DIARRHOEAS

Technical Field

The subject of the invention is the prevention and reduction of diarrhoeas, particularly in travellers going to countries of elevated risk.

Background Art

Travellers' diarrhoea is a complex of medical conditions of a similar course and infection aetiology (bacteria, viruses). It occurs in about 20-50% of tourists from developed countries spending usually 1-2 weeks in countries of lower hygienic standards, especially in hot climate. The main cause of the ailment is high sensitivity to local intestine pathogens, usually absent in the country of the sick, but widely spread in the country of their stay due to the environmental pollution, insufficient access to clean water and improper sanitary and hygienic conditions.

Travellers' diarrhoea typically regresses by itself and needs no causal treatment, but it may impair the functioning of the individuals affected and prevent them from carrying out their professional or holiday plans. Nevertheless, the clinical course of the ailment varies greatly and is individual in character. In some, especially more sensitive persons (children, the elderly, presence of other diseases), it may take on very acute forms.

Travellers' diarrhoea is a common condition experienced by people travelling to countries of warm and hot climate. The population at risk of travellers' diarrhoea in Poland is about 2 million people annually; it is estimated that 300 000 - 500 000 people actually develop it. In the vast majority of cases the treatment is based on preparations taken from home or purchased by the sick at the place of their stay.

The market lacks an efficient substance that would decrease the risk of developing travellers' diarrhoea and have a therapeutic effect in case of the occurrence of the condition.

The preparations in use decrease the symptoms (peristalsis-paralysing drugs) In some cases their short-term beneficial effects are accompanied by negative effects on the whole system or on the saprophytic intestinal flora (antibiotics).

An optimal preparation should exert an immediate effect (absorb toxins, decrease secretion of water and electrolytes into the intestinal lumen, eliminate pathogens) accompanied by anti-inflammatory and regenerative activity in the area of the mucous membrane of the intestines.

The classic definition to diagnose travellers' diarrhoea includes the presence of three or more unformed or poorly formed stools over 24 hours co-occurring with at least one accompanying symptom, such as: nausea, vomiting, stomach-ache, rectal tenesmus, fever, bloody stool [1]. According to the definition of UNICEF WHO, in order to diagnose travellers' diarrhoea it is enough to find three or more unformed stools with or without additional symptoms [2,3]. The presence of one or two loose stools is defined as mild diarrhoea [1 ,3].

Most frequently, however, travellers' diarrhoea is diagnosed intuitionally, by patients themselves. This usually means that every complex of discomfort symptoms in the gastrointestinal tract that is normally absent in the place of residence of the sick person and is associated with the change of the place of stay is diagnosed as the said condition. Popular names of travellers' diarrhoea ("Pharaoh's curse", "Montezuma's revenge") prove that these ailments are associated with trips to hot countries, typically characterized by lower hygienic standards.

Travellers' diarrhoea is a poly-aetiological disease and the number of microorganisms capable of inducing it is very large. Bacteria are responsible for more than 80% of its cases. Usually they are enterotoxic strains of Escherichia coli (ETEC), which cause 30- 50% of the cases, depending of the area. Other aetiological bacterial factors are: enteroaggregative strains of E.coli (EAggEC) - constituting globally the second cause in terms of frequency; enteropathogenic strains of E.coli (EPEC), diffused adhesion E.coli (DAEC), Campylobacter, Shigella, Salmonella, Aeromonas, Plesiomonas, Vibrio. The presented composition differs significantly in South-East Asia where campylobacterioses constitute up to 40% (probably about 20%) of all infections; there is also a much higher risk of various parasitoses, which is reflected in pharmacotherapy [4].

Other pathogenic microorganisms causing traveller's diarrhoea (such as Salmonella, Shigella, Campylobacter) are also widely spread in the world. The list of pathogenic microorganisms which may be the cause of intestinal infections is constantly growing. The number of infections with Caliciviridaea viruses and protozoons (Cryptosporidium, Cyclospora) is increasing. Therefore essential are epidemiological tests which show the current distribution of particular aetiological factors of diarrhoeas in various geographical regions.

An instance of an infection factor that is very widely spread in the world - both in the mild and tropical climate - producing the symptoms of gastroenteritis are rotaviruses. Although rotaviruses cause diarrhoea mostly in children, there are also acute instances in adults (caused mainly by rotaviruses B and C). Rotaviruses along with noroviruses, adenoviruses type 40 and 41 and astroviruses may cause 5-10% of travellers' diarrhoeas [1 ,4].

Much less frequently the cause are protozoons, such as: Giardia lamblia, Entamoeba histolytica and Cryptosporidium.

In about 40% of the cases no aetiological factor is found; also mixed infections have been identified.

Table 1

Aetiological factors of travellers' diarrhoea in selected holiday destinations (tropical countries). Significant diversification of pathogens depending on location can be seen [4].

Another noteworthy fact is that every time in about half of the cases the pathogen was not found.

Traveller's diarrhoea is diagnosed in 20-40% of people travelling to high-risk areas (Central America, South America, South Asia, most of Africa) and in 10-15% of people visiting the Middle East, China, Russia or the south of Europe [5]. Apart from tourists and business people, another group particularly at risk are soldiers on war and peace missions [6] and pilgrims visiting sanctuaries in the Middle East or India [7].

A number of features directly influencing the chance of contracting the condition has been identified. These include: nationality, age (children are particularly susceptible), current state of health, blood group (group 0 favours infections with Shigella and novoviruses). The most important single risk factor remains geographical location of the destination. An important issue is the character of the planned trip; those travelling on their own, staying at hostels, B&B inns and campings are more at risk. The peak of infections falls in the summer period [1].

Such a high frequency of the problem, along with the ever-growing number of travellers, has very serious economic implications. Every day of indisposition due to travellers' diarrhoea generates $ 500 m losses annually in the tourist industry alone. According to estimates, the total state-borne expenses related to the treatment of this indisposition are about $ 650 m in the United States and€ 450 m in the European Union. It is difficult to estimate the financial expenses borne by the sick who usually treat the illness on their own, using medicines purchased at home and taken with them or available at their destination. These must be, however, high amounts, and according to various polls almost all tourists going to warm and tropical countries take "diarrhoea medicines' with them [data from web sites of pharmaceutical and tourist companies].

The number of inhabitants of Poland going to countries of elevated risk of contracting travellers' diarrhoea is systematically increasing; this tendency has been stable for many years now. Below are data from the Institute of Tourism [http://www.intur.com.pl] for tB^2000's. In red are countries of elevated risk of contracting travellers' diarrhoea.

Tabled

Number of Polish tourists in countries (in millions) - type of trip - abroad

Taking into account also increasingly popular trips to other warm and hot climate countries (e.g.' _. India, Mexico, Cuba etc.) the total number of people at risk in our country may be evaluated at no less than 2 million. Factors which can contribute to the increased frequency of travellers' diarrhoea in this group are indisputably economic ones - most tourist trips from Poland abroad are based on cheap accommodation and dining offers, which translates into increased risk of travellers' diarrhoea.

Prevention

The disease is prevented by observing elementary rules of hygiene, such ;as washing one's hands before meals. No food or drinks from unknown sources (f ea markets, street sellers) should be consumed. During trips to risk areas it is recommended not to eat meals served in room temperature, fresh vegetables, uncarbonated water, ice cubes, unpeeled fruit. As relatively safe is considered the consumption of hot meals, bakery products and other dry foods as well as warm or carbonated drinks. '

A separate group are people who have a higher risk of contracting travellers' diarrhoea and experiencing acute complications of this disease. Here belong mostly, ill people with decreased immunity, those after stomach operations, taking inhibitors of proton pump, ill with unspecific intestinal inflammation; sickle cell anaemia and diabetics treated with insulin. Those persons are advised to use preventive therapy with antibiotics throughout their stay abroad. Similar treatment is recommended in case of people who for professional reasons cannot afford to fall ill while they are away (business people, politicians). Medicines used in the prevention of diarrhoea are administered in lower doses. The risk of contracting travellers' diarrhoea is also decreased as a result of taking preparations of metals, mainly bismuth. Research is being conducted in order to evaluate the beneficial effect of probiotics in the prevention of travellers' diarrhoea, although their effectiveness seems to be low, several percent only [9]. The literature stresses the significance of vaccinating infants against rotaviruses before planned trips [10]. Works are currently in progress on a vaccine against enterotoxic strains of E.coli [11]; recommendations have also been published regarding a vaccine against Vibrio cholerae which (to a smaller degree) also protects against ETEC and travellers' diarrhoea related to that strain [11].

Nevertheless, it should be firmly stressed that therapy with antibiotics is not recommended as a standard procedure in trips to countries characterized by higher risk of contracting travellers' diarrhoea. This recommendation has been clearly formulated by Centers for Disease and Disease Prevention operating in the structures of the US Department of Health and Human Services: "CDC does not recommend antimicrobial drugs to prevent TD"

Treatment - most frequently used methods and preparations

The vast majority of people with travellers' diarrhoea treat its symptoms on their own, using preparations and medicines taken from home or purchased at their destination. If the symptoms do not last very long, but regress quickly, no treatment is necessary. The most important element of the therapy is to drink a lot liquids in order to prevent dehydration and electrolytic disorders. It is also important to adjust the diet to the symptoms in order to eliminate them more quickly.

Hydration. It prevents the loss of electrolytes and water, helps compensate for their deficits. Widely available measures that can be used for that purpose include soups and mineral water with added sugar served with salty snacks. Oral rehydratation salts (ORS) are recommended for small children, the elderly and patients with more intense symptoms. Diet. Often a light diet is recommended based on cooked starch (rice, pastas, wheat, oat) and groats. The meals can also include bananas, vegetables, yoghurts, soups, cooked white meat and crackers. What is to be avoided are meals that are hard to digest and milk with added sugar (large osmotic load). The ill person should eat often but in small portions. When the consistence of stool is improved, regular diet can be returned to.

Peristalsis-limiting medicines (Loperamid, Imodium, Stoperan). The group of medicines that is used by ill individuals most often, also the most frequently ones taken on holiday as "diarrhoea medicines". It should be stressed, however, that medicines from this group have solely symptomatic effects and in a certain time range only. By limiting the peristalsis they favour the propagation of pathogens (by extending the passage time, the time of "stay" of a pathogen in the organism) which would be discharged more quickly without these drugs. Therefore, using medicines from this group may only temporarily inhibit the symptoms, but at the same time it prolongs the total duration of their presence and may constitute a risk factor for more severe forms of travellers' diarrhoea. They should be used mainly to achieve an immediate effect, in strictly specified indications (such as longer trips in local transportation without a toilet). Unconditional contraindications to their use include symptoms of dysentery, aggravation of IDB, obstruction of the alimentary tract, age under 2 and hypersensitivity to ingredients of the preparation.

Toxin-resorbing and secretion-limiting medicines These include mainly medicinal charcoal (Carbo medicinalis) and compounds of metals such as bismuth preparations popular in the USA and recommended by the Centers for Disease and Disease Prevention, operating within the structures of the US Department of Health and Human Services. These medicines act in an auxiliary fashion, they may alleviate the course of the disease, but have no toxic effect on intestinal pathogens.

Probiotics. Their function is still being examined. On the one hand they seem to shorten the time of symptoms (about 1 day of difference) and limit the risk of long-term infection, on the other they are effective only in a small percentage of patients. It seems that colonization with pathogens changes the intestinal environment (pH, organic gasses, competition for alimentary components, mainly prebiotics, adhesion to cells of the intestinal epithelium) so much that probiotics are usually displaced in the course of competition by pathogenic bacteria. Using probiotics seems to be most reasonable in case of simultaneous therapies with antibiotics (and after them!), it must be also based on preparations containing several probiotic bacterial cultures in a proper dose.

Antibiotics. Normally the use of antibiotics is reserved for persons in serious condition and exposed to an increased risk of complications. First line medicines in the therapy of adults are quinolones and of children cotrimoxazole. Moreover, doxycycline is also used and in regions where quinolone-resistant Campylobacter is present - azithromycin. Another drug registered for the treatment of travellers' diarrhoea is rifaximin (Xifaxan). Research has proven the effectiveness of that drug also in cases without a specified aetiological factor.

Another stressed fact is the more and more frequently observed antibiotic resistance of bacteria causing traveller's diarrhoea. What is essential, therefore, is to prevent intestinal infections during one's stay in tropical and sub-tropical zones and to observe the proper time and doses of antibiotics used for therapy (according to the rules). A very frequent mistake influencing the above-mentioned antibiotic resistance takes place when patients use antibiotics without a consultation with a doctor, in too small or too big doses, irregularly and most of all for too short - 2-3 days - until the symptoms recede. Antibiotic therapy may also constitute an independent risk factor for intestinal inflammations (pseudomembranous enterocolitis) often described by patients as "postantibiotic diarrhoeas". If the pathogens are resistant to a given antibiotic, it may eliminate only the saprophytic flora and significantly increase the symptoms and course of diarrhoea.

Using some antibiotics can also result in hypersensitivity to sunlight (e.g. Doxycyclinum) and require avoiding exposure to light and high temperatures, which in case of trips to tropical countries may be very difficult if not impossible.

Hospitalization. The most frequent reasons to commence hospital treatment is significant dehydration (>10% mc.) or symptoms of hypovolaemic shock. Observation is performed in case of patients in a serious general condition who do not respond to regular treatment or have growing complications. Typhoid fever, paratyphoid fever (A, B, C) and cholera are typically treated in special centres only.

An effective treatment of travellers' diarrhoea should focus on restoring the balance of intestinal flora while decreasing the intensity of adverse clinical symptoms. Taking into account the growing resistance to antibiotics and potential dangers and problems related to antibiotic therapy, antibiotics should not constitute the first line procedure. Only rifaximin (Xifaxan) can, while observing the doses and time of therapy, be a first choice drug.

It nevertheless seems that the basis of treatment should be activities aiming at restoring the physiological composition of the saprophytic intestinal flora accompanied with antiinflammatory and regenerative effect on the epithelium of the mucous membrane of the intestine.

Also, as a potential therapy of travellers' diarrhoea in the future a vaccine against ETEC is mentioned. Although initial results of research seem to be promising, an important problem is multi-antigenicity and antigenic variability of strains of this bacterium, as well as the possibility of travellers' diarrhoea being caused by many other bacteria and viruses.

Unexpectedly, it turned out that it is possible to create such a composition of a mixture of protected sodium butyrate and a mixture of protected organic acids of controlled release processes, in such a proportion that a synergism effect of bactericidal activity was obtained, whilst excluding the development of resistance of the bacteria to the active substances applied, accompanied by a strong energy supply of the cells of the mucous membrane (all of the organic acids used in the preparation enter into Krebs cycle).

Unexpectedly, it turned out that it is possible to create such a composition of active substances showing selective (a mixture of protected sodium butyrate and protected organic acids of controlled release processes) and non-selective bactericidal properties (silicon dioxide of a relevant degree of dispersion) and in such a proportion that a synergism effect of bactericidal activity was obtained, whilst excluding the development of resistance of the bacteria to the active substances applied, accompanied by a strong and lasting effect of sorption of medium-molecular proteins, which means the effective removal of bacterial toxins from the intestinal environment. Summary of Invention

Summary of the invention is the application of a mixture of sodium butyrate and organic acids to produce a preparation used to improve the activity of intestinal immunological barrier in the prevention and treatment of travellers' diarrhoea.

Summary of another invention is the application of a mixture of sodium butyrate, organic acids and silicon dioxide to produce a preparation used to improve the activity of intestinal immunological barrier in the prevention and treatment of travellers' diarrhoea.

It is beneficial in the invention if a mixture of sodium butyrate is used in such a way that the preparation contains a mixture of protected sodium butyrate of a specified dynamics of release, in particular along the lower section of the gastrointestinal tract, in a relevant proportion with a proper mixture of protected organic acids of a specified dynamics of release, in particular along the lower section of the gastrointestinal tract, selected from fumaric acid and/or citric acid and/or malic acid and/or sorbic acid for use in the prevention and treatment of travellers' diarrhoea in people.

It is also beneficial if in the invention the preparation contains sodium butyrate and a mixture of acids in triglyceride matrices and if triglyceride matrices have a certain dynamics of release of the active substances they contain, in particular along the lower section of the gastrointestinal tract, moreover if about 1 % of the mass of each triglyceride matrix goes all the way along the human gastrointestinal tract.

It is convenient in the invention if the preparation contains sodium butyrate in the amount of 100-200 mg/g and a mixture of organic acids selected from fumaric acid in the amount of 0-150 mg/g and/or citric acid in the amount of 0-90 mg/g and/or malic acid in the amount of 0-60 mg/g and/or sorbic acid in the amount of 0-80 mg/g and a triglyceride matrix of up to 1 g, particularly if the preparation contains sodium butyrate in the amount of 150 mg/g, fumaric acid in the amount of 100 mg/g, citric acid in the amount of 60 mg/g, malic acid in the amount of 40 mg/g, sorbic acid in the amount of 50 mg/g and triglyceride matrix of up to 1 g.

It is also beneficial if the preparation is packed in hard gelatin or cellulose capsules, preferably cellulose ones containing 500 mg of the product each and the dosage of the preparation is two capsules three times a day, and the dosage of the preparation in more severe diarrhoeas is increased twice, while the preparation is recommended for prevention and to use directly after the symptoms of diarrhoea regress, although it is recommended for use during diarrhoea as well.

Summary of another invention is a preparation containing a mixture of protected sodium butyrate of a specified dynamics of release, in particular along the lower section of the gastrointestinal tract, in a relevant proportion with a proper mixture of protected organic acids of a specified dynamics of release, in particular along the lower section of the gastrointestinal tract, selected from fumaric acid and/or citric acid and/or malic acid and/or sorbic acid and silicon dioxide for use in the prevention and treatment of travellers' diarrhoea in people. It is beneficial in this other invention if the preparation contains sodium butyrate and a mixture of acids in triglyceride matrices and if triglyceride matrices have a certain dynamics of release of the active substances they contain, in particular along the lower section of the gastrointestinal tract, and the applied silicon dioxide has a specified degree of dispersion and if about 1% of the mass of each of the triglyceride matrices goes all the way through the human gastrointestinal tract.

It is also convenient in this other invention if the preparation contains sodium butyrate in the amount of 50-100 mg/g and a mixture of organic acids selected from fumaric acid in the amount of 0-75 mg/g and/or citric acid in the amount of 0-45 mg/g and/or malic acid in the amount of 0-30 mg/g and/or sorbic acid in the amount of 0-40 mg/g, triglyceride matrix of up to 600 mg/g and a highly dispersed silicon dioxide in the amount of up to 600 mg/g, particularly if the preparation contains sodium butyrate in the amount of 75 mg/g, fumaric acid in the amount of 50 mg/g, citric acid in the amount of 30 mg/g, malic acid in the amount of 20 mg/g, sorbic acid in the amount of 25 mg/g and triglyceride matrix in the amount of 300 mg/g and a highly-dispersed silicon dioxide in the amount of 500 g/g.

In this other invention it is beneficial if the preparation is packed in sachets containing 1000 mg and 2000 mg of the product each and the preparation is administered in water to drink, while the dosage of the preparation in children is twice a day one sachet of 1000 mg and in adults - twice a day one sachet of 2000 mg, the dosage in more severe diarrhoeas is increased twice and if the preparation is recommended for use mainly during the diarrhoea and immediately after its symptoms regress.

Using sodium butyrate in accordance with the invention as well as using sodium butyrate in accordance with another invention allowed to establish that sodium butyrate released all the way along the lower section of the gastrointestinal tract and properly selected organic acids are responsible for the activation of the following mechanisms:

• cytoprotective mechanisms (protection of cells against harmful endo- and exogenous factors)

• stimulation of proliferation (stimulating cells of the mucous membrane of the large intestine to grow and divide by activating the physiological track of kinases MAPK, MAPKK)

• activation of apoptosis (ending the life cycle of a cell by the so-called programmed death of the cell - an essential mechanism reducing the risk of developing a malignant tumour)

• stimulation of neoangiogenesis (activating the cells of the mucous membrane to produce VEGF - a factor directly stimulating the growth of the blood vessels)

• stimulation of growth of saprophytic bacterial flora, decrease of adhesion of pathogens to the intestinal walls, activation of the immune system, including macrophages and mastocytes in tissues, anti-inflammatory and anti-oxidizing activity [14,15]

• normalization of the intraintestinal environment by, among others, achieving the proper pH in the whole lower section of the gastrotestinal tract.

It also unexpectedly turned out that it is possible to create such a composition of active substances showing selective (a mixture of protected sodium butyrate and protected organic acids of controlled release processes selected as above) and non-selective bactericidal properties (silicon dioxide of a relevant degree of dispersion) and in such a proportion that a synergism effect of bactericidal activity was obtained, whilst excluding the development of resistance of the bacteria to the active substances applied, accompanied by a strong and lasting effect of sorption of medium-molecular proteins, which means effective removal of bacterial toxins from the intestinal environment.

Apart from the effects mentioned above, an addition of a properly dispersed silicon dioxide showing large affinity with medium-molecular proteins means in reality a strong and effective sorption of bacterial toxins and non-selectively of bacteria. The sorption capacity of toxins of silicon dioxide prepared in such a way is up to 800 mg/g and its sorption capacity of microorganisms - up to 100 microbes/g.

An additional advantage of both proposed compositions is their high tolerance, low toxicity, no negative effect on the alimentary tract.

According to another invention the mixture of protected sodium butyrate and a proper mixture of protected organic acids selected from fumaric acid and/or citric acid and/or malic acid and/or sorbic acid in the form that is slowly released along the gastrotestinal tract, is used for the production of a preparation to prevent travellers' diarrhoea and for use immediately after the symptoms of diarrhoea recede.

According to another invention the mixture of protected sodium butyrate and a proper mixture of protected organic acids selected from fumaric acid and/or citric acid and/or malic acid and/or sorbic acid in the form that is slowly released along the gastrotestinal tract with an addition of properly dispersed silicon dioxide, is used for the production of a preparation to prevent and reduce diarrhoea in particular in travellers going to countries of elevated risk.

Description of Embodiments

Example 1.

Making the preparation used in diarrhoeas in people.

In a homogenizer melt the proper fraction of triglycerides, preferably of vegetable origin, allowed for consumption by humans, add sodium butyrate in the amount of 150 mg/g, fumaric acid in the amount of 100 mg/g, citric acid in the amount of 60 mg/g, malic acid in the amount of 40 mg/g, sorbic acid in the amount of 50 mg/g and triglyceride matrix of up to 1g and send it all to the spray and cooling tower to obtain micro-granules of specified physical and chemical properties. The semi-finished product obtained in such a manner is analysed against compliance with the relevant norm.

In a homogenizer melt successively the proper fraction of triglycerides, preferably of vegetable origin, allowed for consumption by humans, add the relevant amount of organic acids selected from fumaric acid and/or citric acid and/or malic acid and/or sorbic acid along with auxiliary substances and send it all to the spray and cooling tower to obtain micro-granules of specified physical and chemical properties. The semifinished product obtained in such a manner is analysed against compliance with the relevant norm.

The micro-granules obtained in such a manner are mixed in specified proportions, analyse and then send to produce hard gellatin and/or cellulose capsules containing 500 mg of the product each.

Example 2. Making the preparation used in diarrhoeas in people.

In a homogenizer melt the proper fraction of triglycerides, preferably of vegetable origin, allowed for consumption by humans, add sodium butyrate in the amount of 75 mg/g, fumaric acid in the amount of 50 mg/g, citric acid in the amount of 30 mg/g, malic acid in the amount of 20 mg/g, sorbic acid in the amount of 25 mg/g and triglyceride matrix of up to 300 mg/g and highly-dispersed silicon dioxide in the amount of 500 g/g and send it all to the spray and cooling tower to obtain micro-granules of specified physical and chemical properties The semi-finished product obtained in such a manner is analysed against compliance with the relevant norm..

In a homogenizer melt successively the proper fraction of triglycerides, preferably of vegetable origin, allowed for consumption by humans, add the relevant amount of organic acids selected from fumaric acid and/or citric acid and/or malic acid and/or sorbic acid along with auxiliary substances and send it all to the spray and cooling tower to obtain micro-granules of specified physical and chemical properties. The semifinished product obtained in such a manner is analysed against compliance with the relevant norm.

Micro-granules obtained in such a way are homogenized in specified proportions, adding highly-dispersed silicon dioxide in the amount of 500 g/g and the homogenous product is analysed against compliance with the relevant norm.

The product that is compliant with the norm is then packed into sachets of 1000g and

2000g.

Literature:

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15) Dianzani C, Cavalli R, Zara GP, Gallicchio M, Lombardi G, Gasco MR, Panzanelli P, Fantozzi R.: Cholesteryl butyrate solid lipid nanoparticles inhibit adhesion of human neutrophils to endothelial cells. Br J Pharmacol. 2006 Jul; 148(5):648-56.