KRILL OIL CONCENTRATE the Phospholipid Factor That Sets Krill Oil Apart

Lena Burri, Ph.D KRILL OIL CONCENTRATE The phospholipid factor that sets krill oil apart

The next generation Omega-3 Phospholipid product from Antarctic krill Foreword

To effectively maintain our bodily functions and to keep cells and body in balance, we need a continuous supply of nutrients. Whereas in diseased conditions, it is also important to address the fluctuations often seen in nutrient uptake and utilization. There is excitement and growing evidence surrounding krill oil as an important nutrient based on its major components:

• Long-chain omega-3 fatty acids (EPA/DHA) • Phospholipids and specifically its choline via its phosphatidylcholine- “lecithin” component

Krill oil is unique in that it is a combination of these naturally occurring nutrients. And when ingested, it delivers EPA/DHA, phospholipids, and choline to the body, where they work individually and in combination; i) as vital constituents in the structure and functioning of cells, ii) for assisting in the balancing of bodily functions, and iii) to address their deficiencies in health conditions related to the heart, brain, inflammation, immunity, liver, etc. Omega-3 fatty acids are one of the most recognized and studied compounds with more than 20.000 scientific papers published. Research has shown that when omega-3 fatty acids from marine sources, such as fish and krill, are ingested, they incorporate into cells to affect cell structure and function. Unlike fish oil, krill oil not only delivers EPA and DHA, but also phospholipids into cells, where they also exert their effects on re- modeling cells’ structures, fluidity, and functions. With this, and their ability to affect cholesterol metabolism, phospholipids may trigger downstream pathways and mechanisms into meaningful health benefits. This book identifies some of the research that addresses several of these pathways and mechanisms important for conditions of e.g. the heart, brain, and liver. It details

2 KRILL OIL CONCENTRATE the above-mentioned aspects of krill oil, its components, and how science and research points to its beneficial health effects while drawing comparisons to fish oil where applicable. The latest product development from Antarctic krill is also introduced. It is a highly concentrated and purified product developed on the premise that a higher omega-3 phospholipid concentration may deliver better health benefits across various health conditions than its predecessors. Thus, the next generation omega-3 phospholipid product is a krill oil concentrate with more omega-3 fatty acids being efficiently delivered to tissues via more phospholipids high in choline.

Lena Burri, Ph.D Aker BioMarine Antarctic AS Oslo, Norway

3 KRILL OIL CONCENTRATE Lena Burri, Ph.D

Lena Burri, Ph.D. has been involved in fundamental research, has published peer-reviewed articles in leading journals, and contributed review articles, as well as book chapters on subjects including omega-3 fatty acids. Lena earned her Master of Science from the University of Basel (Switzerland) and her Ph.D. at the Ludwig Institute for Cancer Research (Switzerland). Her post-doctoral education included stays at Melbourne University (Australia), University of British Columbia (Canada), and University of Bergen (Norway). She now works as R&D Director for Aker BioMarine Antarctic AS.

ISBN: 978-82-690452-0-8 All rights reserved. No part of this book may be reproduced in any form or by any means electronic or mechanical, including photocopying, recording or by any information storage and retrieval system without permission in writing from the copyright holder and the publisher.

4 KRILL OIL CONCENTRATE Summary

• Krill oil is extracted from Antarctic krill.

• Antarctic krill lives in the Southern Ocean around Antarctica.

• The Convention of the Conservation of Antarctic Marine Living Resources (CCAMLR), an international treaty, regulates krill harvesting in a sustainable way. In Area 48 in the Southern Ocean, which is the only area where krill fisheries can operate, the krill industry is allowed to catch one per cent of the estimated 60 million tons of krill.

• Krill oil is rich in omega-3 phospholipids and therefore a good source o f both omega-3 fatty acids and choline from the phospholipid head group.

• What differentiates krill oil from fish oil is the molecular form, in which the omega-3 fatty acids are bound to, namely phospholipids in krill oil and triglycerides in fish oil.

• The long-chain omega-3 fatty acids EPA (eicosapentaenoic acid) and DHA (docosahexaenoic acid) promote healthy heart, brain and visual function and contribute to reducing and maintaining normal levels of inflammation, blood triglycerides, and blood pressure.

• Choline is vital for many biological functions (e.g. membrane structure, nerve signaling, methylation reactions, lipid transport, etc.) and deficiency can lead to fatty liver, muscle damage, atherosclerosis, etc.

• The typical Western diet contains inadequate amounts of omega-3 fatty acids and choline. This contributes to the approximately 90% of the American population who are deficient in omega-3 fatty acids and choline [1,2].

• Krill oil supplementation can help to increase levels of omega-3 fatty acids and choline in the body, thus reducing the deficiencies of these important nutriments.

• Several clinical studies have shown health benefits for supplementation with krill oil.

5 KRILL OIL CONCENTRATE Table of Contents

Summary...... 5 Krill background...... 8 Krill oil concentrate...... 11 Latest krill oil studies...... 15 Sport...... 15 Skin...... 18 Phospholipids...... 22 What is a phospholipid?...... 22 Functions of phospholipids in the body...... 23 Cell membranes...... 23 Lipid transport and cholesterol metabolism...... 24 Uptake of phospholipids...... 26 Phospholipid sources...... 26 Phospholipid deficiency...... 27 Phospholipids in health and disease...... 27 Liver...... 27 Brain...... 30 Choline...... 31 What is choline?...... 31 Functions of choline in the body...... 32 Nerve signaling...... 32 Cell signaling...... 32 Methyl donor...... 32 Water balance...... 33 Uptake of choline salt versus phosphatidylcholine...... 33

6 KRILL OIL CONCENTRATE Choline deficiency...... 34 Choline in health and disease...... 35 Brain...... 35 Sport...... 36 Omega-3 fatty acids...... 37 What are omega-3 fatty acids?...... 37 Functions of omega-3 fatty acids...... 37 Membranes...... 38 Gene transcription and enzyme activity...... 38 Eicosanoids...... 39 Endocannabinoids...... 39 Omega-3 deficiency...... 40 Omega-3 fatty acids in health and disease...... 41 Heart...... 41 Brain...... 42 Krill oil: Phospholipids, choline, and omega-3 fatty acids all in one...... 44 Why krill oil?...... 44 Uptake of krill oil in the body...... 45 Krill oil in health and disease...... 46 Omega-3 Index...... 46 Conclusion...... 51 Acknowledgements...... 51 References...... 52

7 KRILL OIL CONCENTRATE Krill Background

Euphausia superba (E. superba) is the scien- tures are one of the world’s largest swarm- tific name for ‘Antarctic krill’. ing animals. They often aggregate in large, dense swarms stretching for tens of kilome- The shrimp-like crustaceans from the Eu- ters and measuring 30 meters deep. phausiacea family are commonly referred They tend to swarm and migrate to as ‘krill’ and consist of 86 species [3]. vertically as methods of avoiding attacks Euphausia superba (E. superba), also called from predators. They also swim up to the ‘Antarctic krill’, is the most dominant krill sea’s surface to feed and reproduce main- species in the pristine oceans surrounding ly during the night and then swiftly swim Antarctica [4]. Even though Antarctic krill down into deeper waters to avoid predators, are only about the size of your smallest fin- such as birds, penguins, seals, squid, fish or ger, they are a keystone species of the Ant- whales. When larger, swarm-hunting animals arctic marine ecosystem. Krill is considered attack, they quickly scatter by swimming to be at the bottom of the food chain, since backwards (as fast as 60 cm per second) in they feed on phytoplankton and are food for all directions to confuse the predator. An- many marine animals, such as whales, seals, other defense from smaller attackers is to penguins, squid, and fish. Of all multi-cellular leave their exoskeleton behind as a food dis- animal species on Earth, Antarctic krill is the traction. most abundant species with one of the largest biomasses of around 500 million metric Some krill swarms may consist only of tons. young krill or either females or males.

Compared with other marine life, Antarctic Krill are bioluminescent, meaning krill gathers in the largest groups [5] and that their bodies can emit light by chemical also has the most potent known digestive reactions involving oxygen molecules. These enzymes on Earth [6]. reactions take place in photocytes, lumi- nescent cells assembled into light organs. These small pinkish-red transparent crea- Krill use muscle contraction and relaxation

8 KRILL OIL CONCENTRATE Krill Background

Antarctic krill is found in the Southern Ocean

Krill swarm

for the regulation of the intensity of the light. dark brown compound eyes, and like with in- When special closure muscles relax, the light is sects, they consist of many little eyes. This emitted, which is suspected to be due to more gives krill the ability to see in many differ- oxygenated blood reaching the photocytes. ent directions at the same time, which helps It is believed that this light emission them to locate food and detect predators. is important for recognizing members of the However, in comparison to human eyes that same species and keeping them together in can see in only one direction, the vision quali- a swarm [7]. Though suspected, but unknown, ty of each little eye in krill is lower. this feature may also be valuable in defending Krill can survive up to 200 days with- against detection from predators or in mating. out food by shrinking and using their body’s Another characteristic of krill are their large bio-material for storing energy in the form of

9 KRILL OIL CONCENTRATE Antarctic krill’s biophysical make cialized filtering apparatus in their front up is important for their survival. legs to help them feed on the tiniest of They withstand the fridge cold wa- plants (phytoplankton and zooplankton) ters of the Antarctic and expend mostly in the summer, and algae from large amounts of energy constantly under sheets of ice in the winter, when swimming up and down the depths otherwise food may be scarce. More- and lengths of the Antarctic Ocean. over, krill can live up to seven years. Fe- If Antarctic krill stops swimming, males are sexually mature by two years their higher than water body density of age and males by three [9]. Krill shed would make them sink [8]. their outgrown exoskeleton known as a moulting process. They moult in order to lipids. Some krill species hold their lipid grow, while their new shell is still soft and reserves in wax esters and others pre- in times when they shrink due to lack of dominantly in triglycerides. However, food[10]. members of the Euphausiid family are the only known species in which phospholipids are used as their energy depot. In particular, Antarctic E. Superba krill uses phosphatidylcholine containing omega-3 fatty acids as their lipid reserve [3]. E. superba krill uses its spe-

Euphausia superba (Antartic krill)

10 KRILL OIL CONCENTRATE Krill Oil Concentrate

Krill oil is a pure, natural source of can be taken with krill oil compared to eicosapentaenoic (EPA) and docosa- fish oil in order to get the same amount hexaenoic (DHA) omega-3 fatty acids, of omega-3 health benefits. Moreover, phospholipids (with choline), and astax- there is evidence which points to phos- anthin. This combination of important pholipids (and choline) itself having nutrients extracted from Antarctic krill many health benefits. distinguishes it from fish oil and is the basis for krill oil’s uniqueness. Its natu- Astaxanthin contained in krill is rally occurring antioxidant astaxanthin a highly potent antioxidant and is responsible for the dark red color of accounts for krill oil’s red color krill oil. In contrast, fish oil contains no [11]. It assists in keeping the ome- astaxanthin, and their omega-3 fatty ac- ga-3 fatty acids in krill oil stable. ids are incorporated into triglycerides. In cells, it provides protection There is increasing evidence against free radical attack and that the differences between the mo- it has been shown to normalize lecular forms of omega-3 fatty acids oxidative stress in persons such (triglycerides and ethyl-esters in fish as those that are smokers or are oil, and phospholipids in krill oil) are im- overweight. As a result, astaxan- portant. The phospholipid form (from thin has been linked to health bene- krill oil) has been shown to facilitate fits such as anti-inflammatory and the incorporation of omega-3 fatty pain-relieving effects, faster re- acids into tissues in a more effective covery from exercise, UV light pro- and efficient manner compared to tri- tection in the skin [12], aging and glycerides and ethyl-esters (from fish age-related diseases, liver, heart, oil) [14]. Therefore, it is thought that a eye, joint, and prostate health [13]. lesser amount of omega-3 fatty acids

11 KRILL OIL CONCENTRATE Phospholipids have two fatty acids Cellular dysfunction has been linked to bound to a glycerol backbone, while health conditions of the heart, brain, triglycerides, or triacylglycerols, are a liver, joints, etc. Therefore, it is thought combination of three fatty acids with a that providing more phospholipids and glycerol backbone. Fatty acid ethyl es- omega-3 fatty acids with krill oil con- ters are derived by a chemical process centrate will result in better cell func- to exchange the glycerol backbone of a tioning and ultimately better health triglyceride with ethanol, an alcohol. benefits compared to other krill and fish oil products. Each of krill oil’s major components Indeed, the importance of the (phospholipids, choline, and omega-3 amount of phospholipids in krill oil was fatty acids) have shown to have effects shown in a comparison study of two krill in the body’s cells and tissues. This is oils containing either a low (600 mg; LPL) thought to be the basis of its health or a high (1200 mg; HPL) phospholipid benefits in many bodily systems and amount with the same omega-3 fatty acid health conditions. content of 600 mg [15]. LPL, HPL, or con- Fatty acids are important nu- trol oils were given to healthy male and tritional elements of cells, and in partic- female volunteers for 4 weeks. At study ular phospholipids and omega-3 fatty end, both plasma and red blood cell fatty acids are essential in cell membrane acid compositions were compared for the structure and function. They also play different treatment groups. While both an important role in the formulation of the LPL and HPL groups showed signifi- lipoproteins, while their metabolites cantly increased plasma omega-3 levels serve as vital molecules within path- when compared to the control group, ways of the body. For example, they can there was no statistical difference be- modify molecules involved in inflam- tween the LPL and HPL groups. However, mation (cytokines), eicosanoids, gene when incorporation into membranes was expression, and plasma triglyceride compared, then LPL only changed EPA synthesis, to name a few. Both omega-3 in red blood cells, whereas HPL changed fatty acids and phospholipid deficien- both EPA and DHA. And the red blood cell cies can be associated with damaged membrane EPA and DHA incorporation cell structure and decreased fluidity, was significantly higher in the HPL group which can result in cell dysfunction. when compared to the LPL participants.

12 KRILL OIL CONCENTRATE KRILL OIL FISH OIL

Omega-3 fatty Omega-3fatty Phospholipids Astaxanthin Triglycerides acids EPA/DHA acids EPA/DHA

Phosphatidylcholine

Choline

Major components of krill oil and fish oil

Table 1: Typical content values (g/100g oil) for krill oil and krill oil concentrate.

Composition Krill oil Krill oil concentrate

Total phospholipids ≥40 ≥56 Choline ≥5 ≥7 Total omega-3 fatty acids 24 ≥27 EPA ≥12 ≥15 DHA ≥5,5 ≥7

Increasing the amount of phospholipids uct advancement of a highly purified in krill oil increases the incorporation of and concentrated version of krill oil EPA and DHA into membranes [15]. was developed. It was made possible by a specialized ion-exchange tech Since incorporation into red blood cells nology, Flexitech™, that also reduces relates to long-term and tissue absorp- the TMA/TMAO (an osmolyte and pro- tion in general [16], and therefore health tein stabilizer) and the salt content of benefits, an increased amount of ome- the oil. This results in a product with a ga-3 phospholipids will improve the ef- concentrated amount of omega-3 fatty fects of krill oil. acids, phospholipids, and astaxanthin, Hence, to profit from increased mem- which is almost taste- and smell-free brane incorporation rates, a new prod- (see Table 1).

13 KRILL OIL CONCENTRATE Sourced from the pristine waters of the Southern Ocean around Antarctica and due to its low position in the food chain, krill oil is virtually free of toxins and heavy metals. Superba™ krill oil is certified by the Marine Steward- ship Council (MSC) as being sustainable and 100% traceable, from sea to shelf, with the GPS coordinates to prove it.

Trimethylamine (TMA) has a ‘fishy’ odor and gives the characteristic odor to seafood. Humans can convert TMA in the liver to trimethylamine oxide (TMAO).

14 KRILL OIL CONCENTRATE Latest Krill Oil studies

Several articles on health benefits of tem function is reduced. Such a compro- krill oil supplementation in both ani- mised immune system can occur after mals and humans have been published heavy training and sports competitions, over the years and positively changed where the risk for infection is increased, health parameters are presented in especially for upper respiratory tract in- more detail in the section on Krill oil fections of athletes. in health and disease. The very latest krill oil science has emerged in the new It is true that moderate exercise in areas of sports nutrition and skin. comparison to a sedentary lifestyle lowers the risk of upper respiratory SPORT tract infections (URTI). However, high The immune system is a complex inter- intensive exercise can have immune play of cells, tissues and organs to sup- system modulating effects that weak- port tissue repair and prevent invasion of en the defense mechanisms of a recov- the body by bacteria, parasites, viruses, ering body [6]. and fungi. Omega-3 fatty acids enhance The relationship between exercise in- the immune function by mixing into im- tensity and risk of URTI was s hown in a mune cell membranes, which reduces the graph by Professor Nieman [17]. amount of the pro-inflammatory fatty acid, arachidonic acid. The incorporation of omega-3 fatty acids instead promotes the formation of the less potent EPA and DHA-derived inflammatory mediators. The immune-enhancing properties of omega-3 fatty acids might particularly help in situations where the immune sys-

15 KRILL OIL CONCENTRATE The immune system changes after letes, only one athlete was within the heavy exercise include a decrease of in- optimal omega-3 target range that low- terleukin 2 (IL-2) and interferon gamma ers the risk for cardiovascular events (IFN-γ), molecules that can regulate the and suboptimal brain function (reaction activity of immune cells. Likewise, nat- time and executive function) [19]. These ural killer (NK) cell function was shown surprisingly low omega-3 levels, which to be decreased after heavy exercise. were even lower than in heart disease NK cells are the first line of defense by patients [20], may be explained by the reacting quickly to e.g. intruding bacte- high need of energy of an athletes’ body ria and viruses to keep them under con- that might use these essential fatty ac- trol until the antigen-specific immune ids as an energy source. system starts to act. Their activity can Omega-3 supplementation might there- be decreased by up to 60% for several fore be crucial to help athletes to opti- hours after extended exercise [17]. This mize their physical and mental perfor- has led to the view that after intense mance, also in hindsight that EPA and training or sport competitions, there is DHA have the ability to decrease heart an open window of 1-9 hours with low- rate and oxygen consumption during ered host defense, which increases the exercise [21]. likelihood for an infection. The potential of krill oil to strengthen Omega-3 supplementation has the immune function after a simulat- been shown to increase IL-2 and INF-γ ed cycling time trial has been tested production, as well as NK cell function in both male and female participants and might therefore help to increase host [22]. The study was conducted at the Uni- protection after exercise. versity of Aberdeen, Scotland under the It has been recommended by supervision of Dr Stuart Gray, Senior Lec- Simopoulos that athletes at a leisure turer in Exercise Physiology. level should eat up to 2 grams of fish oil The Omega-3 index, a mea- per day in a 2:1 ratio of EPA to DHA [18]. sure of the percentage of EPA and Athletes at a competition level should DHA of total fatty acids in red blood probably consume even more omega-3 cells, was measured after 6 weeks of fatty acids to fully benefit from the im- either 2 grams daily krill oil or placebo mune-modulatory effects. In a study of consumption. The results showed that 106 German winter elite endurance ath- those participants administered krill oil

16 KRILL OIL CONCENTRATE had a statistically significant increase that supplementation of the diet with 2 of their Omega-3 index. grams per day of krill oil for 6 weeks can After 6 weeks of study prod- significantly increase the production of uct supplementation, the volunteers IL-2 and increase the toxic effect of NK performed an incremental maximal ex- cells on other cells in the recovery pe- ercise test on a cycle ergometer. Par- riod after exercise. The effect was gen- ticipants cycled at 70 revolutions per der-independent. minute with workload increasing by 30 Study coordinator, Dr Stuart Gray com- Watts every minute for males, and 20 ments: “Our study is in agreement with Watts every minute for females, until our previous work with fish oil, where volitional exhaustion. we have observed similar results. How- Immune function parameters, like cell ever, the krill oil EPA and DHA dose used signaling molecule production (IL-2, IL- was only a quarter from the dose given in the earlier fish oil study. It remains to be proven if the different structural form (omega-3 phospholipids from krill oil versus omega-3 triglycerides from fish oil) can explain this difference.” Moreover, a previous double-blind 6-week study on krill oil supplementation to the Polish National Rowing Team has shown that krill oil can affect levels of pro- and anti-oxidant balance markers [23]. Study authors found that exercise significantly increased values of oxidative stress parameters Active people need omega-3 fatty acids in both groups, but recovery levels were 4, IL-10, IL-17 and IFN-γ) and the ability significantly lower in athletes receiving to destroy target cells by NK cells were 1 gram krill oil per day compared to the measured at baseline and in the recov- control group. Based on these results ery period after exercise (post-exer- they concluded that supplementation cise, 1h and 3h). with krill oil in trained rowers dimin- The results demonstrated ished post-exercise oxidative damage

17 KRILL OIL CONCENTRATE to red blood cells during recovery. can amount to as much as 3.6 kg of dead Overall, several sport studies skin cells in a year. Under the epidermis including omega-3 fatty acids highlight lies the dermis as an inside layer in- the importance of an adequate ome- cluding sweat glands, hair follicles, and ga-3 fatty acid intake for athletes. nerve endings needed to feel tempera- The ability of krill oil to positively influ- ture, pressure, and pain. ence oxidative stress and immune func- The stratum corneum is im- tion shows that regular consumption portant in preventing water loss to of omega-3 phospholipids from krill oil keep the underlying living cells moistur- might be an effective nutritional strate- ized. If its barrier function is disturbed, gy to help athletes in the post-exercise the skin can become dry and flakey. Hot recovery phase. water, soaps, medication, low humidity, and medical conditions can all lead to Krill oil supplementation has beneficial dry, irritated skin. For example, eczema effects on oxidative stress and immune is characterized by dry, red skin patches function and can help to improve a bal- that can be very itchy, which might be anced sports nutrition for athletes. due to an overactive immune system, but the exact causes are unknown. Also Skin in the case of psoriasis it is unknown Skin is the organ that not only protects why the immune system is not working against microbes, pollution, and phys- properly, which leads to the formation ical assaults, it is further important of excess skin cells that build up in for sensory perception and in maintaining the body’s water content and temperature. It consists of several layers. The epidermis is towards the outside, with the top cell layer called the stratum corneum, which mainly consists of dead cells and oils. The body sheds the dead cells with a rate of up to 40 000 cells per hour. This

18 KRILL OIL CONCENTRATE scales and red, itchy patches. which can lead to discomfort or even infection. Healthy skin permits water loss only In general, the skin holds to a limited amount. Damaged skin, about 30% of the body’s water and the such as in atopic eczema, is character- dead cells of the stratum corneum and ized by increased water loss. Krill oil the lipids ‘waterproof’ the human body. was shown to alleviate water loss and Water loss and hydration of the stra- dry skin in a 12-week clinical study in tum corneum is linked to its lipid con- healthy persons. tent, the generation of new lipids in the skin, and ultimately the level of damage Both omega-3 fatty acids and to the skin barrier function. A good bal- phospholipids are important for skin ance between the skin’s water content health. Deficiency leads to scaling and and the amount of water passing influ- dryness of the skin and enhanced tran- ences skin elasticity, smoothness, and sepidermal water loss (TEWL) [24,25]. roughness and is crucial in maintaining TEWL describes the water loss over the healthy skin [26]. surface of the skin that happens by pas- sive diffusion. The outer layer of dead cells makes the skin flexible and elastic, when it contains enough water. Howev- er, when the TEWL is high and hydration low, it becomes hard and rough. Hence, skin function is negatively affected,

19 KRILL OIL CONCENTRATE Changed lipid levels in the skin Phospholipid effect Increased hydration Decreased water loss

KRILL OIL Changed lipid levels in the skin Modulation of inflamation Increased hydration Omega-3 effect Decreased water loss Improved elasticity Improved smoothness

The skin has several functions, but one Krill oil is an attractive dietary of the most critical ones is to maintain supplement that might help in the mainte- the body’s water levels and to limit wa- nance of skin health and treatment of skin ter loss into the surroundings. Water disorders by both its omega-3 and phos- makes up 75 (infants) to 55% (elder- pholipid components (see above ). ly) of the body’s weight and is vital for To explore the effects of krill biological processes and life. A way to oil on skin health, 31 volunteers (mid- measure water loss is to assess trans- dle-aged men and women with normal epidermal water loss (TEWL). skin) were included in an open label, two-armed clinical trial (unpublished re- In contrast to e.g. saturated sults). The subjects were randomized to fatty acids and cholesterol, EPA and DHA take 3 grams daily krill oil for 13 weeks. cannot be made in the skin and must be All skin measurements were done on the obtained from outside sources. The skin upper forearm and showed that krill oil lacks the enzymes needed to convert intake resulted in a significant increase short-chain omega-3 fatty acids into the in skin hydration and elasticity, as well longer chain EPA and DHA [27]. Omega-3 as in a significant reduction in TEWL. fatty acid levels in human skin are rather In addition, a significant cor- low with under 2% of total skin fatty ac- relation for the change in hydration ids. However, significant increases can and the changes both for elasticity and be achieved by supplementation and 3 TEWL were found. Meaning that the months of 4 grams daily EPA increased volunteers that had a high change in skin EPA 8 times [28]. skin hydration also experienced a high

20 KRILL OIL CONCENTRATE change for both elasticity and TEWL. By using a digital camera, the amount of wrinkles (as a measure for roughness) and width and size of wrinkles (as a measure for smoothness) were assessed at the start and end of the trial. For both parameters, i.e. roughness and smoothness, there was a significant beneficial change observed after krill oil supple- Krill oil improves skin parameters mentation. By increasing the amount of omega-3 fatty acids in the body, krill oil has the ability to not only influence skin hydration and elasticity, but also the amount and size of wrinkles. The measure of how much omega-3 accumulates in the body, given as the Omega-3 index, was increased significantly in the study and correlated with the above mentioned parameters. These results indicate that the increases in Omega-3 index by krill oil supplementation directly relate to the positive changes in skin parameters observed among study subjects.

21 KRILL OIL CONCENTRATE Phospholipids Carbohydrates, nucleic acids, proteins, and lipids are the four main biological compounds that determine life. Whereas, phospholipids are part of the lipid group, which are the very basic units of life.

WHAT IS A PHOSPHOLIPID? A phospholipid molecule consists of two fatty acids, which are long chains of carbon and hydrogen molecules. They are attached to a glycerol backbone that is further linked to a phosphate group. The phosphate group has a head group, such as choline, resulting in phosphatidylcholine (PC). The word lecithin originates from the While choline, phosphate, and glycerol Greek lekithos, egg yolk. Lecithin’s make up the hydrophilic (water-friend- chemical name is phosphatidylcholine, ly) side of the molecule, the fatty acid while commercially it is a phospholipid chains are the hydrophobic (water-fear- mixture. Most lecithin products consist ing) part of the molecule. of phosphatidylcholine, phosphatidyl- The first phospholipid was ethanolamine, phosphatidylserine, and identified in 1847 by Theodore Nicolas phosphatidylinositol [29]. Gobley, a French chemist and pharma- cist who analysed lecithin (a rich source Phospholipids in the form of lecithin of PC) in egg yolk. are used as emulsifiers and stabilizers in foods, in cosmetics and paints or for Phospholipids, as the name indicates, therapeutic reasons, and as nutraceuti- are made up of the mineral phosphorus cal supplement. In 1933, lecithin already and lipids (fats). appeared in the Italian medicine dictio- nary Medicamenta, for conditions such as, diabetes, tuberculosis, depression,

22 KRILL OIL CONCENTRATE and recovery from infectious diseases Both the inside and the outside of [30]. cells is water. When phospholipids are exposed to water, they arrange them- FUNCTIONS OF selves into a two-layered sheet (a bi- PHOSPHOLIPIDS layer) with all of their hydrophobic tails IN THE BODY pointing towards the center of the sheet Phosphorus, found in the head group and the hydrophilic heads towards the of phospholipids, contributes to the surrounding water. normal function of cell membranes, energy-yielding metabolism, and in bones The amount of phospholipids in the hu- and teeth [31]. The phospholipid mol- man body is enormous! The body con- ecule has many functions and uses in tains over 100 trillion cells with mem- the body. For example , as a structural branes, consisting of phospholipids, element in cell membranes, a source of and proteins. The liver itself consists choline for the neurotransmitter acetyl- of membranes that can fill four soccer choline, an important factor in energy fields. production and storage, assisting in blood clotting, antioxidant protection, and About 60% of what is made in a cell is cholesterol solubility to name just a few. needed for membrane structures, either Moreover, it acts as surface-active wet- for channels and receptors inside the ting agent that lines the outside of liver, membrane or proteins attached on the lungs, gastrointestinal tract, and kidney outside of membranes [33]. Membranes cells. not only give a defined volume to a cell, they also allow for internal structures, CELL MEMBRANES such as organelles (e.g. nucleus) and The structural skin around cells and transport vesicles between organelles. their organelles is called a membrane. It is important that our cells maintain a sufficient amount of phos- A phospholipid membrane is only five pholipids to ensure optimal cell func- millionths of a millimeter thick [32]. tion. Otherwise many cell functions can 10,000 membranes on top of each other be compromised, such as their ability to have the same thickness as a piece of control the exit of waste and the entry paper. of nutrients into cells, communication

23 KRILL OIL CONCENTRATE between cells, membrane-bound en- and contain phosphate for energy pro- zyme functions, binding of molecules duction. Moreover, they are needed in (neurotransmitters, antigens, antibod- fat emulsification in the stomach and ies, etc.) to receptors, and more. blood clotting. Two of the many factors which may affect the amount of phospholipids in cells are, i) as we age phospholipid LIPID TRANSPORT AND amounts in cells decrease, and ii) cells CHOLESTEROL METABO- in skin, lungs, liver, heart, and blood LISM vessels can become vulnerable to at- In addition to the central role of phos- tack from free radicals and toxins [34]. pholipids in membrane function, they Therefore, damaged and lost phospho- are, together with apoproteins, part of lipids must be continuously replaced to the outer layer of transport vehicles, ensure optimal cell function and health. called lipoproteins, because they are made of fat (lipid) and proteins. This Phospholipids with their unique po- combination of solubilizers allows the sition in cell walls are essential for transport of cholesterol in the wa- cell growth and the generation of new ter-based bloodstream. Cholesterol is cells. They promote molecule trans- important for cell membrane function port across membranes, binding to and as precursor for other compounds. receptors and enzyme activities and However, in high amounts, it will accu- determine the fluidity of membranes. mulate on and in blood vessel walls and They are a source of messengers in cell make them narrower and stiffer. The signaling, choline, omega-3 fatty acids hardening of arteries via the buildup

24 KRILL OIL CONCENTRATE of cholesterol and fat plaques is called bile) or made by the liver. The absorp- atherosclerosis. If these plaques burst, tion rate of cholesterol in the gut, can they can lead to blood clots that can vary substantially [36]. It is influenced cause heart attacks and stroke. by the amount of phospholipids avail- By forming monolayers, phos- able, since phospholipids are needed for pholipids surround all lipoproteins that the intestinal uptake of cholesterol, but are classified according to their density high amounts of phospholipids reduce in chylomicrons, VLDL (very low-den- cholesterol absorption through molec- sity), LDL (low-density), IDL (interme- ular interactions. Indeed, it was found diate-density) and HDL (high-density) that a dose of 15mg lecithin can inhibit particles. Whereas large lipoproteins cholesterol absorption by 50% in rat have low density and contain more fat guts [37]. There is also evidence for re- than protein. In short, cholesterol is duced intestinal cholesterol absorption taken up in the small intestine and is in the gut of humans after phospholipid delivered to the liver in packages called supplementation [38-40]. chylomicrons. VLDL and LDL are se- creted by the liver and are risk factors Research has shown that dietary phos- for atherosclerosis and heart disease pholipids may help to reduce blood LDL by providing cholesterol to plaques. particles, the ‘bad’ cholesterol and re- LDL is therefore often called the ‘bad’ duce the risk of heart disease by affect- cholesterol. In contrast, HDL, the ‘good’ ing intestinal cholesterol uptake. cholesterol, has a protective effect by removing excess cholesterol from the arteries and bringing it back to the liver on the ‘reverse transport pathway’.

Every 1% increase in blood HDL levels has been associated with a 2-3% reduction in overall heart disease risk [35].

The levels of cholesterol in blood pri- marily depends on the amount of cholesterol that is ingested (from diet and

25 KRILL OIL CONCENTRATE which later join the HDL blood pool. Phospholipids move into cells and their membranes by selective and whole particle uptake routes. In liver cells, about two-thirds of the phospholipid uptake is by selective entry routes. Thereby, phospholipids are transferred directly from lipoproteins into the cell membrane. In addition to selective uptake pathways, cells can incorporate entire lipoproteins by endocytosis, which results in an unspecific incorporation of everything that was part of the particle, including phospholipids. Incoming phospholipids can be metab- olized to triglycerides and can be used in storage or energy generation, when not used in membranes. “Illustration provided and modified courtesy of the National Heart, Lung, and Blood Institute’s The Heart Truth® Program.” PHOSPHOLIPID SOURCES UPTAKE OF Daily phospholipid intake is usually be- PHOSPHOLIPIDS tween 2-8 grams, which corresponds Dietary phospholipids are efficiently to 1-10% of the daily fat consumption (above 90%) taken up into the cells of the [41]. Phospholipids can be found in high intestinal wall after they are hydrolyzed quantities in egg yolk, soybean, meat, to lysoPC and free fatty acids by lipases fish and internal organs, while fruits, in the small intestine. In the intestinal vegetables and grains have a much low- cells, they are reassembled and included er phospholipid content [42]. However, into chylomicrons for further transport the richest sources usually also have a via the lymph and blood. Some studies high content of fat and cholesterol. suggest that a fraction of the dietary Also krill contains high amounts of phospholipids is directly taken up into phospholipids and E. superba is known HDL particles already in the intestine, to use them as energy storage form. In

26 KRILL OIL CONCENTRATE the extracted krill oil concentrate, over plementation are presented in Table 56% of the oil consists of phospholip- 2 that underlie their liver-protective, ids. anti-inflammatory, immune changing, anti-obesity, memory enhancing, an- PHOSPHOLIPID ti-depressant, and anti-tumor effects DEFICIENCY [45]. Their effects on liver health and Not much is known about the optimal physical performance, are outlined in intake levels of phospholipids, since more detail below. phospholipid deficiency symptoms are unknown, except the ones associated LIVER with choline deficiency that are de- The liver is one of the organs that get scribed later on. However, due to refine- the most membrane damage because ment of oils and fats, cleaned raw ma- of its blood filtering function. It deals terials, and changes towards a nutrition with incoming food nutrients and ei- low in fat and cholesterol, modern diets ther stores or prepares them for fur- contain only about one third of phos- ther transport to other organs. It is pholipids than what they contained a also responsible for filtering the blood century ago [43]. In particular, high-risk from harmful substances such as tox- groups for deficiency, such as the elder- ins, alcohol, medication, fatty food, ly, athletes and diseased persons might waste products, viruses, etc. These are all therefore benefit from phospholipid stressors for the integrity of the hepatic supplementation. membrane and fat accumulation or alcohol misuse can take its toll on the liver PHOSPHOLIPIDS and eventually lead to fatty liver disease IN HEALTH AND that can progress into cirrhosis and liver DISEASE failure. Although dietary phospholipids are While studying 700 obese present in many foods, purified forms children, it was found that 15% (10% can help to treat various health con- girls, 22% boys) of the children already cerns, including fatty liver, arthritis, had fatty liver disease [56]. Overall, the heart disease, cachexia, hypercholes- prevalence of suspected nonalcoholic terolemia, cancer, and more [44]. Major fatty liver disease lies at 50% of obese health benefits of phospholipid sup- males in the U.S. [57]. While 45% of

27 KRILL OIL CONCENTRATE Table 2: Dietary phospholipid (PL) and choline health benefits. Health area Benefit Explanation Referenc- es Heart disease Reduced PL improve parameters related [46-48] cardiovascular to heart and cardiovascular risks diseases: • Blood lipid profiles (reduction of total cholesterol, LDL and TG levels and increased HDL levels) • Reduced hypertension • Reduced platelet aggregation and the risk of arteriosclerosis Immune Improvement of PL-induced improvement in [49] system immunological phagocytosis, arachidonic acid function concentration and neutrophil killing activity. Infant Improved brain Choline supplements during [50] development development of pregnancy have been shown fetus to affect brain development of fetus and improve lifelong memory characteristics (animal studies). Liver disease Improved hepat- Reduced alcohol-induced liver [51] ic disorders damage and hepatic damage caused by toxins and virus infections (hepatitis). Physical Improved phys- PL stimulates acetylcholine [52] performance ical synthesis, which triggers the performance release of neurotransmitters by reduction of in the brain, thus improving physical stress physical performance. Stomach and Reduction of PL-induced protection of the [53] gastrointesti- stomach pain in- GI by reduced gastric mucosal nal tract (GI) duced by gastric lesions. Typical GI side effects acid of analgetic drugs (NSAIDS) are reduced, likely due to an increased production of cyto- protective mucosal PGE2.

28 KRILL OIL CONCENTRATE Stress and Reduction in Reduced production of stress [54] depression stress-related hormones linked to strenuous mood symptoms exercise and eased stress-related mood symptoms. Ulcerative Protective and Supplementation of PC has [55] colitis healing effects been shown to protect and in inflammatory restore the intestinal lining of ulcerative colitis the gastrointestinal wall.

GI, gastrointestinal tract; HDL, high-density lipoprotein; LDL, low-density lipoprotein; NSAIDS, nonsteroidal anti-inflammatory drug; PC, phosphatidylcholine; PGE2, prostaglandin E2; PL, phospholipids type-one diabetics have fatty liver [58], Whole phospholipid, in contrast to up to 85% of type-two diabetics are af- choline and omega-3 fatty acid supple- flicted with the disease [59,60]. mentation alone, has been shown to influence liver fat metabolism by either Liver disorders include fatty liver due affecting cholesterol and bile acid gen- to cellular fat accumulation in people eration, degradation of fatty acids or that are e.g. overweight, have diabetes, the secretion of lipoproteins from the or drink too much alcohol leading to an liver [61]. As shown in animals, a diet rich enlarged liver. Cirrhosis, the inflamma- in phospholipids reduces liver fat by tion and scarring of liver tissue, can be means of inhibiting intestinal fat uptake due to hepatitis infection or excessive and influencing the activity of liver en- alcohol consumption, which can also zymes that regulate lipid metabolism. lead to cancer. Autoimmune liver disor- Moreover, liver damage was found to be ders damage liver cells by an abnormal- alleviated by phospholipids. Reduced ly high amount of immune cells. phospholipid levels in liver membranes, as for example found after alcohol consumption, can be counteracted by phos-

29 KRILL OIL CONCENTRATE pholipid supplementation. Accordingly, fatty acid synthesis, enhancement of fatty a study on chronic alcohol consumption acid degradation, and increase of serum ad- in baboons that were given either ethanol iponectin levels [66]. or ethanol together with lecithin over 6.5 years reported that PC protects against liv- BRAIN er fibrosis and cirrhosis [62]. But also other Phospholipids play a central role in brain liver damages like e.g. from viral infections, function and around 60% of the brain by can benefit from phospholipid supplemen- weight consists of phospholipids. Phos- tation to decrease disease activity [63]. pholipids are particularly enriched in dendrites and synapses and it has been Fat accumulation in the liver will often not shown in vitro that nerve growth increas- lead to symptoms indicating that some- es the demand for phospholipids. Nerve thing is wrong. Inasmuch, fatty liver silently growth factor, a small protein controlling increases heart disease risk 3 times in men, nerve growth and maintenance, also stim- 14 times in women and up to 10 times in ulates phospholipid generation [67]. type-one diabetics [64]. The omega-3 fatty acid DHA can only to a very little extent be made by the The majority of the studies performed brain, and must therefore be supplied by with phospholipids, did not include ome- the blood and imported over the blood- ga-3-containing phospholipids, indicating brain barrier. Phospholipids are of utmost that phospholipids in general have benefi- importance in the transport of DHA, since cial effects. However, other studies have the recently discovered DHA transporter demonstrated that phospholipids contain- (Mfsd2a, major facilitator super family ing omega-3 fatty acids have more potent domain containing 2a) accepts DHA only effects on liver and blood plasma lipid if it is bound to phospholipids; to be exact levels, compared to phospholipids without to lysoPC [68]. Mice that are genetically omega-3s [65,66]. engineered to not have this transporter, In particular, Shirouchi and col- have very low DHA amounts in the brain leagues have concluded in their study on leading to neuronal cell loss and cognitive rats that the combination of omega-3 fatty deficits associated with severe anxiety. acids with phospholipids in comparison to In addition, humans identified with muta- egg phospholipids alone can alleviate liver tions in Mfsd2a present defective brain steatosis better through the suppression of growth and intellectual disability due to

30 KRILL OIL CONCENTRATE insufficient DHA-lysoPC uptake into the brain [69,70]. LysoPC in combination with DHA from the blood is therefore vital for normal brain growth and function.

The cells of the small blood vessels com- ing into the brain are tightly stitched together by tight junctions forming the Mfsd2a transports DHA over the blood-brain barrier Illustration adapted with permission from Christer Betsholtz Dr. so-called blood-brain barrier to restrict free passage of molecules. This allows But not only bring phospholipids DHA for the protection of e.g. bacteria infect- into the brain, they also provide choline, ing the brain. Even DHA needs a special an essential nutrient [71], which makes transporter that will transport DHA only up about 15% of the PC molecule. Like when it is bound to lysoPC. DHA, choline is important for brain development and nerve signaling and thereby influences cognition [72]. Hence, a 47% reduced risk for getting dementia was found in the elderly that had the highest PC levels in blood [73]. Choline WHAT IS CHOLINE? Choline is an essential vitamin-like nutrient that is crucial for normal cellular function.

31 KRILL OIL CONCENTRATE FUNCTIONS OF NERVE SIGNALING CHOLINE IN THE BODY Choline is converted to acetylcholine, Choline as a component of phospha- a neurotransmitter, in the nervous sys- tidylcholine, choline plasmalogen and tem. Acetylcholine is of importance in sphingomyelin is a major component of learning, breathing, memory, sleep, and cell membranes. muscles metabolism. Whereas in heart tissue, acetylcholine has an inhibitory 95% of choline in the body is found in effect and leads to reduced heart rate, phospholipid form as phosphatidylcho- in skeletal muscle it has the opposite line (PC). effect.

The derivatives of choline are ver- CELL SIGNALING satile and include functions such as Choline is a part of the phospholip- neurotransmitters (acetylcholine), ids phosphatidylcholine and sphin- cell membrane signaling (phospholip- gomyelin. They are the precursors of ids), lipid transport (lipoproteins), and diglycerides and ceramides, which are methyl-group metabolism (homocys- lipid-signaling molecules in cells. Lipid teine reduction) [74]. Besides, a fetus messengers can freely cross over mem- depends on choline for brain and membranes and can therefore not be stored ory development [75] and to reduce the in vesicles. They act locally on recep- risk for neural tube defects [76]. A lot of tors and enzymes to induce a specific the maternal choline goes to the fetus, cellular response. which might deplete the mother’s choline reserves. METHYL DONOR Choline is required for the metabolism of nucleic acids and amino acids and is an important source of methyl (-CH3) groups over the generation of S-ad- enosylmethionine (SAMe). Up to 50 chemical reactions in mammals depend on SAMe as a methyl donor [77]. These methylation reactions are of importance in lipid biosynthesis, regulation Choline is important in nerve signaling

32 KRILL OIL CONCENTRATE Brain Functions CHOLINE Memory Acetylcholine Neurotransmitter

Betaine

Dimethylglycine Heart Health

Homocysteine Related to heart disease

S-adenosyl Methionine homocysteine

S-adenosyl Brain Health methionine Important in control of DNA Liver Health replication and cell growth Phosphatidyl- choline Phosphatidyl- Courtesy of www.cholinecouncil.com Important componment of cell menbranes ethanolamine of metabolic pathways, and detoxifica- Betaine is further used to convert ho- tion. Changes in DNA methylation, that mocysteine, a risk factor for heart dis- affects gene expression, have been ease, to methionine. linked to cancer. UPTAKE OF CHOLINE SALT WATER BALANCE VERSUS PHOSPHATIDYL- Choline is converted to betaine, which CHOLINE is an osmoregulator, meaning that it can Dietary choline is taken up by choline regulate the volumes and water con- transporters in the intestine. Most of the tent of cells [78]. Cells keep water lev- choline is converted to PC and used in cell els constant by allowing ions to enter, membranes. The liver can recycle choline which makes water to follow and move and the intestine, lungs, and kidneys will into cells. Betaine is an organic water send choline to the brain and liver in time attractant, in particular in cells with of need. high osmotic pressure such as kidney or Administration of choline will intestinal cells. significantly increase blood choline con-

33 KRILL OIL CONCENTRATE centrations [79,80]. It was found that cho- liver, beef, and milk. Although most di- line in the form of PC, is 12 times better ets are considered to provide sufficient in raising human blood choline amounts choline, persons at risk of choline defi- compared to choline salt [79,80]. While ciency exist. Specifically, the National choline salt show maximum levels after Health and Nutrition Examination Sur- 30 minutes (86% increase, 4 hours until vey in 2003-2004 has concluded that normal), PC intake raises choline by 265% 90% of the American population has an and takes 12 hours until normal again [80]. inadequate intake of choline [1]. It was suggested that 60% of the choline in inorganic salts, such as choline chloride, 90% of the U.S. population is not con- choline citrate, and choline bitartrate, is lost suming enough choline to ensure opti- to conversion to TMA by intestinal bacteria mal membrane function, neurotrans- [23]. In comparison, dietary PC leads to mission, and methyl donor availability. three times less TMA [24]. The adequate intake level for choline CHOLINE DEFICIENCY was set at 550 mg/day for men and 425 Only little choline can be made by the mg/day for women in sight of prevent- body itself, the major part needs to be ing liver damage. Choline deficiency-in- taken up over the diet. Choline is found duced fatty liver can occur, when not in higher amounts in e.g. soybeans, eggs, enough choline is present for the for- Courtesy of www.cholinecouncil.com

34 KRILL OIL CONCENTRATE mation of PC molecules that are needed for the formation of VLDL particles. If the fat becomes stuck in the liver, it accumulates and will eventually lead to liver damage. Excessive oxidative stress that damages lipids and DNA in- duces cell death and inflammation that can ultimately culminate in end-stage Choline supplementation might liver disease [81]. Choline deficiency is improve brain functions also the only nutrient deficiency that can lead to cancer. choline, improves brain function in persons affected by memory impairments Genetic predisposition, gender, and due to aging or pathological conditions. age influences ones choline needs and Inasmuch, Alzheimer’s and Parkinson’s care should be taken to ensure an ade- disease patients have a reduction of quate choline intake, since choline defi- cholinergic neurons in the grey matter ciency is linked to an increased risk for of their forebrains [83]. Also elderly fatty liver, muscle dysfunction, athero- persons have reduced forebrain cholin- sclerosis, and neurological disorders ergic neurons and acetylcholine [84]. On [82]. the other hand, choline supplementation increases the amount of choline in CHOLINE IN HEALTH the blood, with a concomitant increase AND DISEASE of choline-containing compounds in the With its many diverse roles, it is not brain [85]. However, in an analysis of surprising that choline deficiency will all available literature on studies with cause disease in humans. The function lecithin given to individuals with Alzhei- of many organs such as liver, muscle, mer’s disease, Parkinsonian dementia kidney, pancreas, and brain all depend and subjective memory problems, no on an adequate choline intake. clear benefit for memory performance could be found for patients with Alz- BRAIN heimer’s disease and Parkinsonian de- It was suggested that supplementation mentia [86]. Yet, people with subjective with choline, as a precursor for acetyl- memory difficulties, gained a very sig-

35 KRILL OIL CONCENTRATE nificant benefit in one of the analysed choline salts alone [79]. studies [87]. Thus, while the overall If, during exercise, enough choline is analysis does not support lecithin for available in the body, then also sufficient dementia patients, people with sub- acetylcholine is produced. Since acetyl- jective memory complaints may profit choline is a signaling molecule important from lecithin supplementation. for muscle contractions, sportive performance might therefore be optimized by PC supplementation.

Acetylcholine is a neurotransmitter important for muscle contractions made from choline. The available free choline influences acetylcholine synthesis rate. Choline is important for muscle contractions

If no choline is supplemented, mara- thon runners can manifest up to a 40% SPORT decrease in blood choline levels [16,17], Intense physical activity challenges which is similar to the choline reductions cellular functions, which are essential found in cyclists [5,18]. Besides, a study to maintain for optimal sports perfor- with lecithin supplementation found fast- mance. Excessive radical formation and er recovery after a bicycle ergometer tri- trauma to the muscles during high-in- al, when compared to the control group tensity exercise leads to inflammation. [26,27] It has been suggested that supplementation with phospholipids, such as During intense exercise, free choline is PC might be beneficial to endurance reduced, which might impact on acetyl- athletes, because of the choline head choline release and therefore sportive group [88]. Indeed, decreased blood performance. choline levels were shown in endurance athletes such as cyclists and runners [89]. Supplementation with PC significantly increases circulating choline levels; in fact twelve times better than

36 KRILL OIL CONCENTRATE Omega-3 Fatty Acids Omega-3 fatty acids played an essential role in human evolution and development. A turning point for the evolution of human intelligence was the diet change from red meat of animals mainly consumed by the Neanderthals to the inclusion of coastal seafood and inland freshwater sources [90]. This might have initiated the growth of the brain about two million years ago.

atoms from the methyl end. In compar- WHAT ARE OMEGA-3 ison, an omega-6 fatty acid has its first FATTY ACIDS? double bond at 6 carbon atoms from The main bioactive omega-3 fatty acids the methyl end. that have been described extensively are eicosapentaenoic (EPA or 20:5n- FUNCTIONS OF 3) and docosahexaenoic acid (DHA or OMEGA-3 FATTY ACIDS 22:6n-3). EPA consists of 20 carbons Aside from being a source of nutritional and 5 double bonds and can be convert- energy, the functions of omega-3 fatty ed enzymatically into DHA. DHA is the acids have a molecular basis. Inasmuch, longest fatty acid chain, with 22 carbons the omega-3 fatty acids have the ability and 6 double bonds. They are called to change membrane fatty acid com- omega-3 fatty acids, because they have position and function, regulate gene their first double bond at three carbon transcription, and alter metabolic and

37 KRILL OIL CONCENTRATE signal transduction pathways. This in- volves a wide array of mechanisms that overlap and interplay in complicated metabolic networks that maintain the body’s equilibrium.

MEMBRANES Most importantly, omega-3 fatty ac- Omega-3 fatty acids influence gene expression ids, as well as the overabundant omega-6 fatty acids, are crucial as building GENE TRANSCRIPTION AND blocks of membrane structures and a ENZYME ACTIVITY cell’s development, integrity and func- When omega-3 fatty acids reach cells, tion. The flexible structure of omega-3 they can activate transcription factors fatty acids determines the fluidity of that stimulate the expression of certain membranes. Membrane fluidity is im- genes [91-94]. This means that omega-3 portant for the correct functioning of fatty acids indirectly influence the pro- membrane proteins such as receptors, duction of functional gene products ion channels, transporters, and en- (often proteins) in our cells. The ome- zymes. A change in fluidity due to al- ga-3 fatty acids thereby influence tered membrane fatty acid composition which proteins are made in a cell and, will change activities and movements ultimately, how the cellular metabolic of these proteins. It will also affect how functions are impacted. It is still un- extracellular signals are transmitted clear to which extent EPA and DHA bind from receptors to intracellular signal- specifically to different transcription ing networks, which is important in e.g. factors, but there are at least some in- neurons, cardiac cells and hormone-se- dications that there are preferences as creting cells. Additionally, membrane to which fatty acid can regulate which permeability increases with the number gene expression [91]. However, not only of double bonds present in fatty acid can fatty acids change the expression chains of membrane phospholipids. of genes, they can also modulate en- The amount of omega-3 fatty acids that zyme activities by directly binding to constitute membrane phospholipids them [95,96]. can be influenced by diet.

38 KRILL OIL CONCENTRATE EICOSANOIDS ENDOCANNABINOIDS The ratio of omega-6 to omega-3 fatty The endocannabinoid system is based acids is important since they compete on the action of endocannabinoids for the same metabolic enzymes that (ECs) that are hormones made from convert them into eicosanoids (i.e. omega-6 fatty acids. The binding of ECs prostaglandins, prostacyclins, throm- to receptors influences the expression boxanes, and leukotrienes). Eicosa- of proteins in organs (e.g. liver, skeletal noids are hormone-like compounds, muscle, pancreas, intestine, bone, and called cellular hormones that control adipose tissue) and affects the action key functions in the body, such as the of the central nervous system. There- central nervous system, immunity, and by they can influence not only enzyme inflammation. activities, but also appetite, energy Whereas prostaglandins, leu- balance, mood, memory, pain percep- kotrienes, and lipoxins play a regulatory tion, stress response, anxiety, immune role in inflammation, thromboxanes and functions, and reproductive processes. prostacyclins are important for con- An overactive EC system is believed to trolling bleeding. Resolvins and protec- promote increased fat mass and var- tins, metabolites of EPA and DHA, help ious markers of metabolic syndrome reduce inflammatory responses [97]. [98]. In general, eicosanoids from The quantities of ECs made, ul- omega-3 fatty acids are less inflam- timately depend on the amounts of ome- matory than those from omega-6 fatty ga-6 arachidonic acid available in mem- acids. Hence, a disturbed ratio with high branes, which depends on the amount of omega-6 versus omega-3 fatty acids arachidonic acid ingested. Increased in- and a low omega-3 intake in general take of fatty acids of the omega-3 family will tip the balance towards the produc- positively influences the ratio of omega-3 tion of pro-inflammatory eicosanoids. to omega-6 fatty acids in blood and or- Therefore, a replacement of the ome- gans. In this way, less arachidonic acid is ga-6 arachidonic acid by EPA or DHA incorporated into phospholipids, result- may lead to a less inflammatory active ing in decreased conversion of arachidon- eicosanoid profile, decreasing the risk ic acid to ECs. Thus, dietary fatty acids for inflammatory diseases. constitute a means to change the body’s fatty acid composition and thereby EC

39 KRILL OIL CONCENTRATE levels, ultimately affecting membrane Nowadays, the ratio between omega-6 signaling events and leading to changed to omega-3 can be as high as 10-20:1, energy metabolism (food intake and en- whereas historically it was as low as ergy processing). 1-2:1 [101]. The recommendations today call for a ratio of 5:1 [102]. Some studies OMEGA-3 DEFICIENCY further indicate that the optimal ratio Both omega-3 and omega-6 fatty acids may depend on the disease state [102]. are needed for optimal health. Howev- When it comes to heart disease, a ratio er, since there is an abundance in the of 4:1 was linked to a reduction of 70% Western diet of omega-6 fatty acids in total mortality. Rectal cell prolifera- compared to omega-3 fatty acids, the tion in patients with colorectal cancer balance between the two is highly dis- were reduced at a ratio of 2.5:1, while a turbed [99,100]. The underlying reason ratio of 2-3:1 suppressed inflammation is the vast increase in the consumption in patients with rheumatoid arthritis. A of vegetable oils rich in omega-6 fatty ratio of 5:1 was needed to induce bene- acids, which are present in corn, sun- fits for asthma patients, whereas a 10:1 flower seeds, cottonseed and soybean ratio had negative effects. and industrially produced meat. At the The right amount of omega-6 same time, the consumption of omega- and the right balance between omega-6 3-rich fish has decreased markedly. to omega-3 fatty acids is therefore essential for health, as otherwise too The human body has evolved on a del- many pro-inflammatory molecules are icate balance of omega-6 to omega-3 produced from omega-6 fatty acids fatty acids. Cells in the body rely on a that can affect disease outcomes. specific ratio of these fatty acids in Most people do not consume order to function properly. If out of bal- the recommended twice a week of ance, health is affected and it is of no fatty fish and hardly reach the recom- surprise that our modern unbalanced mended daily intake doses of EPA and diet promotes health issues such as DHA. Although the recommended daily heart disease, arthritis, depression, dose varies highly between different and dementia. countries (1000mg in Japan and South Korea, 500mg in France, and 450mg in Norway), most countries recommend

40 KRILL OIL CONCENTRATE 250mg (Austria, Belgium, Czech Re- balance, vision, and many more pro- public, Denmark, Finland, Germany, cesses in the body [105-107]. Greece, Iceland, Ireland, Italy, Luxem- bourg, Netherlands, Portugal, Slovakia, HEART Slovenia, Spain, Sweden, Switzerland, Most established is the influence and United Kingdom). While Australia of EPA and DHA on cardiovascular and New Zealand recommend 160mg of risk, since they can modify blood tri- EPA/DHA per day, other countries like glycerides, HDL cholesterol, plaque de- Canada, Hong Kong, Israel, Singapore, velopment, heart rate, and heart muscle Taiwan, and the United States do not functions [108]. By doing so, EPA and provide an official recommendation. DHA reduce the risk for cardiac death. Moreover, a meta-analysis that sum- Most people have mean intake levels be- marizes all available research on heart low the optimal daily EPA and DHA dose health found a connection between EPA of 250mg recommended by the World and DHA and a reduction in blood pres- Health Organization (WHO) and the Euro- sure (decrease of systolic blood pres- pean Food Safety Authority (EFSA) and sure of around 4.5mm Hg and diastolic could therefore benefit from omega-3 blood pressure of 3mm Hg), especially supplementation. in those with high blood pressure [109].

Already a decrease of 2 mm Hg blood OMEGA-3 FATTY ACIDS IN pressure is of importance, since it is HEALTH AND DISEASE linked to a 6% reduced stroke mortal- More than 20,000 studies on EPA and ity, 4% reduced coronary heart dis- DHA have assessed the health bene- ease mortality, and 3% reduced total fits of omega-3 fatty acids [103,104]. mortality [110]. Through their influence on membrane integrity, gene expression, the balance with omega-6 fatty acids and the generation of eicosanoids and endocannabinoids, they have the ability to modify heart disease risk, inflammatory response, neurological and psychiatric

41 KRILL OIL CONCENTRATE BRAIN two years of structural brain aging in Omega-3 fatty acids have been shown dementia-free study participants [116]. to be essential for mental health and DHA was further associated with better brain development and function [111]. visual memory, executive function, and Research has indicated that the high- abstract thinking. Generally, reduced er EPA and DHA intakes, the lower the brain volume is linked to cognitive de- risk for Alzheimer’s disease, age-relat- cline and dementia. Already at the age ed cognitive decline, depression, ag- of 30, the brain volume starts to decline gression, and more [112-114]. Omega-3 due to the normal aging process, but fatty acids may improve mental health at twice the speed, if mild dementia is by influencing nervous system activity, present, indicating the need for optimal memory, serotonin and dopamine lev- omega-3 intake throughout life. els, neurotransmission, and the forma- Besides their positive effect tion of synapses between neurons [115]. on heart and brain health, EPA and DHA It has been shown that indi- have been linked to a variety of health viduals with low blood EPA and DHA concerns of which a selection is listed in levels have smaller brains and that the Table 3. lower brain volume corresponded to

Table 3: Omega-3 fatty acid health benefits. Health area Benefit References Brain health DHA is highly concentrated in the brain and [117-121] is important for cognitive (memory and brain performance) and behavioral function. Effects were reported for: Mood Depression Cognitive function Emotional distress ADHD symptoms Alzheimer disease Memory Age-related cognitive decline Self-harm and suicidal thinking Cancer Mainly in cancer with gastrointestinal origin [122,123] work omega-3s as an anti-inflammatory agent.

42 KRILL OIL CONCENTRATE Eye health Omega-3 supplementation are reported to [124,125] have a positive effect on macular degeneration (a serious age-related eye condition that can progress to blindness) and dry-eye syndrome. Heart health Reduced risk of heart disease by: [126-129] ↓ Cholesterol ↓ Triglyceride ↓ Plaque development ↓ Arteriosclerosis ↓ Blood pressure ↑ HDL (“good cholesterol”) ↑Omega-3 index

Inflammatory Reduced inflammatory responses in a range [106, diseases of diseases such as rheumatoid arthritis, 130-132] inflammatory bowel diseases, systemic lupus erythematosus, and childhood asthma. The effect of omega-3s is due to a reduction in the production of many pro-inflammatory mediators such as: ↓ C-reactive protein ↓ Interleukins ↓ Prostaglandins ↓ Tumor necrosis factor alpha ↑Omega-3 index Metabolic By reducing triglycerides and inflammatory [133] disorders markers and improving insulin sensitivity, omega-3s can affect non-alcoholic fatty liver disease. Skin health Maintenance of skin health by increasing [134,135] hydration, reducing sunburn, aging, and skin cancer via photo-protective effects Women Reduced menstrual pain [136] health

ADHD, attention deficit hyperactivity disorder; DHA, docosahexaenoic acid; HDL, high-density lipoprotein

43 KRILL OIL CONCENTRATE Krill Oil: Phospholipids, Choline, And Omega-3 Fatty Acids All In One

WHY KRILL OIL? rine Living Resources) on sustainability To fulfill the global issues. In addition, Aker BioMarine con- demand for omega-3 tinues to be the most proactive player fatty acids with grow- in the krill fishery and has implemented ing pressure on fish voluntary measures. This includes mea- stocks, new sustain- sures such as 100% observer coverage able sources are need- and real-time reporting procedures to ed. Krill oil is such a ensure the continued sustainability of source and WWF-Nor- this fishery. By permitting scientists way clearly says: “Man- onboard at no cost, they are also con- agement of krill is sus- tributing to science and research. The tainable.” krill fishery has been going on for more Nina Jensen, CEO of than 30 years and the catches have WWF-Norway: “WWF-Norway and Aker been relatively stable and low. Howev- BioMarine have worked together since er, WWF-Norway remains concerned 2008 for a sustainable management of that while the precautionary catch limit krill resources in the Antarctica. As a represents a smaller catch limit than result of the partnership and the MSC for most fisheries, it may fail to protect (Marine Stewardship Council) certifi- krill predators at the local scale. It is cation, Aker BioMarine has started an therefore important to have industry extensive mapping of fish larval by- catch, and is working with WWF-Nor- way to document the potential overlap/ conflict between the fishery and land based predators. WWF-Norway and Aker BioMarine are also working ac- tively towards CCAMLR (Commission for the Conservation of Antarctic Ma- Krill live in a very pure environment

44 KRILL OIL CONCENTRATE players, like Aker BioMarine, to commit contaminants, which are often found in to further research to understand the marine life higher in the food chain. complex marine ecosystem in the Ant- The quality of krill oil has arctica.” been further increased by reducing TMA/TMAO and salt contents giving The Marine Stewardship Council (MSC) an almost taste- and smell-free krill oil is an international nonprofit organiza- concentrate. Phospholipid, choline, and tion that focuses on the health of ocean omega-3 fatty acid concentrations are stocks and how they are managed, in increased, providing a special combina- addition to assessing the effects of tion of these important nutrients in the fisheries on the wider ecosystem. The krill oil concentrate. Aker BioMarine krill fishery has been evaluated by an independent assess- UPTAKE OF KRILL OIL IN ment team, and the evaluators have THE BODY certified that the fishery is well man- The evidence that phospholipids are aged, that the krill stock is healthy, and a more effective delivery molecule of that Aker BioMarine’s krill fishery is fatty acids to organs than triglycerides sustainable with minimal impact on the is rising [137-140]. Of particular interest ecosystem. is the accumulation of DHA into the brain, since it makes up about 30-40% While krill oil comes from a sustainable of the fatty acids in the gray matter of source, it has also been found to be pure the cortex and is particularly concen- by extensive analysis for the presence trated in synaptic membranes [141]. In- of contaminants, such as dioxins, fu- deed, in preclinical studies, when given rans, dioxin-like PCBs (polychlorinated single-doses of radiolabeled DHA, the biphenyls), organochlorine pesticides, DHA brain accumulation was twice as PBDEs (polybrominated diphenyl high, when DHA was supplied in phos- ethers), heavy metals, PAHs (polycyclic pholipid rather than triglyceride form aromatic hydrocarbons), arsenic spe- [137,138]. cies, fluoride, trans fatty acids, and marine algal toxins. The krill’s place at the Omega-3 fatty acids from krill oil are bottom of the food chain and its clean efficiently transported and integrat- habitat impede accumulation of these ed into cell membranes, because they

45 KRILL OIL CONCENTRATE are bound to phospholipids. As part of Steady-state, but also in- cell membranes, EPA and DHA have the creased levels of the Omega-3 Index ability to influence fluidity of the mem- after supplementation have been branes, signaling processes, and meta- shown to directly correlate with EPA bolic parameters in the cell. and DHA levels in human cardiac tissue [132,162,163]. In contrast to plasma fatty KRILL OIL IN HEALTH AND acid measurements that reveal short- DISEASE term omega-3 intake [164], the Omega-3 Animal studies that found benefits for Index is believed to mirror overall tis- krill oil supplementation include mod- sue EPA and DHA levels and therefore els for ulcerative colitis [142], depres- a person’s health status. An Omega-3 sion [143], obesity [144-149], myocardial Index of 8% or above is considered op- infarction [150], chronic inflammation timal [165], while a low Omega-3 Index [151], rheumatoid arthritis [152], and glu- indicates a higher risk of sudden cardi- cose tolerance [153]. Highlights from ac death. human studies with krill oil include benefits for a variety of parameters sum- marized in Table 4 .

OMEGA-3 INDEX The Omega-3 Index that has been pro- Since increases in EPA and posed as a novel biomarker for cardio- DHA levels correlate with sudden cardi- vascular risk and is defined as the per- ac death [166], researchers have recent- centage of EPA and DHA in red blood ly turned their investigations toward cell (RBC) fatty acids [161]. krill oil to see if it can increase omega-3 RBC levels. In an unpublished clinical

46 KRILL OIL CONCENTRATE Table 4: Benefits of krill oil in clinical studies.

Parameters Explanation Effects of krill oil References ADHD Attention deficit hy- ↑ Improvement in Unpublished data symptoms peractivity disorder; a behavior and daily neurodevelopmental function capacity psychiatric disorder in children characterized by attention deficits, hyperactivity, and impulsiveness. Cachexia Loss of body mass ↑ Improved blood Unpublished data that cannot be lipids in advanced reversed nutritionally cancer patients (cancer patients) with cachexia and is characterized (TG reduced, by anorexia, muscle HDL increased, wasting, and lipid LDL reduced). profile disorder. Endocannabinoid system parameters normalized and inflammation markers reduced CRP C- reactive protein; ↓ Reduced CRP, [155] CRP blood levels rise inflammation, and in response to inflam- arthritic symp- mation. toms (WOMAC scores) Endocanna- Lipid signaling ↓ Reduced endo- [156] binoids molecules made from cannabinoids in omega-6 fatty acids obese persons involved in a variety of physiological processes including appetite, pain-sensa- tion, mood, and memory. Obese persons have an overactive endocannabinoid system.

47 KRILL OIL CONCENTRATE HDL High-density lipo- ↑ Increased [157,158] protein; the “good” HDL (> 50%) in cholesterol, since subjects with it removes excess elevated blood cholesterol from the fat levels. HDL/ blood and brings it TG ratio improve- to the liver for other ment in healthy uses. subjects Immune Prolonged training ↑ Increased [22,23] function periods may cause a immune function down-regulation of after intense exer- the immune system cise via higher NK by decreasing natural cell and IL-2 levels killer (NK) cells (first ↓ Reduced line of defense to e.g. oxidative damage bacteria and viruses) to red blood cells and interleukin-2 during recovery (IL-2), a signaling mol- from rowing in ecule in the immune national team response. rowers Inflammation Arthritis is an auto- ↓ Reduced [155] immune disease char- inflammation in acterized by inflam- subjects with ar- mation, joint pain, and thritis symptoms swollen joints.

LDL Low-density li- ↓ Decrease or no [157,158] poprotein; LDL is change of LDL often called the “bad cholesterol”, since it picks up cholesterol and fat from the liver and delivers it to artery walls causing atherosclerosis (hardening of the arteries), thereby increasing the risk for heart disease.

48 KRILL OIL CONCENTRATE Omega-3 Percent EPA and ↑ Significant [127,159] index DHA in red blood increase of cells. Correlates with Omega-3 index amount of EPA and DHA in other tissues. Low Omega-3 index is linked to higher risk of heart disease. PMS Premenstrual ↓ Significant re- [160] symptoms syndrome; PMS duction in physical syndrome might have and emotional an inflammatory symptoms related connection. to PMS measured as breast tender- ness, stress, irri- tability, depression, joint pain, bloating, swelling, abdominal pain, and weight gain ↓ Significant reduction in amount of painkillers used for painful cramps Triglyceride A lipid molecule ↓ Significant [157-159] consisting of glycerol reduction of with three fatty acids triglycerides in bound to it. The fatty blood acids can be saturated or unsaturated.

ADHD, attention deficit hyperactivity disorder; CRP, C-reactive protein; DHA, docosahexaenoic acid; EPA, eicosapentaenoic acid; HDL, high-density lipoprotein; IL-2, interleukin-2; LDL, low-density lipoprotein; NK, natural killer; PMS, premenstrual syndrome; TG, triglyceride; WOMAC, Western Ontario and McMaster Universities Arthritis Index

49 KRILL OIL CONCENTRATE a central biomarker of heart disease risk [167]. In this study, a total of 300 volunteers were divided into five groups and supplemented with krill oil at either 0.5, 1, 2 or 4 grams per day or placebo (olive oil). The individuals included in the study had blood triglyceride values between 150 and 499 mg/dL. Blood lipids were measured at baseline, 6 weeks and 12 weeks of treatment. study, healthy volunteers took either 2 Relative to subjects in the grams of Superba™ krill oil for 8 weeks placebo group, those administered or 2 grams of an omega-3 enriched krill oil had a statistically significant fish oil. The goal of the study was to 10% reduction in serum triglycerides. compare the delivery of omega-3 fatty Moreover, LDL cholesterol levels were acids, PLs versus TGs, to see if the mo- not increased in the krill oil groups rela- lecular form influences the increase in tive to the placebo group, an important Omega-3 Index. The results of the study finding considering an increase in LDL showed that krill oil increased the Ome- cholesterol has been observed in some ga-3 Index significantly more than fish fish oil trials [159]. Additionally, the sub- oil after 8 weeks of supplementation. jects taking 4 grams of krill oil per day In fact, krill oil increased the Omega-3 raised their Omega-3 Index from 3.7% Index about 70% more than fish oil at the end of study after dose adjustment between the two treatment groups. These results go hand in hand with another study, which investigated the effect of 12 weeks daily Superba™ krill oil intake in volunteers with “borderline high” or “high” blood triglyceride levels [159]. Triglycerides have an important role in lipid metabolism and are

50 KRILL OIL CONCENTRATE to 6.3%. Comparable increases in the mize omega-3 fatty acid, phospholipid, Omega-3 Index have been linked to de- and choline status in the body. The all- creased risk for sudden cardiac death in-one advantage of krill oil delivering in previous studies – in a prospective simultaneously omega-3 fatty acids, cohort study by about 80% [168] and in phospholipids, and choline gives it a a case control study by 90% [169]. unique standpoint in the omega-3 mar- Thus, there seem to be ad- ket. However, the full elucidation of how vantages of krill oil, when it comes to much each component benefits human raising a person’s Omega-3 Index. Most health must await further study. importantly, the mentioned studies show that krill oil more effectively rais- ACKNOWLEDGEMENTS es the Omega-3 Index compared to fish I am very grateful for editing help from oil, even though krill oil delivers lower Gunhild Yksnøy and Michel Lockhart amounts of EPA and DHA on a gram per and to everyone proofreading the text. gram basis compared to fish oil. Clear A special thanks for the contribution health benefits, in particular for heart of tables from Tove Julie Evjen and health, have been shown by raising one’s Wenche Rasch. Omega-3 Index higher than 8%.

CONCLUSION While Superba™ krill oil has demonstrated its health benefits in numerous studies, the new krill oil concentrate holds even more promise in particular in relation to health concerns that are characterized by phospholipid and/or choline deficiency. Reduced phospholipid and choline levels are seen in e.g. liver and brain disorders, heart disease, intestinal inflammation, lung and skin diseases, or after intense exercise. Krill oil supplementation can help to heal membranes and to opti-

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63 KRILL OIL CONCENTRATE 64 KRILL OIL CONCENTRATE