WO2013173971A1 - Use of fructus ligustri lucidi in the manufacture of a medicament for the prevention or treatment of a condition linked to weak bones in a young subject - Google Patents
Use of fructus ligustri lucidi in the manufacture of a medicament for the prevention or treatment of a condition linked to weak bones in a young subject Download PDFInfo
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- WO2013173971A1 WO2013173971A1 PCT/CN2012/075870 CN2012075870W WO2013173971A1 WO 2013173971 A1 WO2013173971 A1 WO 2013173971A1 CN 2012075870 W CN2012075870 W CN 2012075870W WO 2013173971 A1 WO2013173971 A1 WO 2013173971A1
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- Prior art keywords
- ligustri lucidi
- fructus ligustri
- young
- bones
- bone
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Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/63—Oleaceae (Olive family), e.g. jasmine, lilac or ash tree
- A61K36/638—Ligustrum, e.g. Chinese privet
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P19/00—Drugs for skeletal disorders
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P19/00—Drugs for skeletal disorders
- A61P19/08—Drugs for skeletal disorders for bone diseases, e.g. rachitism, Paget's disease
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P19/00—Drugs for skeletal disorders
- A61P19/08—Drugs for skeletal disorders for bone diseases, e.g. rachitism, Paget's disease
- A61P19/10—Drugs for skeletal disorders for bone diseases, e.g. rachitism, Paget's disease for osteoporosis
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
Definitions
- the present invention relates to Fructus Ligustri Lucidi for use in the prevention or treatment of a condition linked to weak bones in young subjects.
- the invention also relates to a non-therapeutic use of Fructus Ligustri Lucidi for increasing calcium content in bones of young subjects. Unhealthy lifestyles that are lacking balanced nutrition, physical exercise or sunshine are putting more and more
- Bones are important as they give shape and support the body. They are living tissues that rebuild constantly throughout life. During the growth phase of a body, i.e. childhood and adolescence of a human subject, the body adds new bone mass faster than it removes old one, while about after the age of between 20 and 30 years, bone mass can be lost faster than it is actually build up.
- Bone mineral density is thereby a good and accepted test for measuring the strength and health of bones.
- a low BMD value indicates that bones are weak and more likely to break.
- low BMD may lead to a lower peak bone mass, which is generally achieved by early adulthood. Peak bone mass is an important determinant of an adult risk of further
- osteoporosis or osteopenia later in life.
- cholesterol level may have adversely affected adolescent bone mass. This may also explain the
- healthy diet including foods rich in calcium and vitamin D, should do regular physical exercise e.g. a sports activity and should reduce their body weight to normal where overweight.
- estrogen especially when started soon after menopause, can help maintaining bone density.
- the object of the present invention is to address the problem as stated above, to improve the state of the art and to provide a food and/or medical solution to prevent or treat conditions linked to weak bones in children and adolescents and which helps them to increase the calcium content and strength of their bones e.g. for building-up an increased peak bone mass before entering adulthood.
- the present invention provides in a first aspect Fructus Ligustri Lucidi for use in the prevention or treatment of a condition linked to weak bones in a young subject.
- the invention relates to a non-therapeutic use of Fructus Ligustri Lucidi for increasing the calcium content in bones of a young subject.
- a third aspect of the invention pertains to a food product or a food supplement comprising an alcohol extract from Fructus Ligustri Lucidi.
- a still further aspect of the invention pertains to a pet food product comprising Fructus Ligustri Lucidi.
- mice received a diet supplemented with about 700 mg/kg body weight of a Fructus Ligustri Lucidi ethanol extract per day. Body weight and food intake of the rats was weekly monitored. After 12 weeks of the study period the rats were sacrificed, the femur and tibia collected and analysed.
- the present invention pertains to Fructus Ligustri Lucidi for use in the prevention or treatment of a condition linked to weak bones in a young subject, particularly wherein the condition is rickets, bone atrophy, bone tumor or cancer, brittle bone disease, osteogenesis imperfecta, vitamin D deficiency or growth deficiency.
- Fineness Ligustri Lucidi also known as glossy pivet fruit is the dried ripe fruit of Ligustrum lucidum (Oleaceae family) , a wild-growing ever-green small tree native in China and Korea The dried ripe fruit is the part of the plant used in
- Rickets is among the most frequent childhood diseases in many developing countries .
- Bone atrophy is a decrease in bone mass. Bone atrophy may result from inactivity when calcium is withdrawn from the bone making it less dense, weaker, and more susceptible to
- Osteogenesis imperfecta is the most common disease causing fractures in childhood. Osteogenesis imperfecta is a genetic disorder usually resulting from abnormalities of the genes, which control the production of a protein called collagen. This is the main protein in bone and essential for its strength .
- “Growth deficiency” refers to a slow or flat rate of growth in children which is a condition in which a child is not growing as expected.
- the Fructus Ligustri Lucidi for use as of the invention pertains to a young subject, wherein the young subject is a child or growing-up adult from 1 to 20 years of age,
- the young subject is still in the growing-up phase and at an age where he or she has not yet completely build-up the peak bone mass and/or where the bones are still growing.
- the Fructus Ligustri Lucidi for use as of the invention pertains to a young subject, wherein the young subject is a young animal, preferably a young cat or a young dog. Very similar as young humans, also young animals can suffer from weak bones as has been reported for certain livestock animals, chickens and animals kept as pets such as young dogs or young cats.
- the Fructus Ligustri Lucidi for use as of the invention the Fructus Ligustri Lucidi is provided in the form of an extract, preferably an extract with an alcohol, even more preferably an extract with ethanol .
- the dried fruit of Fructus Ligustri Lucidi is not very attractive in a direct use in a food product for human consumption because of organoleptic reasons.
- the active preferably an extract with an alcohol, even more preferably an extract with ethanol .
- ingredients can be extracted from the Fructus Ligustri Lucidi dried fruits by extraction for example with water or alcohol.
- a water extract of those fruits in the form of a medical tea or decoction is well known mainly from the traditional Chinese medicine.
- the Fructus Ligustri Lucidi for use as of the invention is provided in a dose of from 4 to 20 g/day, preferably from 6 to 15 g/day to a young subject.
- the identified dose of from 4 to 20 g/day or preferably from 6 to 15 g/day of the original fruit or herb provides sufficient active ingredient from the Fructus Ligustri Lucidi to a young subject for the claimed health benefit to be effective.
- the Fructus Ligustri Lucidi is provided in the form of an extract, wherein the extract is adjusted to a dose which is equivalent to a consumption of 4 to 20 g/day, preferably of 6 to 15 g/day of said Fructus fruit.
- the Fructus Ligustri Lucidi for use as of the invention is provided in combination with calcium.
- Fructus Ligustri Lucidi improves a.o. the
- the calcium is thereby provided in a daily dose of from 500 to 1' 500 mg, preferably from 800 to 1'300 mg .
- the invention pertains to a non-therapeutic use of Fructus Ligustri Lucidi for increasing the calcium content in bones of a young subject, preferably for increasing the strength of the bones and/or improving growth in length of the bones.
- the invention is not only
- the invention pertains to a food product or a food supplement comprising an alcohol extract from Fructus Ligustri Lucidi. This has the advantage that the food product does not need to contain whole fruit matter of the Fructus Ligustri Lucidi per se which is not very
- an alcohol extract of said fruit product comprises the active ingredient in a concentrated form which can be easily added or
- the alcohol is not present anymore in the extract when the extract is used in the manufacturing for example of a food product.
- the invention pertains also to a food product or food supplement comprising an alcohol extract of Fructus Ligustri Lucidi, wherein the alcohol extract is in a dry or powdered form.
- a still further aspect of the invention pertains to a pet food product comprising Fructus Ligustri Lucidi.
- Fructus Ligustri Lucidi For many animals the taste or organoleptic property of the whole fruit of
- Fructus Ligustri Lucidi may not be a problem for consumption.
- pet food products comprising such Fructus whole fruit matters can be produced more easily and cheaper as when first extracts from the Fructus fruit need to be prepared and then incorporated into such pet foods .
- Example 1 Weaning female Sprague-Dawley (SD) rats, purchased from the Department of Laboratory Animal Science, Peking University Health Science Center, were used for the experiment. The animals were of specific pathogen-free (SPF) degree. They were housed in temperature and humidity controlled rooms (22 ⁇ 2°C, 50-60% humidity) with a 12h:12h light/dark cycle. Animals were acclimatized for one week before formal experiment. After one week, animals were randomized into 2 groups (12 per group) : A) Group 1: fed by AIN-93G diet. This Group is control group. B) Group 2: fed by AIN-93G diet with Fructus Ligustri Lucidi (FLL) ethanol extract (700 mg/kgBW)
- FLL Fructus Ligustri Lucidi
- the serum calcium and phosphorus content was determined by colorimetry using commercially available test kits (Sigma, St).
- Coupled Plasma mass spectrometer (ICP-MS) (PE-Sciex DRC, PerkinElmer, USA) . Calcium intake was estimated from food intake. Urinary Ca was determined by colorimetry using
- Ca absorption rate (%) (Ca intake - fecal Ca) / Ca intakexlOO;
- Ca balance Ca intake- (urinary Ca + fecal Ca) ;
- Ca retention rate (%) Ca balance / Ca
- the femur and tibia from right side were heated in oven at 105 ° C to constant weight and the dry weight was measured by analytical balance.
- the length and diameter at mid-point location of femurs were measured by a vernier caliper.
- lanthanum oxide After making up to the volume, calcium content was determined according to the steps of instrument manual of the atomic absorption spectrophotometer used (PE— Zeemam-5100, PerkinElmer, USA) .
- the BMD was measured at the distal end of the femur (region 1 cm in length from the most distal point of the bone) , femoral neck and femoral diaphysis (mid-diaphyseal region) ,
- left femurs were used for a three-point bending mechanical test (FPZ100, Germany) to
- the average food Ca content in control and FLL group was 5.65mg/g and 5.63mg/g, respectively.
- Ca intakes in the intervention group were higher than in that of the control group.
- Ca absorption rate and retention rate in the FLL group were significantly higher than that of the control group (Table 1) .
- the femoral dry weight of FLL group was significantly higher than that of the control group (Table 2) .
- BMP Femoral bone mass density
- the BMD of neck, distal end and femoral diaphysis of the femurs in the FLL group were all significantly higher than those of the control group (Table 5) .
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- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Physical Education & Sports Medicine (AREA)
- Chemical & Material Sciences (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- Medicinal Chemistry (AREA)
- General Health & Medical Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- Pharmacology & Pharmacy (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Natural Medicines & Medicinal Plants (AREA)
- Orthopedic Medicine & Surgery (AREA)
- Rheumatology (AREA)
- Alternative & Traditional Medicine (AREA)
- Biotechnology (AREA)
- Botany (AREA)
- Medical Informatics (AREA)
- Microbiology (AREA)
- Mycology (AREA)
- Epidemiology (AREA)
- Coloring Foods And Improving Nutritive Qualities (AREA)
- Medicines Containing Plant Substances (AREA)
Abstract
Use of Fructus Ligustri lucidi in the manufacture of a medicament for the prevention or treatment of a condition linked to weak bones in a young subject is disclosed. Non-therapeitic use of Fructus Ligustri lucidi for increasing calcium content in bone of young subjects, as well as a food product or food supplement comprising an alcohol extract of Fructus Ligustri lucidi and a pet food comprising said Fructus Ligustri lucidi are also provided.
Description
USE OF FRUCTUS LIGUSTRI LUCIDI IN THE MANUFACTURE OF A MEDICAMENT
FOR THE PREVENTION OR TREATMENT OF A CONDITION LINKED TO WEAK BONES IN A YOUNG SUBJECT
The present invention relates to Fructus Ligustri Lucidi for use in the prevention or treatment of a condition linked to weak bones in young subjects. The invention also relates to a non-therapeutic use of Fructus Ligustri Lucidi for increasing calcium content in bones of young subjects. Unhealthy lifestyles that are lacking balanced nutrition, physical exercise or sunshine are putting more and more
children and young adolescents at risk of developing weak bones, and exposing them thereby to susceptibility of bone fractures and bone diseases in young age. Only in the USA and still today, millions of children are not building bones as strong as they should be, which leaves them more vulnerable to fractures, rickets and other bone conditions. Certain bone diseases such as e.g. osteogenesis imperfecta can also make bones weak and brittle that easily fracture or break.
Bones are important as they give shape and support the body. They are living tissues that rebuild constantly throughout life. During the growth phase of a body, i.e. childhood and adolescence of a human subject, the body adds new bone mass faster than it removes old one, while about after the age of between 20 and 30 years, bone mass can be lost faster than it is actually build up.
Strong bones are very important for health of a subject. Bone mineral density (BMD) is thereby a good and accepted test for measuring the strength and health of bones. A low BMD value, for example indicates that bones are weak and more likely to break. Furthermore, low BMD may lead to a lower peak bone mass,
which is generally achieved by early adulthood. Peak bone mass is an important determinant of an adult risk of further
medical conditions such as osteoporosis or osteopenia later in life.
Recently, there has been growing evidence that childhood
obesity negatively affects bone development (N. Pollock et al . , 2011, J Pediatr. 158: 727-734). It has been observed that children being overweight with metabolic abnormalities, such as insulin resistance, low high-density lipoprotein
cholesterol level, and visceral adiposity, may have adversely affected adolescent bone mass. This may also explain the
observed more frequent skeletal bone fractures in overweight versus normal children (P. Dimitri et al . , 2009, J Bone
Mineral Res. 25:527-536). Furthermore, pre-diabetes diagnosed in pre-pubertal overweight children was reported to be
associated with lower bone mass and weaker bones in those children, which may be at risk for poor skeletal development (N. Pollock et al., 2010, J Bone Mineral Res. 25:2760-2769).
Hence, today there is an increasing number of children and adolescents with weak bones, with associated risks of bone fractures, and risk of bone diseases some of which may
manifest already at a young age some of them only very late in life. The National Institute of Health (NIH) and many other Organisations recommend that young children should eat a
healthy diet including foods rich in calcium and vitamin D, should do regular physical exercise e.g. a sports activity and should reduce their body weight to normal where overweight.
Besides those recommendations, only very limited medical
interventions are available today to address the problem of weak bones in children and helping them to strengthen their
bones and e.g. building-up the BMD while still in the growing- up age.
On the other hand, medical and food based solutions for the prevention or treatment of bone weakening in adult subjects particularly of osteoporosis, such as e.g. for menopausal women, are available. The goal of a treatment of osteoporosis in adults is the prevention of bone fractures by reducing the loss of bone due to the aging process. Osteoporosis results when the rate of bone resorption exceeds the rate of bone rebuilding. Besides recommendations for patients suffering from osteoporosis to adapt their lifestyle to more regular physical exercise, a balanced diet with calcium and vitamin D, less smoking and drinking alcohol etc., medical treatments are available such as different anti-resorptive agents which decrease the removal of calcium from the bones,
bisphosphonates or hormone-related therapies. Thereby, estrogen, especially when started soon after menopause, can help maintaining bone density.
Further solutions which are based on traditional foods or food supplements to treat or prevent bone disorders and
particularly osteoporosis in aging adults have been proposed (Review article: S. Putnam et al., 2007, Phytother. Res. 21: 99-112) . Thereby good evidence has been provided that certain dietary components and herbal products can influence the balance between mineralization and resorption of calcium from bones in adult persons. For example, it has been reported that a number of vegetables, including onion, garlic and parsely, can inhibit bone resorption in ovariectomized rats. Essential oils from sage, rosemary, thyme and other herbs inhibit osteoclast activity and thereby lead to increased bone mineral density. Soya rich in isoflavones appears to have an estrogen-
like effect and has shown promising results in maintaining bone health. Various traditional herbal formulas in Chinese medicine have also demonstrated effects in pharmacological models of osteoporosis. Thereby, for example, an extract of Fructus Ligustri Lucidi has shown a decreased bone loss effect in an ovariectomized rat model (Y. Zhang et al . , 2008, British J Nutrition 99: 494-502). So far, however, none of those traditional foods, food supplements or Chinese herbs have been demonstrated to work also in young subjects during their growth phase and before completion of their peak bone mass in young age. In fact, the use of those foods and herbs has been investigated with the purpose of correcting the balance between the accelerated rate of bone loss due to the age and the re-mineralization of those bones in the aged adult
subjects. Furthermore, most of the currently known plant based extracts are based or are expected to be based on hormone like substances, like e.g. estrogen-like substances, and are not to be expected to be effective or suitable for young subjects for building-up the initial peak bone mass before adulthood.
The object of the present invention is to address the problem as stated above, to improve the state of the art and to provide a food and/or medical solution to prevent or treat conditions linked to weak bones in children and adolescents and which helps them to increase the calcium content and strength of their bones e.g. for building-up an increased peak bone mass before entering adulthood.
The object of the present invention is achieved by the subject matter of the independent claims. The dependent claims further develop the idea of the present invention.
Accordingly, the present invention provides in a first aspect Fructus Ligustri Lucidi for use in the prevention or treatment of a condition linked to weak bones in a young subject.
In a second aspect, the invention relates to a non-therapeutic use of Fructus Ligustri Lucidi for increasing the calcium content in bones of a young subject.
A third aspect of the invention pertains to a food product or a food supplement comprising an alcohol extract from Fructus Ligustri Lucidi.
A still further aspect of the invention pertains to a pet food product comprising Fructus Ligustri Lucidi.
It has been surprisingly found by the inventors that when an extract of Fructus Ligustri Lucidi is provided in the food to young weaning female rats during their first 4 months of life, i) calcium provided in the food is much better retained by those animals, ii) the bones were thicker and heavier, iii) the calcium content in those bones was increased, and iv) the biomechanical strength of those bones were increased, in comparison to a control group kept under same conditions and same food supply but without the above mentioned extract. In particular, the weaning rats used in the experiment were randomized into groups already four weeks after birth and then fed with the supplemented food for the experimental period which lasted 12 weeks. These 1 to 4 months of age of those animals tested correspond to about 3 to 12 years of human age (Australian Rat Information Forum) . The rats of the
experimental group received a diet supplemented with about 700 mg/kg body weight of a Fructus Ligustri Lucidi ethanol extract per day. Body weight and food intake of the rats was weekly
monitored. After 12 weeks of the study period the rats were sacrificed, the femur and tibia collected and analysed.
Thereby it was determined that the dry weight as well as the calcium content of the femurs of the experimental group was significantly increased over that of the control group.
Details of the animal study and obtained results are further disclosed under the Example section below.
The present invention pertains to Fructus Ligustri Lucidi for use in the prevention or treatment of a condition linked to weak bones in a young subject, particularly wherein the condition is rickets, bone atrophy, bone tumor or cancer, brittle bone disease, osteogenesis imperfecta, vitamin D deficiency or growth deficiency.
"Fructus Ligustri Lucidi" also known as glossy pivet fruit is the dried ripe fruit of Ligustrum lucidum (Oleaceae family) , a wild-growing ever-green small tree native in China and Korea The dried ripe fruit is the part of the plant used in
traditional Chinese Medicine.
"Rickets" is a softening of bones in children due to
deficiency or impaired metabolism of vitamin D,
magnesium, http://en.wikipedia.orR/wiki/Rickets - cite note-0 phosphorus or calcium, potentially leading to fractures and deformity.
Rickets is among the most frequent childhood diseases in many developing countries .
"Bone atrophy" is a decrease in bone mass. Bone atrophy may result from inactivity when calcium is withdrawn from the bone making it less dense, weaker, and more susceptible to
fractures.
"Brittle bone disease" also sometimes refered to as "Osteogenesis imperfecta" is the most common disease causing fractures in childhood. Osteogenesis imperfecta is a genetic disorder usually resulting from abnormalities of the genes, which control the production of a protein called collagen. This is the main protein in bone and essential for its strength .
"Growth deficiency" refers to a slow or flat rate of growth in children which is a condition in which a child is not growing as expected.
The Fructus Ligustri Lucidi for use as of the invention pertains to a young subject, wherein the young subject is a child or growing-up adult from 1 to 20 years of age,
preferably from 3 to 18 years of age or preferably from 3 to 15 years of age. Particularly, the young subject is still in the growing-up phase and at an age where he or she has not yet completely build-up the peak bone mass and/or where the bones are still growing.
In a further embodiment, the Fructus Ligustri Lucidi for use as of the invention pertains to a young subject, wherein the young subject is a young animal, preferably a young cat or a young dog. Very similar as young humans, also young animals can suffer from weak bones as has been reported for certain livestock animals, chickens and animals kept as pets such as young dogs or young cats. In a preferred embodiment, the Fructus Ligustri Lucidi for use as of the invention, the Fructus Ligustri Lucidi is provided in the form of an extract, preferably an extract with an alcohol, even more preferably an extract with ethanol . The
dried fruit of Fructus Ligustri Lucidi is not very attractive in a direct use in a food product for human consumption because of organoleptic reasons. However, the active
ingredients can be extracted from the Fructus Ligustri Lucidi dried fruits by extraction for example with water or alcohol. Thereby, a water extract of those fruits in the form of a medical tea or decoction is well known mainly from the traditional Chinese medicine. The extraction of the active ingredients from those fruits with an alcohol, and
particularly with ethanol, seems to be more efficient than the extraction with water and allows to more concentrating those still undefined active ingredients for consumption. The extraction and concentration with ethanol has thereby the advantage that it provides a very efficient extraction method for those active ingredients and that the use of ethanol for extraction purposes for use in the manufacturing of food products is very safe and well accepted by today's consumers.
In a preferred embodiment, the Fructus Ligustri Lucidi for use as of the invention is provided in a dose of from 4 to 20 g/day, preferably from 6 to 15 g/day to a young subject. The identified dose of from 4 to 20 g/day or preferably from 6 to 15 g/day of the original fruit or herb provides sufficient active ingredient from the Fructus Ligustri Lucidi to a young subject for the claimed health benefit to be effective.
Preferably, the Fructus Ligustri Lucidi is provided in the form of an extract, wherein the extract is adjusted to a dose which is equivalent to a consumption of 4 to 20 g/day, preferably of 6 to 15 g/day of said Fructus fruit.
In a further preferred embodiment, the Fructus Ligustri Lucidi for use as of the invention is provided in combination with calcium. As it has been found by the inventors and disclosed
herein, that Fructus Ligustri Lucidi improves a.o. the
incorporation of calcium into the bones of young subjects. It is therefore advantageous to provide the necessary and
sufficient level of calcium to said subjects directly together with the Fructus fruit as part for example of the daily diet. Preferably, the calcium is thereby provided in a daily dose of from 500 to 1' 500 mg, preferably from 800 to 1'300 mg .
In another aspect, the invention pertains to a non-therapeutic use of Fructus Ligustri Lucidi for increasing the calcium content in bones of a young subject, preferably for increasing the strength of the bones and/or improving growth in length of the bones. Advantageously, the invention is not only
applicable to young subjects with a medical condition linked to weak bones, but can also be applied to any healthy young subject for helping him or her to building-up the calcium content of the bones. This leads a.o. to stronger bones in the young age which are less at risk to fracture or break; it supports a healthy growth and development of the bones during the growth phase of the subject; it supports and improves the building-up of a maximal peak bone mass after the young subject has reached the end of his or her active growth phase and enters adulthood, and thereby also leads to stronger bones during adulthood with a lesser risk of fractures or bone breaks later in life.
In a still further aspect, the invention pertains to a food product or a food supplement comprising an alcohol extract from Fructus Ligustri Lucidi. This has the advantage that the food product does not need to contain whole fruit matter of the Fructus Ligustri Lucidi per se which is not very
attractive to human consumption because of taste and other organoleptic properties of these fruits. Instead, an alcohol
extract of said fruit product comprises the active ingredient in a concentrated form which can be easily added or
incorporated into a food product or food supplement without affecting its taste or organoleptic quality too much.
Preferably, the alcohol is not present anymore in the extract when the extract is used in the manufacturing for example of a food product. This has the advantage that such a food product can be safely used also for younger children below the age of 16 years. Therefore, the invention pertains also to a food product or food supplement comprising an alcohol extract of Fructus Ligustri Lucidi, wherein the alcohol extract is in a dry or powdered form.
A still further aspect of the invention pertains to a pet food product comprising Fructus Ligustri Lucidi. For many animals the taste or organoleptic property of the whole fruit of
Fructus Ligustri Lucidi may not be a problem for consumption. Advantageously, pet food products comprising such Fructus whole fruit matters can be produced more easily and cheaper as when first extracts from the Fructus fruit need to be prepared and then incorporated into such pet foods .
Those skilled in the art will understand that they can freely combine all features of the present invention disclosed herein. In particular, features described for the medical use of the present invention may be combined with the non-therapeutic use of the present invention and vice versa. Further, features described for different embodiments of the present invention including the food products and supplements may be combined. Further advantages and features of the present invention are apparent from the figures and examples .
Example 1 :
Weaning female Sprague-Dawley (SD) rats, purchased from the Department of Laboratory Animal Science, Peking University Health Science Center, were used for the experiment. The animals were of specific pathogen-free (SPF) degree. They were housed in temperature and humidity controlled rooms (22±2°C, 50-60% humidity) with a 12h:12h light/dark cycle. Animals were acclimatized for one week before formal experiment. After one week, animals were randomized into 2 groups (12 per group) : A) Group 1: fed by AIN-93G diet. This Group is control group. B) Group 2: fed by AIN-93G diet with Fructus Ligustri Lucidi (FLL) ethanol extract (700 mg/kgBW)
Animals were provided deionised sterile water throughout the research. Both food and water were administered ad libitum. During the experimental period, the body weight and food intake were measured once a week. After 12 weeks, rats were placed into metabolic cages for 4 days : 1-day adaptation and 3-day sample collections of urine and feces. At the end of the metabolic experiment, blood was withdrawn from arteria
femoralis under the anaesthesia with Pentobarbital sodium (30 mg/kg BW) , and then the rats were sacrificed. Serum was prepared and stored at -80°C for biochemical determinations. The femur and tibia of both sides were collected, cleaned of all soft tissue, and stored at -20°C for analysis.
Methods
Analysis of serum markers of bone metabolism:
The serum calcium and phosphorus content was determined by colorimetry using commercially available test kits (Sigma, St
Louis, MO, USA) .
Calcium balance experiments:
Each animal was put in the metabolic cage individually for 4 continuous days. The urine and feces were collected separately from day 2 to day 4. Food intake of each animal was recorded
every day. The feces were dried at 120 °C and milled. The feces ash and food samples from each group were dissolved by 6 M HC1. Then calcium content was measured by Inductively
Coupled Plasma mass spectrometer (ICP-MS) (PE-Sciex DRC, PerkinElmer, USA) . Calcium intake was estimated from food intake. Urinary Ca was determined by colorimetry using
commercially available kit (Sigma, St Louis, MO, USA) . The absorption and retention rates of calcium were calculated by the following equations: Ca absorption rate (%) = (Ca intake - fecal Ca) / Ca intakexlOO; Ca balance = Ca intake- (urinary Ca + fecal Ca) ; Ca retention rate (%) = Ca balance / Ca
intakexlOO
Dry weight, length and diameter of bones:
The femur and tibia from right side were heated in oven at 105 °C to constant weight and the dry weight was measured by analytical balance. The length and diameter at mid-point location of femurs were measured by a vernier caliper.
Calcium content of femur:
The dried right femur was digested in a flask. According to the calcium content of the sample, 0.300 - 1.00 g sample was weighed, placed in 150 mL Erlenmeyer flask, added mixed acid (nitric acid: perchloric acid = 4:1) 15 - 20 mL, covered with a small funnel and heated on a electric hot plate until white smoke was generated and the digestion liquid was colorless. A few milliliters of deionized water was added to the colorless digestive juice, then boiled to drive out the remaining acid, and this was repeated twice. Made sure the last volume of digestive juice was no more than 1 ml. During the digestion process, blank control was done at the same time, which was equivalent volume of mixed acid and processed under the same conditions as the samples. Sample solution for detection, standard solution and blank was all diluted using 0.5%
lanthanum oxide. After making up to the volume, calcium
content was determined according to the steps of instrument manual of the atomic absorption spectrophotometer used (PE— Zeemam-5100, PerkinElmer, USA) .
Bone densitometry:
After soft tissue cleaning, the left femur of each rat was subjected to densitometric measurements of areal mineral
density (dual-energy X-ray absorptiometry; Norland, CRANBURY, NJ, USA) . For the measurements, the bones were placed on their posterior surfaces in a thin-walled plastic container filled with 0.9% NaCl and scanned with built-in small animal mode.
The BMD was measured at the distal end of the femur (region 1 cm in length from the most distal point of the bone) , femoral neck and femoral diaphysis (mid-diaphyseal region) ,
respectively. The coefficient of variation (CV) for the
measurements (determined by 30 separate scans) was < 3%. All measurements were performed by the same investigator.
Biomechanical testing of the femurs:
After bone densitometry analysis, left femurs were used for a three-point bending mechanical test (FPZ100, Germany) to
determine maximal load (N, a measure of the maximum force that the bone withstood before fracture) and stiffness (N/mm, a measure of the extrinsic rigidity of the bone) . Each specimen was placed on two supports spaced 15 mm apart and the load was applied to the bone midway between the supports at a
deformation rate of 2 mm/min until fracture occurred. The femurs were positioned such that bending occurred about the medial-lateral axis. Load-deformation curves were recorded during bending process. The parameters ultimate load, ultimate stress and stiffness were determined using Series IX (v8.08.00) software. In this study, ultimate load was the weight which caused the bones failure or breakage.
Results :
Calcium (Ca) balance
The average food Ca content in control and FLL group was 5.65mg/g and 5.63mg/g, respectively.
Ca intakes in the intervention group were higher than in that of the control group. Ca absorption rate and retention rate in the FLL group were significantly higher than that of the control group (Table 1) .
Table 1: Effect of FLL extracts on Ca retention
* P < 0.05 versus control
Dry weight, length and diameter of bones
The femoral dry weight of FLL group was significantly higher than that of the control group (Table 2) .
Values are expressed as mean
** P < 0.01 versus control
Femoral calcium contents and bone mineral content
The femora calcium content within the FLL group were significantly higher than that within control group (Table 3 and 4) . Table 3: Effect of FLL extracts on femoral calcium contents
Values are expressed as mean ± SD.
** P < 0.01 versus control
Values are expressed as mean ± SD.
** P < 0.01 versus control
Femoral bone mass density (BMP)
The BMD of neck, distal end and femoral diaphysis of the femurs in the FLL group were all significantly higher than those of the control group (Table 5) .
Values are expressed as mean ± SD.
*P<0.05 versus control ** P < 0.01 versus control
Biomechanical testing of the femurs
The results of the three-point bending test showed that ultimate load and stress of the FLL group was significantly increased versus control group (Table 6) .
Table 6: Results of biomechanical on each group
Values are expressed as mean
*P<0.05 versus control ** P < 0.01 versus control
Claims
Fructus Ligustri Lucidi for use in the prevention or treatment of a condition linked to weak bones in a young subj ect .
The Fructus Ligustri Lucidi for use as of claim 1, wherein the condition is rickets, bone atrophy, bone tumor or cancer, brittle bone disease, osteogenesis imperfecta, vitamin D deficiency or growth deficiency.
The Fructus Ligustri Lucidi for use as of claim 1 or 2, wherein the young subject is a child or growing-up adult from 1 to 20 years of age, preferably from 3 to 18 years of age or preferably from 3 to 15 years of age.
The Fructus Ligustri Lucidi for use as of claim 1 or 2, wherein the young subject is a young animal, preferably a young cat or a young dog.
The Fructus Ligustri Lucidi for use as of one of the above claims, wherein the Fructus Ligustri Lucidi is provided in the form of an extract, preferably an extract with an alcohol .
The Fructus Ligustri Lucidi for use as of claim 5, wherein the alcohol is ethanol .
The Fructus Ligustri Lucidi for use as of one of the above claims, wherein the Fructus Ligustri Lucidi is provided
in a dose of from 4 to 20 g/day, preferably from 6 to 15 g/day .
The Fructus Ligustri Lucidi for use as of one of the above claims, wherein the Fructus Ligustri Lucidi is provided in combination with calcium.
The Fructus Ligustri Lucidi for use as of one of the above claims, wherein the calcium is provided in a daily dose of from 500 to 1'500 mg, preferably from 800 to 1'300 mg .
10. Non-therapeutic use of Fructus Ligustri Lucidi for
increasing the calcium content in bones of a young subj ect .
11. The non-therapeutic use of Fructus Ligustri Lucidi as of claim 10, for increasing the strength of bones.
12. The non-therapeutic use of Fructus Ligustri Lucidi as of claim 10 or 11, for improving length growth of bones in a young subject.
13. The non-therapeutic use of Fructus Ligustri Lucidi as of one of claims 10 to 12, wherein the young subject is in the growth phase.
14. A food product or food supplement comprising an alcohol extract from Fructus Ligustri Lucidi.
15. A pet food product comprising Fructus Ligustri Lucidi.
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