iodized with

100ppm iodine as potassium iodate, were provided by the Micronutrient Initiative (Ottawa, ON)

and assessed for dual fortification. Fine, refined salt (3-500µm) from Kensalt, Kenya and was

used in iodine stability analysis. A coarser (~2 mm) salt was by The Tamil Nadu Salt Company

and was used for the assessment of organoleptic properties.

28

3.2 Experimental Overview

Figure63.1: Experimental Overview

3.3 Determination of Iron Content

Total and ferrous iron was determined by spectrophotometry as a complex with 1, 10-

phenanthroline as described by Oshinowo et al. [55] [56].

3.4 Spray drying Conditions

All samples were spray dried using a Buchi B290 mini-spray drier. The spray dried products of

both aqueous and suspended ferrous fumarate feed were evaluated in terms of ferrous iron yield.

Ferrous fumarate solubility was maximized using acetic acid. Ferrous fumarate solubility was

measured in 0.5, 1.0, 1.5, 2.0, 2.5, 3.0, 30 and 40% v/v acetic acid in water; the highest solubility

(12g/L) was obtained with 2% acetic acid.

29

The effect of temperature and flowrate on yield was determined by spray drying 100 mL samples

of a ferrous fumarate suspension (10% w/v) containing dissolved HPMC E15 (2% w/v) and

Dextrin DE7 (8% w/v). Parameters were first adjusted to reduce spray chamber fouling; later,

small adjustments were made to determine the precise parameters required to achieve optimal

yield. The evaluated range of inlet temperatures was 125 – 170OC; feed flowrates were varied

between 0.36 and 0.90 L/h. All feed solutions and suspensions were agitated during spray

drying.

The average droplet size was estimated by analyzing the particles produced from spraying a

highly concentrated feed stream. High solid feeds form particles which are approximately the

same size as the droplets [34]. The stream contained: 18% w/v sodium fumarate, 2% w/v HPMC

and 10% w/v ferrous fumarate in water.

3.5 Particle Imaging and Surface Analysis

Particles were imaged using a Hitachi S-2500 Scanning Electron Microscope (SEM) after being

sputter-coated in gold. The operating conditions were: an accelerating voltage of 15kV; a

working distance of 10mm; at a tilt angle of 15O.

Time-of-Flight Secondary Ion Mass Spectrometry (ToF-SIMS) was used to confirm ferrous

fumarate encapsulation using the ION-TOF ToF-SIMS IV instrument at Surface Interface

Ontario, Toronto, Ontario.

To determine the extent of encapsulant coverage on the capsule surface, exposed iron was

leached into a pH 7 EDTA solution. The capsules were added to 25 mL of a 5% w/v Na2EDTA

solution that has been adjusted to pH 7 using 10% w/v NaOH in water. Samples were stirred at a

constant rate using a magnetic stirrer. After 5 min., solutions were filtered (0.45 µm membrane

30

filter) and analyzed for iron using Inductively Coupled Plasma spectrophotometry (ICP AES

Optima 7300 DV, ANALEST, Toronto, Ontario). Detection was achieved at wavelengths of

238.204, 239.562 and 259.939 nm; obtained values were averaged.

3.6 Experimental Set-up

Initially, several categories of formulation variables were investigated, including: the molecular

weight of encapsulating material, the active material loading, the type of encapsulating material

and the excipient loading (Table 3.1). The colour of the most promising formulations was

adjusted using various concentrations of white excipients and titanium dioxide (Table 3.1).

These samples were evaluated for segregation in salt as well as colour.

Table83.1: Microencapsulated ferrous fumarate formulation design

Nominal Iron Content (w/w) Encapsulating Material(s) (w/w)

Control – Ferrous fumarate with no encapsulating material

33% None

Various Molecular Weights

27% CMC 7M8

27% CMC 3000

27% CMC 15

27% CMC 1000

Various Iron Loadings – Adjusted by manipulating dextrin concentrations

26% 20% Dextrin (DE7) / 80% HPMC (E15)

23% 56% Dextrin (DE7) / 44% HPMC (E15)

20% 69% Dextrin (DE7) / 31% HPMC (E15)

16% 80% Dextrin (DE7) / 20% HPMC (E15)

13% 87% Dextrin (DE7) / 13% HPMC (E15)

31

Table 3.1 Continued:

Various Encapsulants

16% HPMC (E-15)

16% Gum Arabic

16% 80% Dextrin (DE7) / 20% HPMC (E15)

16% CMC 15

Various Excipient Loadings

16% HPMC (E-15)

16% 75% HPMC (E15) / 25% sodium fumarate



Excipient Types

16% 50% HPMC (E15) / 50% sodium chloride


Samples Evaluated for Colour and Adhesion

16% 20% HPMC (E15) / 80% TiO2

16% 60% HPMC (E15) / 40% TiO2

16% 80% HPMC (E15) / 20% TiO2

16% HPMC (E-15)






3% 7% HPMC (E15) / 69% sodium fumarate / 24% TiO2



32

3.7 Double Fortified Salt Preparation and Stability Test

KenSalt was blended with iron capsules at a target iron concentration of 1000µg/g. A bench-

scale, Les Industries All-Inox Inc., ribbon blender operating for 15 min. at 22 rpm was used. The

formulations of capsules used in stability testing are summarized in Table 3.1.

The resulting salt samples were stored under two conditions for six months: at 40OC, 40-60%

relative humidity (RH) in an environmental chamber, and at room temperature, ~20% RH in a

closed box; samples were analyzed for iodine monthly.

3.8 Iodine Analysis

Iodine content in salt was determined by iodometric titration, following the AOAC official

method [7] [57].

The method entails reducing iodiate to I2. Iodine is then detected with starch and titrated using a

sodium thiosulphate solution. A minimum of four replicates were taken for each sample.

3.9 Adhesion Testing

Coarse Tamil salt with an initial moisture level of 0.8% w/w (determined gravimetrically) was

blended with ferrous fumarate capsules, as described in 2.4. Moisture levels were adjusted to

1.0% w/w and 2.4% w/w using deionized water applied as a spray during blending. The

formulations of capsules used in adhesion analysis are summarized in Table 3.1.

33

The dual fortified salt samples were then sieved using Ro-Tap Testing Sieve Shaker Model B

(Tyler Consulting Engineers Inc.) for 5 min., with 30, 45, 60, 140, 270 and 400 US mesh sieves.

The mass and iron content in each sieved fraction was determined.

3.10 In-vitro Bioavailability

The iron solubility of the capsules was determined by adding premixes at 5mgiron to 500 mL of

pH 1 hydrochloric acid, maintained at 37OC and shaken at 90 rpm, within a dual action shaker.

Every 15 min., 5mL samples were withdrawn, filtered (0.45µm membrane filter) and analyzed

for iron [58].

3.11 Colour evaluation

The colour of selected capsules were compared by analyzing the relative red, blue and green

signals produced in digital photographs of the samples. Photographs were taken against a white

background with a light source directly above the sample.

34

4 RESULTS AND DISCUSSION

4.1 Process Considerations

4.1.1 Iron Oxidation

Oxidation reduces the bioavailability of ferrous iron [4]; therefore it is important to limit

oxidation during the encapsulation process. Both feed suspensions and solutions were evaluated

to determine the extent of oxidation during the process. An investigation of ferrous fumarate

dissolution techniques found best results with solutions of acetic acid. A maximum solubility of

12g/L was obtained with 2% acetic acid, a significant improvement from the ~1g/L obtained

with 0.1N hydrochloric acid (Table 4.1).

Table94.1: Ferrous fumarate solubility in acetic acid

Concentration

of acetic acid

in water

(% v/v):

0.5 1.0 1.5 2.0

Value Standard

Deviation Value S. D. Value S. D. Value S. D.

Solubility of

ferrous

fumarate

(g/L): 5.7 0.3 7.6 0.6 9.4 2.0 12 1

Concentration

of acetic acid

in water

(% v/v):

2.5 3.0 30

Value S. D. Value S. D. Value S. D. Solubility of

ferrous

fumarate

(g/L):

9.5 0.3 10 1.0 8.1 0.3

Values are

averages

from four

trials

35

Dissolved ferrous fumarate in aqueous acetic acid was spray dried with encapsulating materials.

The capsules were compared with those obtained with suspensions of ferrous fumarate in

aqueous solutions of the coating material. Approximately 55% of the ferrous fumarate dissolved

in aqueous acetic acid was oxidized during spray drying, while only 4% of the iron was oxidized

when the ferrous fumarate was suspended in water. In suspension, the interior of the ferrous

fumarate particles are not exposed to the solution; only the outer most layers are prone to

oxidation. This is an indication that the use of suspended particles result in a less oxidized

product.

4.1.2 Iron Throughput

Maximizing feed concentration is also desirable as increasing the feed concentration lowers the

energy required to evaporate the carrier fluid. The acceptable feed concentration of suspended

material is limited by the atomizer. An excessive amount of suspended solids could damage the

atomizer. In our system, the maximum acceptable suspended solid concentration is 10% w/v

[32]. This concentration is much greater than that which could be achieved by dissolving the

ferrous fumarate (~12g/L or 1.2% w/v).

Considering the difference in extent of oxidation and the low solubility of ferrous fumarate, it is

clear that encapsulating suspended ferrous fumarate is more advantageous than spraying it from

a true solution.

Suspensions present some operating challenges. Feed suspensions must be sufficiently viscous

to prevent particle settling within the feed tube. Viscosities of ~15 cps were determined to be

adequate to prevent settling of ferrous fumarate particles in this system.

36

Table104.2: Suspended feed versus dissolved feed

Suspended Ferrous Fumarate

Ferrous Fumarate Dissolved in Acetic Acid

Throughput Concentration: ~100 g/L < 14 g/L

Percent of Iron Oxidized: ~4% ~55%

Solution Viscosity: ≥15 cps No Restriction

4.1.3 Yield Optimization

Up to 150OC, yield increased significantly with temperature (Table 4.2). The yield remained

constant or slightly decreased when the temperature was raised above 150OC. This indicates that

150OC is sufficient to dry the droplets, minimizing the number of particles that foul the spray

chamber, and maximizing the yield.

Table114.3: Yield as a function of Temperature

Temperature (OC): 125 135 150 160 170

Yield (% of total dry solids, w/w): 72 79 81 80 81

Variables held constant: feed rate (0.72 L/h), atomizing gas flow rate

(667 std LN2/h at 90 psi) and aspirator flow rate (maximum).

The drying gas serves two functions within spray drying: it provides a sink for evaporating

solvents, and it entrains the droplets. Reduced drying gas flow rate (and, thus the drying gas

velocity) decreases the maximum particle size which could be entrained. Decreasing the drying

gas flow rate below maximum resulted in yield reduction.

37

Large droplets cannot be entrained by the drying gas, and settle out prior to the collection stage.

Small particles, formed by small droplets, cannot be separated from the drying gas via cyclone.

Droplet size affects the yield, and can be controlled by the feed rate; droplet size increases with

feed rate [32]. There is a balance between forming small, uncollectable particles with lower flow

rates and larger, difficult-to-entrain particles formed with higher flow rates. The optimal yield

was found with a feed rate of 0.72 L/h (Table 4.3).

Table124.4: Yield as a function of Feed Rate

Feed rate (L/h): 0.43 0.72 1.01 1.30 1.58

Yield (% of total dry solids, w/w): 76 81 72 66 59

Variables held constant: inlet temperature (150OC), atomizing gas flow rate

(667 std LN2/h at 90 psi) and aspirator flow rate (maximum).

Droplet size is also determined by the atomizing gas flow rate [32]. The expansion of the

atomizing gas provides the energy required to form droplet surfaces. An increase in atomizing

gas flow rate results in a decrease in droplet size [32]. The yield decreased substantially when

the atomizing gas flow rate was reduced below the maximum value of 667 std LN2/h.

Droplet size estimates were achieved by analyzing particles formed from spraying a highly

concentrated feed stream [34]. As seen by electron microscopy, the size of the produced

particles, and thus droplets, had a broad range (Figure 4.1). The majority of the droplets were

less than 5µm in diameter, and substantially smaller than the average ferrous fumarate particle

(~10µm). The ferrous fumarate must be contained in the larger particles, suggesting that the

droplet size is dependent on whether a ferrous fumarate particle is contained within the droplet.

38

Figure74.1: SEM image of particles formed from a feed stream containing 30% w/v solids

Under these conditions, the outlet temperature varied between 68O - 78OC, and yields of 64% -

82% w/w were obtained based on total dry solid. These had a slight increase in the iron-to-

coating material ratio. This iron selectivity is between 112% - 121% w/w for all samples

produced during this study. The moisture content of all formed capsules were <4% w/w under

these conditions. Lost particles were too small to be separated by the cyclone (~5%) or too large

to be entrained (~15%). Both were observed, even under optimal conditions.

Using 10% w/v suspended ferrous fumarate, with HPMC and dextrin, optimal yield was obtained

using an inlet temperature of 150OC; a feed rate of 0.72 L/h; an atomizing gas flow rate of 667

std LN2/h at 90 psi and an aspirator flow rate set to maximum.

4.2 Capsule Evaluation

4.2.1 Particle Imaging

To ensure encapsulation occurred, particles with and without encapsulant were imaged (Figure

4.2). The formed capsules had substantially different shapes than the un-encapsulated particles

(Figure 4.2). Even capsules with 90% ferrous fumarate underwent observable changes in

39

morphology; though this level of encapsulant did not have affect on the shape, at higher

magnification, it is clear that many of the rough surfaces are covered by a smooth polymer film

(Figure 4.2).

a)

b)

c)

Figure84.2: SEM images of various particles; a) Spray dried ferrous fumarate (2,000X); b)

HPMC capsule containing 15% ferrous fumarate (500X), c) HPMC capsule containing

90% ferrous fumarate (1,000X)

The spheroids shown in Figure 4.2b were typical of spray capsules of polymeric materials with

ferrous fumarate levels below 60%. These spheroids are formed by wet shell particle formation.

The polymer is unable to redistribute itself within the bulk of the droplet prior to reaching

saturation locally at the air-droplet interface. This forms a hollow shell which collapse during

drying.

4.2.2 EDTA Leaching

To confirm that spray drying resulted in effective encapsulation, samples were immersed in a

pH7 EDTA solution for 5 min., during which exposed iron was partially leached while the

polymeric portion of the capsules remained intact. Since the amount of iron leached is directly

proportional to the amount of iron exposed, the levels of encapsulation can be inferred.

40

Leaching removed 22% of the iron from the control, while 7% - 13% of the iron was removed

from the encapsulated samples of various ferrous fumarate loadings and compositions. Thus

there is a significant reduction in exposed iron after spray drying with encapsulants. The iodine

stability tests show that this reduction in iron exposure can prevent the iron-iodate interaction

considerably (Section 4.2.4).

With dextrin as an encapsulant, the amount of iron leached was similar regardless of iron

loading; 7 - 13% of the iron was leached from these capsules. This indicates that regardless of

iron content, capsules may have similar iron coverage. Only 7±1% of the iron was leached from

HPMC capsules, indicating HPMC is capable of the best coverage with the evaluated iron

loadings. Better coverage is expected when the iron loading is reduced below the evaluated

13%.

4.2.3 TOF-SIMS

TOF-SIMS was also used to confirm ferrous fumarate encapsulation. The CH3O ion signal,

attributed to organic material, showed greater coverage on the surface of the encapsulated

samples than on the control (Figure 4.3). Small amounts CH3O are expected to be produced

from the fumarate component of the iron salt.

Encapsulation also reduced the surface iron signal substantially; further confirming that spray

drying can create an iron-iodine barrier. Substituting a low molecular weight excipient, sodium

fumarate, does not affect the iron exposure (Figure 4.3). Excipients can alter the particle

formation pathway, and are of interest in this study.

41

The encapsulation is imperfect (Figure 4.3). Although, in the encapsulated samples, the exposed

iron area is much smaller than in the ferrous fumarate particles (~10µm); there is iron showing,

indicating that there must be imperfections that breach the capsule walls (Figure 4.3.f).

a)

b)

c)

d)

e)

f)

Figure94.3: ToF-SIMS images of various capsules. Capsule surfaces on which CH3O ions

and iron ions were detected were coloured blue and red, respectively. The compositions of

imaged samples are listed in Table 3.1: (a) is the control; (b) is dextrin capsules; (c) HPMC

capsules; (d) HPMC with 81% w/w sodium fumarate substitution; (e) HPMC with 50%

w/w sodium fumarate substitution; and (f) HPMC, but viewed at a greater magnification

(4000X versus 1000X).

42

4.2.4 Iodine Stability

Once the encapsulated iron particles are blended into iodized salt, the exposed iron readily reacts

with iodate in the presence of moisture [7]. Therefore, iodine loss can be used as an indirect

measure of encapsulation effectiveness.

After 6 months at 40OC, and 40-60% RH, the majority of dual fortified salt samples showed

good stability, retaining approximately 15% more iodine than the control (fortified with

unencapsulated ferrous fumarate). In a 6 month period, control samples retained ~60% of the

initial iodine while encapsulated samples had iodine retentions ranging from ~70 to ~85%.

Capsules that result in iodine retentions of 80% or greater, under these conditions, are considered

suitable for pilot-scale evaluations.

When stored at room temperature, and at lower humidity, in a five month period, samples

retained ~80% to ~90% of the iodine, an improvement from the ~75% iodine retention observed

with the control samples under these conditions.

Still, the observed iodine loss was greater than that in iodized salt alone, which retained ~85% in

a 6 month period. Even at room temperature, and lower humidity, plain iodized salt lost 3-15%

of the iodine after 5 months. These losses were caused by impurities in the salt, and cannot be

prevented by iron encapsulation [54].

This, combined with the EDTA leaching and TOF-SIMS results, suggest that encapsulation

greatly reduces the amount of exposed iron, but does not eliminate it entirely.

43

4.3 Formula Evaluation

4.3.1 Capsule Size and Shape

Molecular weight is a factor in determining particle formation pathway. Larger molecular

weight polymers have reduced molecular mobility, resulting in earlier shell formation, and

impacting particle morphology [34]. Molecular weight also affects particle size [34] [37].

Various NaCMC capsules were produced to investigate the effects of encapsulant molecular

weight; iodine retention was used as a measure of effectiveness. There was a statistically

relevant decrease in iodine retention for capsules formed with high molecular weight NaCMC

(Figure 4.4). The low molecular weight NaCMC also performed poorly after five months

(Figure 4.4). There appears to be an optimal value for polymer molecular weight which will

result in higher encapsulation effectiveness (Figure 4.4).

Though one formulation performed adequately, electron microscopy showed all formulations

produced collapsed capsules, indicating all evaluated molecular weights, under these conditions,

underwent wet shell particle formation. The collapsed capsules expose the enclosed iron. Other

unfavourable morphologies are also possible under this type of particle formation. There is a

need for an excipient to ensure dry shell particle formation occurs, producing uniformly round

capsules.

The performance of NaCMC 250cps shows that some samples, despite collapse, can perform

adequately. If the shell has ample tensile strength, it is possible that collapse does not cause a

breach in the capsule wall, and the iron will not be exposed. However, both particle size and

tensile strength increase with molecular weight [34] [59]. There appears to be a balance between

reducing the surface area of the capsules and increasing the tensile strength of the shells. The

use of excipients can prevent particle collapse altogether and produces rigid, solid capsules.

44

Figure104.4: Iodine stability in iodized salt blended with ferrous fumarate capsules of

various NaCMC molecular weights/viscosities after 5 months storage at 40OC and 40%-

60%RH. Nominal iron content of all samples: 27% w/w. Error bars: 1 standard deviation.

Sample size of 4.

4.3.2 The Effect of Excipient on Particle Shape and Size

The effect of an excipient on the particle formation process was investigated. The replacement

of 50% w/w of the polymeric encapsulating material (HPMC in this trial) with a white, readily

soluble (solubility >20% w/v in water) excipient - sodium fumarate - resulted in less spherical,

more irregular particles, and did not prevent particle collapse (Figure 4.5). Sodium fumarate,

with low molecular weight (molecular weight of 137 g/mol), is more mobile than the polymeric

material used as encapsulants; sodium fumarate is able to redistribute itself more readily, and

precipitated later, than HPMC. This difference in timing is the likely cause of the irregular

shape.

When replacing 81% w/w of the polymer with sodium fumarate more spherical particles were

formed. Though these capsules had some irregularities, the shape was more uniform than that of

45

samples containing less excipient. With this system, substituting greater amounts of sodium

fumarate for polymeric encapsulant resulted in unacceptable viscosities and particle settling

within the feed tube.

With a more viscous system, containing NaCMC 3000, sodium fumarate substitution of 95%

w/w was achieved. This resulted in regular, spherical particles, indicating that dry shell particle

formation can be achieved by use of high levels of sodium fumarate. The sodium fumarate

prevented capsule collapse and promoted the formation of regularly shaped particles (Figure

4.5). A)

B)

C)

D)

Figure 4.5:11SEM micrographs of capsules containing 16% iron with various excipient

levels; A) 0% sodium fumarate (magnification of 4,000X); B) 50% sodium fumarate

(magnification of 4,000X); C) 81% sodium fumarate (magnification of 4,000X) and D) 95%

sodium fumarate (magnification of 10,000X).

46

Sodium fumarate substitution of 95% resulted in much smaller particles due to the timing of

sodium fumarate precipitation (Figure 4.5). Being white, the addition of sodium fumarate

improves capsule colour.

When using iodine retention as a measure of encapsulation quality, all capsules with sodium

fumarate performed similarly well, all retaining more iodine on average than the base case

without sodium fumarate substitution (Figure 4.6). Though this material impacts the capsule

formation process, varying the level of excipient resulted in little change in iodine retention.

Sodium fumarate itself appears to improve the iodine retention, as using the same levels of

sodium chloride as an excipient resulted in much higher iodine loss in the first months of storage

(Figure 4.7). By the third month, samples containing a 50% sodium fumarate excipient

substitution retained ~81% of the initial iodine while samples containing equal amounts of

sodium chloride as an excipient retained only ~70% of the initial iodine (Figure 4.7). This could

be attributed to the fact that sodium fumarate is slightly basic and iodine reduction requires

protons. Aqueous sodium fumarate also reduces the solubility of ferrous fumarate; this may

reduce the amount of aqueous iron(II) around the salt, and prevent interactions.

47

Figure 4.6:12Iodine stability in iodized salt blended with ferrous fumarate capsules of

various excipient levels after 6 months storage at 40OC and 40%-60%RH. Nominal iron

content of all samples: 16% w/w. Samples feed at maximum ferrous fumarate throughput.

Error bars: 1 standard deviation. Sample size of 4.

Figure 4.7:13Iodine stability in iodized salt blended with ferrous fumarate capsules with

low molecular weight excipients after 5 months storage at 40OC and 40%-60%RH.

Nominal iron content of all samples: 16% w/w. Nominal Excipient content in coating

material: 50% w/w dry basis. Error bars: 1 standard deviation. Sample size of 4.

48

4.3.3 Iron Loading

Along with regular shaped capsules, it is desirable to obtain the highest iron loading possible,

reducing the amount of encapsulation material required. When varying the loading of ferrous

fumarate in dextrin capsules, statistically similar performances were observed until the later

months of storage (Figure 4.8). By the sixth month, samples with ferrous fumarate loadings

greater than 60% w/w had shown substantial iodine degradation (Figure 4.8). Therefore, for

effective fortification, ferrous fumarate levels should not exceed 60% w/w in the microcapsules.

Since statistically similar performances were achieved for the first two months of storage for all

iron loadings, the capsules must have similar levels of exposed iron. The greater degradation

observed in the fifth and sixth month of storage for the capsules of higher iron loadings can be

attributed to the thickness of the capsule walls. Samples with iron loadings of 70 and 80% have

less coating material per capsule, and thinner capsule walls. After a period of five months, the

water around the salt could start affecting the particle integrity of these thinner walled capsules;

the salt contains about 0.8% w/w water, and was stored in ~50% relative humidity. Thinner

walls also provide less resistance to iodate diffusion.

49


various nominal ferrous fumarate loadings after 6 months storage at 40OC and 40%-

60%RH. All capsules comprised of dextrin (DE7) and HPMC E15. Error bars: 1 standard

deviation. Sample size of 4.

4.3.4 Encapsulant Chemistry

The use of sodium fumarate as excipient resulted in dry shell particle formation, favourable

capsule shapes, improved the colour and increased the iodine retention, but a small amount of

polymeric encapsulant is still required to prevent particle settling. Varying the encapsulant

chemistry has shown that there are statistical differences between the performances of various

polymers in iodized salt (Figure 4.9). Similar performances were found in systems using HPMC,

dextrin and gum arabic during six months of evaluation (Figure 4.9). NaCMC capsules behaved

similarly until the fifth month of storage. After this time substantial iodine loss was observed.

NaCMC has been reported to increase the solubility of divalent metals [38]. This would result in

greater amounts of dissolved ferrous fumarate, potentially increasing the amount of exposed iron.

50


various common encapsulating materials after 6 months storage at 40OC and 40%-

60%RH. Nominal iron content of all samples: 16% w/w. Samples fed at one quarter of the

maximum ferrous fumarate throughput. Error bars: 1 standard deviation. Sample size of

4.

Though capsules created with dextrin performed well, the encapsulating system had to include

another material to obtain adequate viscosity. In order to produce dextrin capsules, HPMC was

added to increase the viscosity of the solution. The addition of a rheology modifier represents a

complicating step, which can be avoided by the use of other materials.

Samples encapsulated with HPMC had the highest average iodine retention after 6 months.

Accordingly, HPMC was selected, along with sodium fumarate, to encapsulate ferrous fumarate.

51

4.3.5 Capsule Colour

Despite using a white excipient, sodium fumarate, the capsules had an unacceptably red colour.

Adding titanium dioxide to the spray solution resulted in substantial colour improvement.

Unfortunately, this also resulted in a lower ferrous fumarate loading. As indicated by SEM

imaging, titanium dioxide improved capsule colour without affecting the particle formation

process (Figure 4.10).

Figure 4.10:16SEM Image of capsules containing 7% HPMC (E15), 67% sodium fumarate

and 27% TiO2 with a Ferrous fumarate loading of 6%.

Several formulations with various combinations of HPMC, sodium fumarate, titanium dioxide

and ferrous fumarate were evaluated (Table 4.4). A sample containing a substantial amount of

dextrin also met the visual screening criteria. A good balance between colour and ferrous

fumarate loading was obtained with the formulation containing 7% HPMC, 69% sodium

fumarate and 24% titanium dioxide.

Capsules containing 93% dextrin had similar colour and higher active material loading than

those containing 24% titanium dioxide (Table 4.4). Unfortunately, the dextrin capsules

underwent substantial colour change when added to moist salt making them unacceptable for use

in fortification. Dextrin absorbs water from the salt, and becomes transparent.

52

Table134.5: Colour Evaluation of Various Capsules

Nominal Iron

Content

(w/w)

Percent Red

Normalized to Signal

Strength

Control 33% 18.7%

Tamil Salt® 0% 0.6%

60% HPMC (E15) / 40% TiO2 16% 10.9%

5% HPMC (E15) / 95% sodium fumarate 7% 8.5%

93% Dextrin (DE7) / 7% HPMC (E15) 5% 4.6%

7% HPMC (E15) / 69% sodium fumarate / 24% TiO2 3% 4.9%

7% HPMC (E15) / 67% sodium fumarate / 27% TiO2 2% 1.5%

4.4 Adhesion

It is essential that the iron capsules adhere to the salt and maintain a uniform iron distribution

even after months of transport and storage. Capsule material, size and density affect the ability

of capsules to adhere to the salt. Microcapsules mostly adhere to salt particles within the

imperfections and coarse areas of the salt (Figure 4.11). When viewing double fortified salt

under SEM, large clusters of microcapsules were only found in the irregular areas; very few

capsules were present on the flat surfaces of the salt. Coarse, unrefined salt is expected to have

the largest number of imperfections, resulting in the best microcapsule adhesion.

53

When iron capsules are blended into the salt, some salt agglomerates are sheared apart; this

increases the surface area and the amount of imperfections to which the iron capsules can adhere.

Even after blending, there are a limited number of imperfections, and a limited number of

capsules that can fit into each imperfection. As a result, the number of capsules that can be

effectively carried by this salt is limited.

Smaller salt particles contain more surface area per unit mass. With the evaluated coarse salt,

smaller particles are created when crystals break during blending; this results in more surface

imperfections on the smaller broken shards, and allows for the adhesion of more iron capsules.

Figure 4.11:17Microcapsules adhering to salt

54

After sieving the double fortified Tamil Salt®, more than 35% of the colour appropriate capsules

were lost, while only a small amount of the salt (~2%) was lost due to particle break-down

(Table 4.4). Capsule adhesion to the larger salt particles is dependent on moisture. When water

content of the salt was adjusted to a level typical of inexpensive commercial coarse salt (2.4%),

92% of the iron capsules were retained, an improvement from the 65% retention observed using

salt with lower water content (0.8% and 1%) (Table 4.5).

55

Table 4.6:14Iron content in sieves after 5 min. of sifting double fortified Tamil salt with

1000ppm iron

Percent Iron Adhering to Salt after

Sieving Formulation

Mesh

30

Mesh

45

Mesh

60

Mesh

140

Lost

7% HPMC / 69% sodium fumarate / 24% TiO2** 51% 17% 9% 14% 8%

10% HPMC / 90% sodium fumarate 55% 13% 7% 12% 12%


HPMC 43% 20% 18% 4% 15%



7% HPMC / 69% sodium fumarate / 24% TiO2* 26% 12% 11% 16% 35%

7% HPMC / 69% sodium fumarate / 24% TiO2 34% 11% 6% 12% 37%



* Salts moisture content adjusted to approximately 1.0% w/w

** Salts moisture content adjusted to approximately 2.4% w/w

All other samples had a moisture content of approximately 0.8% w/w

Increasing the moisture content resulted in greater iron concentrations in all sieve fractions

(Figure 4.12). For all samples, the largest salt fraction (>595 µm) had iron concentrations lower

56

than the targetted1000 ppmiron; however the iron concentration in this fraction was substantially

improved by increasing the water content (Figure 4.12).

A) B)

Figure 4.12:18Sieve Fraction Evaluations. A) Iron Concentration in Sieve Fractions; B)

Salt Content in Sieve Fractions.

4.5 Bioavailability

Iron is largely absorbed in the duodenum [4]. Accordingly, it is critical that capsules breakdown

and release the ferrous fumarate prior to reaching this area of the gastrointestinal tract. To

confirm that the spray dried iron premix capsules were likely to be fully bioavailable their

solubility in simulated stomach acid (pH 1 HCl solution) was determined. The highest

concentration observed during dissolution was 34 mg/L; the solubility of ferrous fumarate in this

solution is greater than 100 mg/L.

57

With the most promising formulation, comprised of 9% ferrous fumarate, 6% HPMC (E15), 63%

sodium fumarate, and 22% TiO2, some 75% of the iron was dissolved 30 min., and more than

78% was released in the 2h required to transit through the stomach (Figure 4.12). These capsules

had significantly greater iron release profiles than other colour appropriate capsules; other

capsules only released approximately 40% of the contained iron after 120 min. (Figure 4.13).

The most promising formulation performed in a similar manner to capsules produced via

agglomeration and extrusion, which also resulted in a ~75% iron dissolution in 30 min. and a

~80% iron dissolution in 2h [58]. When ingested with other foods, the premix is expected to

perform even better as the presence of other food material has been shown to increase the

dissolution and subsequent absorption of iron [4].

Figure 4.13:19Relative apparent bioavailablity plot. Percentage iron dissolved versus time

in pH 1 HCl solution, 37OC, agitated at 90 rpm. Formula 1 is 9% ferrous fumarate / 6%

HPMC (E15) / 63% sodium fumarate / 22% TiO2; Formula 2 is 6% ferrous fumarate / 6%

HPMC (E15) / 63% sodium fumarate / 25% TiO2 and Formula 3 is 93% Dextrin (DE7) /

7% HPMC (E15).

Control

Formula 1

Formula 2

Formula 3

58

The sodium fumarate components of the capsules are expected to dissolve quickly as it is readily

soluble under these conditions. Titanium dioxide is not soluble and seems to hinder the

dissolution process. Formulation 2 contains approximately 15% more titanium dioxide than

formulation 1. This explains the significant difference in dissolution kinetics between the two

encapsulation systems.

Titanium dioxide also plays a role in the particles density, which is related to the buoyancy

forces suspending the capsules. The longer particles are suspended, the less likely they are to

agglomerated at the bottom of the vessel. This may also be a contributing factor in the poor

performance of the Formulation 2 capsules.

Capsules formed using Formulation 3 (containing mostly dextrin) tended to agglomerate and

float in the solution. This led to the poor dissolution of iron from these capsules. Dextrin is

expected to hydrolyze readily under these conditions, but the agglomeration hindered the wetting

process. This indicates that these capsules will be problematic when added to cooking water or

liquid foods. The floatation of the agglomerates result in the detection and ultimate separation of

these capsules from the food.

The formulation which consists of 9% ferrous fumarate, 6% HPMC (E15), 63% sodium

fumarate, and 22% TiO2 produced a premix of desirable colour with adequate in-vitro

bioavailability.

59

5 CONCLUSION

This study has demonstrated that ferrous fumarate microcapsules produced via spray drying,

when incorporated in iodized salt produce double fortified salt that is stable for 6 months at

elevated temperature and humidity and is therefore an attractive option for the fortification of

coarse, unrefined salt.

HPMC can create a digestible barrier that prevents iodine-iron interactions. Sodium fumarate,

used as an excipient in the film-forming polymer resulted in uniform, spherical particles, while

high-molecular weight polymers, without the excipient, typically collapsed, exposing the

enclosed ferrous fumarate. Titanium dioxide, added to mask the reddish colour of ferrous

fumarate, did not affect the formation of uniform spherical particles.

Iron loadings of up to 20% were attained. The capsules adhered to salt with moisture levels

typical of coarse, unrefined, commercial salt, and were undetectable on the double-fortified salt.

The in-vitro bioavailability of the iron on salt was acceptable: Nearly 80% of the iron dissolved

in simulated stomach acid within 2h.

The produced capsules have been shown to meet the criteria of being stable in iodized salt; able

to adhere to moist, unrefined, coarse salt; be too small to be detected; have appropriate colour;

and be digestible. This indicates that these capsules are appropriate for the fortification of

unrefined, coarse salt.

The best formulation resulted in rigid, uniform capsules. These capsules had a good balance

between colour and iron loading; the ability to adhere to moist coarse salt; and the best acid

dissolution kinetics. This formulation contained 9% ferrous fumarate, 6% HPMC (E15), 63%

60

sodium fumarate, and 22% TiO2 (dry basis). The ferrous fumarate premix was produced by an

easily-scaled, single-step process, and therefore its processing should be scaled-up in pilot-plant

tests, to produce sufficient quantities for efficacy and effectiveness tests.

61

6 RECOMMENDATIONS

1) Reformulate and produce capsules using a pilot scale spray drier with a rotary atomizer.

The atomizer used in this study restricts the use of suspended materials, limiting the

ferrous fumarate and titanium dioxide concentrations within the feed. It is possible that a

higher iron loading and better coloured capsules can be achieved using this type of

atomizer. Rotary atomizers are the most common type of atomizer used on industrial

scale spray driers.

2) Using the best formulations, measure the iodine stability in double fortified salt, based on

coarse, unrefined salt at various moisture levels. This would determine the highest level

of moisture in which stable dual fortified salt can be produced with these capsules. Data

determining the lowest level of moisture required for capsule adhesion can also be

obtained.

3) Investigate the use of other excipients. The trials with sodium chloride and sodium

fumarate demonstrated that the excipient can have an effect on the stability of iodate in

double fortified salt. GRAS buffer salts which provide a higher pH than sodium fumarate

may further reduce the iron solubility around the capsules, and the amount of aqueous

iron(II) around the salt, preventing iron-iodate interactions.

4) Investigate the use of zinc oxide as a pigment. Zinc oxide can be used to whiten the

capsules, and to address zinc deficiencies.

62

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[40] D. Walton and C. Mumford, “The morphology of spray-dried particles-the effect of process variables upon morphology of spray-dried particles”. Chemical Engineering Research and Design, 1999, vol. 77, pp. 442-460. [41] M. Elslamian, “Experimental and Theoretical Investigation of Micro- and Nano-powder Synthesis by Spray Pyrolysis and Drying”. 2006. Doctor of Philosophy Thesis, Department of Mechanical and Industrial Engineering, University of Toronto. [42] C.S. Handscomb and M. Kraft, “Simulating the structural evolution of droplets following shell formation”. Chemical Engineering Science, 2010, vol. 65, pp. 713–725. [43] M. Mezhericher, A. Levy, and I. Borde, “Theoretical Models of Single Droplet Drying Kinetics: A Review”. Drying Technology, 2010, vol. 28, pp. 278–293. [44] H. Kooijman, “A modification of the Stokes-Einstein Equation for Diffusivities in Dilute Binary Systems”. Ind. Eng. Chem. Res., 2002, vol. 41, pp. 3326-3328. [45] K. Goud, H. Desai and H.J. Park, “Recent Developments in Microencapsulation of Food Ingredients”. Drying Technology, 2005, vol. 23, pp. 1361–1394.

[46] J. Uhlemann, B. Schleifenbaum and H.J. Bertram. “Flavour Encapsulation Technologies: an overview including recent developments”. Perfumer and Flavorist, 2002, vol. 27, pp. 52-61.

[47] Y. Xu, C. Zhan, L.Fan, L.Wang and H. Zheng, “Preparation of dual crosslinked alginate-chitosan blend gel beads and in vitro controlled release in oral site-specific drug delivery system”. International Journal of Pharmaceutics, 2007, vol.3, no.36, pp.329-337. [48] M. George and T. Abraham, “pH sensitive alginate-guar gum hydrogel for the controlled delivery of protein drugs”. International Journal of Pharmaceutics, 2007, vol. 3, no. 35, pp. 123-129. [49] P. Gupta, K. Vermani and S. Garg. “Hydrogels: from controlled release to pH-responsive drug delivery”. Drug Discovery Today, 2002, vol. 7, no. 10. [50] A. Gharsallaoui, G. Roudaut, O. Chambin, A. Voilley and R. Saurel, “Applications of spray-drying in microencapsulation of food ingredients: An overview”. Food Research International, 2007, vol. 40, pp.1107–1121. [51] G. Castro, B. Panilaitis and D. Kaplan, “Emulsan, a tailorable biopolymer for controlled release”. Bioresource Technology, 2008, vol. 99, no. 11, pp. 4566-4571. [52] B. Youan, “Microencapsulation of superoxide dismutase into biodegradable microparticles by spray-drying”. Drug Delivery, 2004, vol.11, no.3, pp.209-214.

66

[53] W. Kolanowski, G. Laufenberg and B. Kunz, “Fish oil stabilisation by microencapsulation with modified cellulose”. International Journal of Food Sciences and Nutrition, 2004, vol. 55, no. 4, pp. 333 -/343. [54] L.L. Diosady, J.O. Alberti, M.G. Venkatesh Mannar and S. Fitzgerald, “Stability of Iodine in Iodized Salt Used for Correction of Iodine-Deficiency Disorders”. Food and Nutrition Bulletin, 1998, vol.29, no.3, pp. 239-249. [55] T. Oshinowo, L.L. Diosady, R. Yusufali and L. Laleye, “Stability of salt double fortified with ferrous fumarate and potassium iodate or iodide under storage and distribution conditions in Kenya”. Food and Nutrition Bulletin, 2004, vol. 25, pp. 264–270. [56] A. Harvey Jr., J. Smart and E. Amis, “Simultaneous spectrophotometric determination of Iron (II) and total iron with 1, 10-phenanthroline”. Analytical Chemistry, 1955, vol. 27, no.1. [57] Association of Official Analytical Chemists (AOAC), “Method 33.149”. Official Method of Analysis, 14th ed., 1984, AOAC, Arlington. [58] Y. Li, L.L. Diosady and A. Wesley, “Iron in vitro bioavailability and iodine storage stability in double-fortified salt”. Food and Nutrition Bulletin, 2009, vol. 30, no. 4, pp. 327-35. [59] M. Hallam, G. Pollard and I. Ward, “Relationship between tensile strength and molecular weight of highly drawn polyethylenes”. Journal of Material Science Letters, 1987, vol. 6, pp. 975-976.

67

8 APPENDICES

8.1 Iron Test Calibration Curve

The following is a sample calibration curve used for determining iron content:

Figure A20: Sample Calibration Curve

8.2 Ferrous fumarate Solubility in Acetic acid Solution

Solubility was determined by stirring 10g of ferrous fumarate to 50mL of acetic acid solution.

Solutions were stirred, and left covered over-night, allowing adequate time for ferrous fumarate

dissolution, as well as the settling of non-dissolved ferrous fumarate particles. Four samples

were drawn from the top, filtered and analyzed for iron content using the method described in

Section 3.

68

Table A15: Sample Solubility Calculation

Sample #

Acetic A

cid C

oncentration (%

v/v)

Volum

e of Sam

ple (mL)

Absorbance 1

Absorbance 2

Average

Absorbance

Solution Fe2+

Concentration

(g/L)

Absorbance 1

Absorbance 2

Average

Absorbance

Solution Fe C

oncentration (g/L)

Solution Fe3+

Concentration

(g/L)

1 2 5 1.25 1.26 1.258 3.77 1.26 1.25 1.253 4.18 0.40 2 2 5 1.41 1.42 1.416 4.21 1.30 1.30 1.302 4.30 0.09 3 2 5 1.19 1.19 1.190 3.56 1.18 1.19 1.187 3.95 0.39 4 2 5 1.36 1.37 1.367 4.11 1.36 1.35 1.352 4.51 0.40 Average: 3.82 Average: 4.23 0.32 Standard Deviation: 0.28 Standard Deviation: 0.24 0.15 Ferrous fumarate solubility: 12.9 g/L Percent Oxidized: 7.6 % w/w

Figure A21: Iron Oxidation versus Acetic Acid Concentration

69

Figure A22: Ferrous fumarate Solubility versus Acetic Acid concentration

Figure A23: Acetic Acid to Soluble Iron Ratio versus Acetic Acid Concentration

70

8.3 Ferrous fumarate Particle Size

Three grades of ferrous fumarate were evaluated for spray drying. The particle size distribution

was determined by use of Malvern Mastersizer. The only grade small enough to pass through

the spray driers atomizer had the following distribution:

Figure A24: Particle Size Distribution of Ferrous fumarate Powder

8.4 Process Oxidation

The oxidation during the process was measured for both suspended ferrous fumarate in water and

dissolved ferrous fumarate in 2% acetic acid solution by taking four samples and analyzing them

using the method described in Section 3. Water contents were determined gravimetrically.

71

Table A16: Oxidation and Water Content of Key Samples

Level of Oxidation

(% w/w)

Standard Deviation

on Level of

Oxidation (%)

Water Content

(% w/w)

Ferrous fumarate

powder 0.82 0.46 0.00

Sample produced by feeding ferrous fumarate:

dissolved in acetic

acid solution 53.8 12.9 18.3

suspended in water 3.93 1.01 3.93

dissolved with acetic

and ascorbic acid*

solution

0.29 0.14 3.68

*Samples prepared with ascorbic acid had a dark colour indicating that ferrous ascorbate was formed

8.5 Sample Dependant Process Conditions

Table A17: Dependant Operating Conditions for All Samples

Nominal

Iron

Content

(w/w) Encapsulating Material(s) (w/w)

Pressure

Drop Across

Filter

(mmHg)

Operating

Temperatur

e (OC)

Control

33% None N/A N/A


72

27% CMC 7M8 61 68

27% CMC 3000 63 71

27% CMC 15 79 72

27% CMC 1000 61 72

Various Iron Loadings

26% 20% Dextrin (DE7) / 80% HPMC (E15) 90 68

23% 56% Dextrin (DE7) / 44% HPMC (E15) 81 69

20% 69% Dextrin (DE7) / 31% HPMC (E15) 81 72

16% 80% Dextrin (DE7) / 20% HPMC (E15) 74 80

13% 87% Dextrin (DE7) / 13% HPMC (E15) 81 78


16% HPMC (E-15) 74 78

16% Gum Arabic 75 75

16% 80% Dextrin (DE7) / 20% HPMC (E15) 74 80

16% CMC 15 79 72


16% HPMC (E-15) 78 89

16% 75% HPMC (E15) / 25% sodium fumarate 76 91



Excipient Types

16% 50% HPMC (E15) / 50% sodium chloride 59 91



16% 20% HPMC (E15) / 80% TiO2 60 86

16% 60% HPMC (E15) / 40% TiO2 60 86

16% 80% HPMC (E15) / 20% TiO2 61 85

16% HPMC (E-15) 89 78




73



Table A3 Continued

3% 7% HPMC (E15) / 69% sodium fumarate / 24%

TiO2

55 77


TiO2

55 72


TiO2

55 71

74

8.6 Sample Yields and Selectivity

The samples selected for this study had the following mass yields and iron selectivity:

Table A18:Yield and Selectivity for All Samples

Nominal Iron

Content (w/w) Encapsulating Material(s) (w/w)

Actual Iron

Content (w/w)

Iron Selectivity

Total Mass

Yield (% of total

solids, dry basis)

Control

33% None 33% N/A N/A


27% CMC 7M8 31% 113% 68%

27% CMC 3000 32% 118% 66%

27% CMC 15 31% 113% 70%

27% CMC 1000 31% 113% 67%


26% 20% Dextrin (DE7) / 80% HPMC (E15) 32% 120% 75%

23% 56% Dextrin (DE7) / 44% HPMC (E15) 28% 121% 77%

20% 69% Dextrin (DE7) / 31% HPMC (E15) 22% 113% 64%

16% 80% Dextrin (DE7) / 20% HPMC (E15) 18% 112% 64%

13% 87% Dextrin (DE7) / 13% HPMC (E15) 16% 118% 68%

75


16% HPMC (E-15) 19% 115% 64%

16% Gum Arabic 19% 118% 67%

16% 80% Dextrin (DE7) / 20% HPMC (E15) 18% 112% 64%

16% CMC 15 19% 115% 64%


16% HPMC (E-15) 19% 115% 64%

16% 75% HPMC (E15) / 25% sodium fumarate 19% 117% 74%



Excipient Types

16% 50% HPMC (E15) / 50% sodium chloride 20% 121% 73%



16% 20% HPMC (E15) / 80% TiO2 20% 121% 82%

16% 60% HPMC (E15) / 40% TiO2 20% 121% 77%

16% 80% HPMC (E15) / 20% TiO2 20% 121% 70%

16% HPMC (E-15) 19% 115% 64%


76






TiO2

4% 121% 77%


TiO2

2% 120% 72%


TiO2

1% 118% 68%

8.7 Iodine Stability Data:

Prior to the iodine titration, the actual strength or concentration of the thiosulphate solution was

determined by titrating for a known amount of KIO3 (Table A5).

Table A5: Sample Thiosulphate Solution Standardization

Replicate

Volume Consumed to neutralize control

(mL)

Strength of Thiosulphate Solution used

(gIodine/mLsolution used) Average Strength SD RSD (%)

1 20.90 28.4

2 20.19 29.4

3 19.00 31.2

4 20.20 29.4

29.58 1.07 3.61

Salt from 12 bags of KenSalt® were mixed and sampled in 4 different spots to determine the

initial iodine content of the salt (Table A6).

77

Table A6: Determination of Initial Iodine Content

Location* Mass

Analyzed (g) Volume Consumed to

neutralize sample (mL) Iodine Content

(ppm) Mean Iodine

Content (ppm) SD RSD (%)

1 5 18.25 117

2 5 17.41 111

3 5 19.70 126

4 5 18.92 121

119 6 5.2

*1 was located at the bottom, right-hand corner of the bag 2 was located at the bottom, left-hand corner of the bag 3 was located at the top, right-hand corner of the bag 4 was located at the top, left-hand corner of the bag

Table A7: Iodine Retention for All Samples, 40OC, 40-60% RH (Months 1, 2 and 3)

Nominal Iron

Content (w/w) Encapsulating Material(s) (w/w)

Iron Retention Month 1

(% of Initial)


(% of Initial)


(% of Initial)

Control 0% Blank 99 ± 3 104 ± 1 93 ± 5

33% None 78 ± 4 70 ± 5 62 ± 3 Various Molecular Weights

27% CMC 7M8 99 ± 4 93 ± 4 93 ± 3 27% CMC 3000 102 ± 7 86 ± 3 88 ± 6 27% CMC 15 101 ± 3 93 ± 4 89 ± 4 27% CMC 1000 104 ± 3 97 ± 7 94 ± 1

Various Iron Loadings 26% 20% Dextrin (DE7) / 80% HPMC (E15) 100 ± 5 93 ± 3 90 ± 3 23% 56% Dextrin (DE7) / 44% HPMC (E15) 89 ± 5 90 ± 2 87 ± 1 20% 69% Dextrin (DE7) / 31% HPMC (E15) 94 ± 3 100 ± 3 80 ± 2 16% 80% Dextrin (DE7) / 20% HPMC (E15) 97 ± 4 96 ± 4 80 ± 6 13% 87% Dextrin (DE7) / 13% HPMC (E15) 100 ± 5 74 ± 7 85 ± 1

Various Encapsulants 16% HPMC (E-15) 100 ± 2 82 ± 4 85 ± 3 16% Gum Arabic 100 ± 2 82 ± 4 85 ± 2 16% 80% Dextrin (DE7) / 20% HPMC (E15) 97 ± 4 96 ± 4 80 ± 6 16% CMC 15 101 ± 4 91 ± 7 86 ± 5

Various Excipient Loadings 16% HPMC (E-15) 94 ± 2 80 ± 3 84 ± 3 16% 75% HPMC (E15) / 25% sodium fumarate 101 ± 3 86 ± 4 75 ± 3

78

16% 50% HPMC (E15) / 50% sodium fumarate 102 ± 8 84 ± 4 81 ± 7 16% 19% HPMC (E15) / 81% sodium fumarate 96 ± 3 88 ± 5 79 ± 3 16% 5% CMC 3000 / 95% sodium fumarate 93 ± 5 85 ± 7 76 ± 2

Excipient Types 16% 50% HPMC (E15) / 50% sodium chloride 80 ± 5 78 ± 4 70 ± 1 16% 50% HPMC (E15) / 50% sodium fumarate 102 ± 8 84 ± 4 81 ± 7

79

Table A8: Iodine Retention for All Samples, 40OC, 40-60% RH (Months 4, 5 and 6)

Nomin

al Iron

Conten

t (w/w) Encapsulating Material(s) (w/w)

Iron

Retention

Month 4

(% of

Initial)

Iron

Retention

Month 5

(% of

Initial)

Iron

Retention

Month 6

(% of

Initial)

Control

0% Blank 92 ± 3 87 ± 2 85 ± 1

33% None 63 ± 3 66 ± 3 60 ± 1


27% CMC 7M8 86 ± 3 88 ± 4 80 ± 5

27% CMC 3000 80 ± 3 69 ± 8 64 ± 8

27% CMC 15 88 ± 4 69 ± 2 76 ± 8

27% CMC 1000 95 ± 8 76 ± 4 60 ± 7


26% 20% Dextrin (DE7) / 80% HPMC (E15) 85 ± 5 79 ± 4 75 ± 5

23% 56% Dextrin (DE7) / 44% HPMC (E15) 83 ± 5 80 ± 3 73 ± 4

20% 69% Dextrin (DE7) / 31% HPMC (E15) 87 ± 6 81 ± 3 83 ± 2

16% 80% Dextrin (DE7) / 20% HPMC (E15) 79 ± 1 80 ± 6 82 ± 2

80

13% 87% Dextrin (DE7) / 13% HPMC (E15) 75 ± 3 79 ± 2 86 ± 3


16% HPMC (E-15) 87 ± 5 80 ± 1 83 ± 1

16% Gum Arabic 73 ± 4 67 ± 5 60 ± 2

16% 80% Dextrin (DE7) / 20% HPMC (E15) 79 ± 1 80 ± 6 82 ± 2

16% CMC 15 82 ± 3 74 ± 3 75 ± 5


16% HPMC (E-15) 86 ± 2 79 ± 4 74 ± 1

16% 75% HPMC (E15) / 25% sodium fumarate 82 ± 2 79 ± 4 79 ± 1



16% 5% CMC 3000 / 95% sodium fumarate 86 ± 2 85 ± 7 76 ± 2

Excipient Types

16% 50% HPMC (E15) / 50% sodium chloride 79 ± 5 68 ± 5 68 ± 5


81

Table A9: Iodine Retention for All Samples, room temperature, ~20% RH

Nominal

Iron

Content


Iron

Retention

Month 1

(% of

Initial)

Iron

Retention

Month 2

(% of

Initial)

Iron

Retention

Month 3

(% of

Initial)

Control

0% Blank 105±2 101±2 90±2

33% None 103±5 89±2 76±3


16% HPMC (E-15) 104±3 104±8 97±5

16% 80% Dextrin (DE7) / 20% HPMC (E15) 100±5 104±8 100±2


16% HPMC (E-15) 104±3 104±8 97±5

16% 50% HPMC (E15) / 50% sodium fumarate 97±7 96±1 99±3


Excipient Types

16% 50% HPMC (E15) / 50% sodium chloride 102±7 100±1 83±4


82

Nominal

Iron

Content


Iron

Retention

Month 4

(% of

Initial)

Iron

Retention

Month 5

(% of

Initial)

Iron

Retention

Month 6

(% of

Initial)

Control

0% Blank 89±6 91±6 83±7

33% None 70±2 75±2 70±2


16% HPMC (E-15) 90±7 85±6 85±3

16% 80% Dextrin (DE7) / 20% HPMC (E15) 92±2 88±3 86±5


16% HPMC (E-15) 90±7 85±6 85±3



Excipient Types

16% 50% HPMC (E15) / 50% sodium chloride 79±1 74±3 72±6


83

8.8 EDTA Leaching

Three methods to quantify the iron content were evaluated: Spectrophotometry as described by

Oshinowo et al. [55] [56]; Atomic Absorption Spectrometry (AAS) used directly on the samples;

and ICP as described in Section 3.

8.8.1 EDTA leaching iron quantification via Spectrophotometry

The iron-1, 10-phenanthroline complex is the species detected by Spectrophotometry. The

EDTA and 1, 10-phenanthroline were competing iron ligands. This resulted in slowed 1, 10-

phenanthroline complexation kinetics (Figure A6). The time required for equilibrium is too long

for this method to be practical.

Figure A25: Iron Detected by Spectrophotometry from samples collected during a timed

EDTA leach of ferrous fumarate particles.

84

8.8.2 EDTA leaching iron quantification via AAS

EDTA suppresses the iron signal when using AAS. The result is low absorption and high

variability in sample reading (Table A10).

Table A10: Iron Detected by AAS from iron-EDTA Solution

Actual Concentration (mg/L)

Average Level Detected from four readings (mg/L)

Change in Detected Level (mg/L)

Standard Deviation from four readings (mg/L)

Standard Error

0.0 -3.1 - 0.39 13% 1.1 -2.8 0.3 0.43 15% 4.1 -2.5 0.3 0.36 14% 6.2 -2.1 0.4 0.25 12% 9.7 -1.6 0.5 0.39 24%

8.8.3 Leaching iron quantification via ICP

ICP showed consistent results for detecting iron in EDTA complexes (Figure A7).

Figure A26: Sample Calibration Curve for Leached Iron Detection via ICP for various

analytes

85

Table A11: Sample Calibration Date Collected prior to each Scan

Sample Analyte Intensity Intensity Average

Intensity Standard Deviation

60 60 576

Fe 238.204

576

318 298

-264 -269 585

Fe 239.562

585

159 492

44 50 686

Calibration Blank

Fe 259.939

686

366 369

46538 49135 35067

Fe 238.204

35067

41452 7449

54816 57903 41781

Fe 239.562

41781

49070 8511

53962 58222 39479

1 mg/L

Fe 259.939

39479

47786 9748

102725 108550 75871

Fe 238.204

75871

90754 17349

121216 128155 90340

Fe 239.562

90340

107513 20031

119384 128935 86114

2 mg/L

Fe 259.939

86114

105137 22309

210095 222898 164021

4 mg/L

Fe 238.204

164021

190259 30744

86

Table A11 Continued

247890 263188 195518

Fe 239.562

195518

225528 35212

244143 264521 186483

Fe 259.939

186483

220407 40046

253896 265525 194395

Fe 238.204

194395

227053 38007

299508 313553 231703

Fe 239.562

231703

269117 43581

294776 315206 222155

5 mg/L

Fe 259.939

222155

263573 48547

525015 551333 415269

Fe 238.204

415269

476721 71768

622391 652186 566441 82707

495593 Fe 239.562

495593

609158 654892 474689

10 mg/L

Fe 259.939

474689

553357 92737

87

Data was collected using ICP:

Table A12: EDTA Leach data

Nominal Iron Content (w/w)

/ Encapsulating

Material(s)

Amount Diluted for analyzing (initial-to-

final volume ratio) Analyte

Intensity Average

Average Iron Concentration dissolved in the leach solution ((mg/L)/giron

added)

Iron Leached Total (%)

Standard error (%)

Fe 238.204 677542 Fe 239.562 801399

33% Iron /

None 2 : 25

Fe 259.939 779562 Fe 238.204 690316 Fe 239.562 816988

33% Iron /

None 2 : 25

Fe 259.939 797787 Fe 238.204 672159 Fe 239.562 795520

33% Iron /

None 2 : 25

Fe 259.939 777122

8433 21 2

Fe 238.204 138674 Fe 239.562 163467

27% Iron /

CMC 7M8 1:10

Fe 259.939 164245 Fe 238.204 109662 Fe 239.562 129131

27% Iron /

CMC 7M8 1:10

Fe 259.939 129832 Fe 238.204 108241 Fe 239.562 127616

27% Iron /

CMC 7M8 1:10

Fe 259.939 128186

4706 12 2

Fe 238.204 163655 Fe 239.562 192830

27% Iron /

CMC 1000 2:25

Fe 259.939 188390 Fe 238.204 153490 Fe 239.562 180924

27% Iron /

CMC 1000 2:25

Fe 259.939 176379 Fe 238.204 135008 Fe 239.562 159185

27% Iron /

CMC 1000 2:25

Fe 259.939 155407

5069 13 0

Fe 238.204 188131 Fe 239.562 221911

27% Iron /

CMC 3000 2:25

Fe 259.939 216627 5174 13 1

88

Table A12 Continued

Fe 238.204 125412 Fe 239.562 147820

27% Iron /

CMC 3000 2:25

Fe 259.939 144208 Fe 238.204 182762 Fe 239.562 215418

27% Iron /

CMC 3000 2:25

Fe 259.939 210035

Fe 238.204 266271 Fe 239.562 313961

27% Iron /

CMC 15 1:5

Fe 259.939 315218 Fe 238.204 112512 Fe 239.562 132553

27% Iron /

CMC 15 1:5

Fe 259.939 133150 Fe 238.204 45122 Fe 239.562 53131

27% Iron /

CMC 15 1:5

Fe 259.939 53431

4620 12 1

Fe 238.204 226740 Fe 239.562 267376

26% Iron /

20% Dextrin / 80% HPMC

1:25 Fe 259.939 268691

Fe 238.204 102100 Fe 239.562 121413

26% Iron /


1:25 Fe 259.939 116747

Fe 238.204 105518 Fe 239.562 125547

26% Iron /


1:25 Fe 259.939 119609

4767 12 1

Fe 238.204 227633 Fe 239.562 271220

23% Iron /


1:10 Fe 259.939 259528 Fe 238.204 229305 Fe 239.562 273233

23% Iron /


1:10 Fe 259.939 260132 Fe 238.204 286182 Fe 239.562 337810

23% Iron /


1:10 Fe 259.939 339444

Fe 238.204 278129 Fe 239.562 328353

23% Iron /


1:10 Fe 259.939 330069

3871 10 1

89

Table A12 Continued

Fe 238.204 212674 Fe 239.562 250538

20% Iron /


1:10 Fe 259.939 244443 Fe 238.204 235951 Fe 239.562 278235

20% Iron /


1:10 Fe 259.939 271520 Fe 238.204 163534 Fe 239.562 192612

20% Iron /


1:10 Fe 259.939 188143 Fe 238.204 262742 Fe 239.562 310122

20% Iron /


1:10 Fe 259.939 302195

4115 10 1

Fe 238.204 278765 Fe 239.562 329134

16% Iron /


1:5 Fe 259.939 321132 Fe 238.204 149188 Fe 239.562 175898

16% Iron /


1:10 Fe 259.939 171563 Fe 238.204 254151 Fe 239.562 299679

16% Iron /


1:10 Fe 259.939 291845 Fe 238.204 177571 Fe 239.562 209404

20% Iron /


1:10 Fe 259.939 204478

3512 9 2

Fe 238.204 139077 Fe 239.562 164158

13% Iron /


1:10 Fe 259.939 159997

Fe 238.204 146239 Fe 239.562 172507

13% Iron /


1:10 Fe 259.939 168167 Fe 238.204 123310 Fe 239.562 145321

13% Iron /


1:10 Fe 259.939 141841

3495 9 1

90

Table A12 Continued

Fe 238.204 147503 Fe 239.562 174100

13% Iron /


1:10 Fe 259.939 169841

Fe 238.204 142311 Fe 239.562 167789

16% Iron /

HPMC 1:5

Fe 259.939 163558 Fe 238.204 71662 Fe 239.562 85247

16% Iron /

HPMC 1:5

Fe 259.939 81462 Fe 238.204 87876 Fe 239.562 103558

16% Iron /

HPMC 1:5

Fe 259.939 104137 Fe 238.204 67125 Fe 239.562 79781

16% Iron /

HPMC 1:5

Fe 259.939 76494

2598 7 1

Fe 238.204 283778 Fe 239.562 338143

16% Iron /

Gum Arabic 1:5

Fe 259.939 321391 Fe 238.204 85540 Fe 239.562 100727

16% Iron /

Gum Arabic 2:25

Fe 259.939 98394 Fe 238.204 72233 Fe 239.562 85958

16% Iron /

Gum Arabic 2:25

Fe 259.939 81840 Fe 238.204 149628 Fe 239.562 150184

16% Iron /

Gum Arabic 2:25

Fe 259.939 176407

3940 10 1

Fe 238.204 35048 Fe 239.562 41243

16% Iron /

CMC 15 2:25

Fe 259.939 41481 Fe 238.204 62602 Fe 239.562 73727

16% Iron /

CMC 15 2:25

Fe 259.939 74180 Fe 238.204 38944 Fe 239.562 45962

16% Iron /

CMC 15 2:25

Fe 259.939 46182

2884 8 1

91

8.9 Salt Moisture Content

Table A13: Moisture Content Calculations

Sample

Mass of Salt Before

Drying (g)

Mass of Salt After

Drying (g)

Water Content (%

w/w)

Tamil Salt 5.00 4.96 0.8

Tamil salt with

added water to

achieve 1% water

content

5.00 4.95 1.0

Tamil Salt with

added water to

achieve 2.5% water

content

5.00 4.88 2.4

92

8.10 Tamil Salt Particle Distribution prior to Blending

Figure A27: Particle Size Distribution for Tamil Salt

93

8.11 Iron Distribution for all Samples Evaluated for Adhesion

Table A14: Iron Content of Sieve Fractions

Percent Iron Adhering to Salt after

Sieving Formulation

Mesh

30

Mesh

45

Mesh

60

Mesh

140

Lost

7% HPMC / 69% sodium fumarate / 24% TiO2** 51% 17% 9% 14% 8%



HPMC 43% 20% 18% 4% 15%



60% HPMC / 40% TiO2 44% 9% 9% 4% 34%

7% HPMC / 69% sodium fumarate / 24% TiO2* 26% 12% 11% 16% 35%

20% HPMC / 80% TiO2 42% 13% 8% 2% 36%

80% HPMC / 20% TiO2 46% 10% 4% 3% 37%





* Salts moisture content adjusted to approximately 1.0% w/w

** Salts moisture content adjusted to approximately 2.4% w/w

All other samples had a moisture content of approximately 0.8% w/w

94

8.12 Pictures of Salt Analyzed for Colour

Table A15: Colour Evaluation

Samples Evaluated for Colour

Iron

Content Encapsulants Photograph

Red

Signal

Green

Signal

Blue

Signal

0% N/A

141 136 141

33% None (Control)

92 62 47

16% HPMC (E-15) 115 87 66

16% 20% HPMC (E15) / 80% TiO2

145 116 96

7% 5% HPMC (E15) / 95%

sodium fumarate 152 130 108

5% 4% HPMC (E15) / 96%

dextrin

130 119 108

3% 7% HPMC (E15) / 69%

sodium fumarate / 24% TiO2

154 139 128

2% 7% HPMC (E15) / 67%

sodium fumarate / 27% TiO2

154 139 126

1% 6% HPMC (E15) / 65%

sodium fumarate / 29% TiO2 163 155 157

95

8.13 In vitro Bioavailability Data

The maximum absorbance was calculated based on the amount of iron added for each sample

(Table A16).

Table A16: Maximum Values for in vitro bioavailablity

Iron Content Encapsulants

Mass iron Added (mg)

Highest concentration in Solution (mg/L)

Highest concentration in test solution (mg/L)

Maximum Absorbance

33% None (Control) 5 9.0 1.08 0.17

5% 4% HPMC (E15) / 96% dextrin 4 7.6 0.91 0.15


6 12.8 1.54 0.25


3 5.7 0.69 0.11

96

Table A16: In vitro Bioavailability Data

15 Minutes 30 Minutes 45 Minutes

Absorbance Absorbance Absorbance


Reading 1

Reading 2

Average

Percent

Dissolved

Reading 1

Reading 2

Average

Percent

Dissolved

Reading 1

Reading 2

Average

Percent

Dissolved

33% None (Control) 0.12 0.12 0.12 71% 0.17 0.17 0.17 100% 0.18 0.18 0.18 103%

3%

7% HPMC (E15) / 69% sodium fumarate / 24% TiO2

0.14 0.14 0.14 58% 0.19 0.19 0.19 75% 0.19 0.19 0.19 75%

1%


0.03 0.03 0.03 28% 0.03 0.03 0.03 31% 0.05 0.05 0.05 49%

5% 4% HPMC (E15) / 96% dextrin

0.07 0.07 0.07 45% 0.06 0.06 0.06 42% 0.06 0.06 0.06 42%

60 Minutes 90 Minutes 120 Minutes

Absorbance Absorbance Absorbance


Reading 1

Reading 2

Average

Percent

Dissolved

Reading 1

Reading 2

Average

Percent

Dissolved

Reading 1

Reading 2

Average

Percent

Dissolved

33% None (Control) 0.17 0.17 0.17 100% 0.18 0.18 0.18 101% 0.18 0.18 0.18 101%

3%


0.19 0.19 0.19 75% 0.19 0.19 0.19 76% 0.19 0.19 0.19 78%

1%


0.05 0.05 0.05 43% 0.06 0.06 0.06 51% 0.06 0.06 0.06 55%

5% 4% HPMC (E15) / 96% dextrin

0.06 0.06 0.06 41% 0.06 0.06 0.06 43% 0.06 0.06 0.06 42%

97

8.14 Viscosity Data

To ensure the evaluated materials had substantially different molecular weights, their

viscosity in 2% solution was measured using a U-tube viscometer (Figure

A28).

Figure A28: Viscometer Readings for CMC family of materials at concentrations of 2 %

w/w in water

spray drying based technologies for the double ......spray drying based technologies for the double...

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