Double fortified salt at crossroads.

Iron Deficiency Anemia (IDA) and Iodine Deficiency Disorders (IDD) are the major public health problems often co-existing in many regions in our country. National Institute of Nutrition (NIN) has promoted the technology of double fortification of common salt with iodine and iron as a strategy to control both deficiencies under food-based approaches. Two other formulations of double fortified salt (DFS) have been subsequently developed by other agencies. NIN formulation & Nutrisalt have a stabilizer/promoter to maintain the stability of iodine in the presence of iron. The Micronutrient Initiative (MI) formulation uses physical separation of iodine by microencapsulation. NIN carried out extensive studies on stability, bioavailability, acceptability, safety and impact (including in community) of DFS. Feasibility both at factory level production and community level implementation have been worked out. MI salt had also undergone stability, acceptability and impact studies. No data is reported on the stability of Nutrisalt except that good stability is claimed in the available reports. In principle, the strategy of double fortification of salt with iron and iodine is sound with uniformly good impact on urinary iodine excretion and prevention of anemia. However, striking increments in hemoglobin (Hb) were not readily demonstrated since the intended purpose of DFS was only to provide iron at maintenance level and not therapeutic level. Complexities in the experimental designs, confounding variables and quality of the ingredients in salts also contributed to difficulties in interpretation of Hb status in studies involving DFS. Along with improvements contemplated in formulation to enhance the stability and bioavailibility, DFS should be able to fulfil the promise and realise its potential in reducing iron and iodine deficiency amongst our poor population in the next few years.

Impact evaluation of iron & iodine fortified salt.

As a novel approach to tackle the problems of iron deficiency anaemia and iodine deficiency disorders (IDDs), which often coexist, the National Institute of Nutrition has developed iron and iodine fortified common salt (double fortified salt-DFS) as a public health measure. This salt has undergone a battery of laboratory and field tests to evaluate its feasibility for use in a national programme. The DFS is designed to provide 1 mg of iron and 15 micrograms of iodine per gram of common salt. This was made possible by the inclusion of a polyphosphate stabilizer, sodium hexametaphosphate (SHMP) at 1 per cent level. The stability of iron and iodine was found to be good up to 6 months. However, the stability of iodine depended upon the quality of the salt used for fortification. The biological effects of long-term consumption of DFS were evaluated in experimental rats and in field trials. Both iron and iodine from the salt were found to be biologically available in regenerating haemoglobin and in increasing excretion of iodine in urine. When this salt was tested in tribal villages endemic for goitre and iron deficiency anaemia, the bioresponse was good with regard to the iodine status but was not uniform in all segments with regard to iron, probably due to confounding variables. In a study carried out in residential school children where such variables did not exist, DFS was found to have significant impact on haemoglobin status in anaemic children and improved their urinary iodine excretion. The consumption of DFS for 2 yr did not have any adverse effects in school children as well as in the tribal population. Parameters related to calcium homeostasis were not altered in children receiving DFS. Histopathological examination of tissues and radiological examination of bone did not reveal any abnormality in DFS fed rats. Similarly serum and urinary parameters related to calcium and phosphorus were not altered in DFS fed rats. Therefore, DFS is presented as a feasible and effective strategy to control the double deficiency of iron and iodine in our community.

Current controversies in carotene nutrition.

Most of the dietary vitamin A is derived from plant foods in the form of pro-vitamin A, the carotenoids. Though in 1930 it was first demonstrated that beta-carotene is the precursor for vitamin A and it is well accepted that 1 mole of beta-carotene is equivalent to one mole of vitamin A, the mechanism of conversion to vitamin A has been controversial. Some of the mechanisms suggested are central cleavage potentially yielding 2 molecules of vitamin A or excentric cleavage producing one molecule of vitamin A from beta-carotene which drastically varied the potency of carotene. A mucosal supernatant from rat intestine was shown to have beta-carotene dioxygenase activity which provided the basis for central cleavage. Many observations on enzyme activity in vitro and efficacy of carotene in vivo did not support the above findings and a re-evaluation of the whole problem was undertaken at the National Institute of Nutrition (NIN), Hyderabad. Intestinal conversion of beta-carotene to vitamin A both in vitro and in vivo in rats and in vivo in children was evaluated. A novel method of obtaining the in vivo conversion of carotene to vitamin A using the ratio of area under plasma vitamin A time curves after a dose of beta-carotene and vitamin A (> 100 micrograms) was developed in rats and later extended to children. In children a dose of 1.5 mg of beta-carotene and vitamin A was used. From these studies intestinal conversion of beta-carotene to vitamin A was found to be an enzymatic reaction involving central cleavage and which needed the presence of oxygen. The substrate was found to bind the enzyme at C-15,15'. The enzyme may be associated with inherent or contaminant enzyme which breaks of other part of the molecule released after central cleavage of carotene. The in vivo conversion of carotene to vitamin A was found to vary from 20 to 80 per cent depending on the nutritional status. Vitamin A deficiency was found to enhance both the in vitro and in vivo conversion and protein deficiency to decrease both. Thus the present results confirm the convertibility of dietary carotenoids to vitamin A and could facilitate further investigations on interactions of different dietary carotenoids on the absorption and cleavage of carotene to vitamin A in children.

Biological efficacy and plasma norethisterone levels of orally administered norethisterone enanthate in rat and hamster.

Norethisterone enanthate (NET-En) is a well known intramuscular contraceptive drug. The long acting nature of this preparation when administered orally was evaluated in female rats and hamsters using fertility inhibition test and from the plasma levels of norethisterone (NET). An oral dose of 20-60 mg NET-En was administered to random groups of six female rats and hamsters and were mated after five and ten days with males of proven fertility. The fertility inhibition rate was determined from vaginal delivery. A dose-dependent reduction in fertility was seen in rats 5 days after oral administration of NET-En. This effect was found to be less pronounced and not significant 10 days after administration of similar doses of NET-En. In hamsters, a similar but less pronounced effect was noted. The decrease in fertility was significant only at the 60 mg dose. The plasma levels of NET estimated by RIA over a period of 15 days, in a different set of treated rats, suggested rapid absorption of NET-En within a day, and drug concentration decreased slowly, the levels on the 4th day ranged from 0.9-2.3 with the 10 mg and 1.0-4.0 ng/ml with the 20 mg dose. Detection of adequate levels of NET in plasma during the estrous cycle in rats, and the fertility inhibition observed in female rats and at higher doses in hamsters, suggest that NET-En is orally active.

Isolation and characterization of monkey liver ferritin.

Monkey liver ferritin was isolated and purified along with human liver ferritin and their physicochemical and immunological characteristics were compared. The apparent molecular weight of monkey liver ferritin was estimated to be 430 kDa as against 450 kDa of human liver ferritin. Both ferritins appeared to be made up of a 22.5 kDa polypeptide under denaturing conditions and the proteins contained neutral sugar (wt/wt) of 2.0% (monkey) and 2.4% (human). By immunoblots both human and monkey liver ferritins showed appreciable cross-reactivity with the polyclonal antibodies raised against either proteins. Monkey liver ferritin, however, was not recognised by the human monoclonal antibody. The amino acid composition of both ferritins was more or less similar. Isoelectric focusing indicated that monkey liver ferritin showed microheterogeneity with three bands at pI 5.4, 5.5 and 5.6, whereas human liver ferritin showed a single band at pI 5.6 confirming the relative acidic nature of monkey liver ferritin.

Plasma levels of estradiol & progesterone at different gestational ages in Indian women.

Plasma levels of estradiol and progesterone were determined at different gestational ages in Indian pregnant women from the low socio-economic group. The levels of estradiol and progesterone progressively increased with gestation and reached maximum values (14.35 +/- 1.092 and 145.9 +/- 7.69 ng/ml, respectively) by term. Though the estradiol values were comparable to those reported in literature, the term values of progesterone were found to be lower (145.9 +/- 7.69 ng/ml) than those reported from Western countries (160 +/- 7.5 ng/ml) at 34-38 wk of gestation. These low progesterone values may have physiological implication in placental function.

Evaluation of a colorimetric method for vitamin A estimation.

A simple colorimetric procedure for plasma vitamin A is evaluated, which does not require sophisticated or expensive equipment. Vitamin A values obtained with 30 human plasma samples and 11 liver samples obtained from lactating rabbits, using a colorimetric procedure based on Carr-Price reaction with ferric chloride and acetyl chloride were compared with those obtained with spectrophotometric and spectrofluorimetric micro methods. With widely varying plasma samples, the values showed a high degree correlation and good agreement. The intraassay variation was 3% which is in the acceptable range. The plasma samples could be analysed within 4 weeks and the reagents, were found to be stable, unlike some batches of trifluoro acetic acid (TFA).

Pharmacokinetics of norethisterone from two different combination contraceptive pills in Indian women.

In view of our previous studies that the plasma elimination of norethisterone (NET) from mini pill is faster in low socio-economic group Indian women, the present studies were contemplated to find the least effective dosage of NET from combination pills. Pharmacokinetics of NET were evaluated in a total of twenty women of low socio-economic group taking pills containing NET-acetate (500 micrograms or 1 mg) and ethinyl estradiol (30 or 50 micrograms respectively) on empty stomach. Blood samples were drawn at different time intervals from 0.5 to 24 hr and plasma NET was estimated by a specific radio-immunoassay. In the women taking 1 mg NET-acetate containing pills peak plasma levels ranging from 6.2 to 20.8 ng/ml were observed at 1 hr whereas with 500 micrograms pill they ranged from 2.0 to 6.5 ng/ml and the peak was noted at 4 hr. Pharmacokinetic parameters of NET were more or less comparable between the two pills. The results suggest that pills containing 500 micrograms NET-acetate and 30 micrograms ethinyl estradiol provide adequate levels of NET even in low-socio-economic group women.