Is there a new biomarker on the block when it comes to measuring zinc status? …Yes & No!

Julie M. Long of the University of Colorado, Denver discusses the exchangeable zinc pool (EZP) as a biomarker of zinc status and reviews the findings from a recently published sub-study of the Zinc in Powders Trial.

While a lot is known about dietary zinc requirements through the lifecycle and the necessity of this trace mineral for optimal growth, development, and immune function, identification of a ‘gold standard’ biomarker for zinc status remains elusive (1). Currently, serum or plasma zinc status or proxy indicators like dietary zinc intake or stunting rates, are used to estimate zinc status, but each has its limitations.

Small, observational studies suggest that the EZP may be a way to measure zinc status. The EZP is defined as the size of the combined pools, such as the liver, in the body that readily exchange with zinc in the plasma within 2–3 days. This rapidly exchanging zinc is estimated to include approximately 10% of total body zinc and is thought to reflect metabolically active zinc (2). Additionally, adult and pediatric studies suggest that EZP correlates well with dietary zinc intake and zinc absorption, and unlike serum or plasma zinc, EZP size may not decrease when systemic inflammation is present (3-5).

Recently a sub-study of the Zinc in Powders Trial (6) allowed researchers to test the usefulness of EZP as a marker of zinc status via a double blinded randomized controlled trial of zinc supplementation. The sub-study prospectively compared the change in EZP from study start to finish in children who were preventively supplemented with 10 mg of zinc for 6 months as part of a multiple micronutrient powder (MNP) consumed with food or as a zinc dispersible tablet versus a placebo powder.

The major findings of this study included:

·       EZP increased for children who were given zinc supplements compared to no change for children that consumed a placebo powder

·       Children who consumed the zinc dispersible tablet had the greatest increase in EZP, double the increase of children in the MNP group

·       EZP of children experiencing systemic inflammation at baseline did not differ from EZP of children without inflammation

While these results are promising, the limitations of EZP as a biomarker include the requirement for use of stable isotope methodology, including careful urine collections and specialized analyses. Thus, it is less “field friendly” than serum zinc and presently may best be viewed as a useful addition to the zinc toolbox.

The implications of this study’s findings indicated that EZP was responsive to changes in zinc intake and was most responsive to the consumption of the more bioavailable zinc in a dispersible tablet. Additionally, unlike serum or plasma zinc, EZP appears to be resistant to changes when inflammation is present, suggesting it has potential to accurately and reliably measure zinc status!

References:

1.     King, J.C.; Brown, K.H.; Gibson, R.S.; Krebs, N.F.; Lowe, N.M.; Siekmann, J.H.; Raiten, D.J. Biomarkers of Nutrition for Development (BOND)-zinc review. J. Nutr. 2016, 146, 858S–885S.

2.     Miller, L.V.; Hambidge, K.M.; Naake, V.L.; Hong, Z.; Westcott, J.L.; Fennessey, P.V. Size of the zinc pools that exchange rapidly with plasma zinc in humans: Alternative techniques for measuring and relation to dietary zinc intake. J. Nutr. 1994, 124, 268–276.

3.     Krebs, N.F.; Westcott, J.E.; Culbertson, D.L.; Sian, L.; Miller, L.V.; Hambidge, K.M. Comparison of complementary feeding strategies to meet zinc requirements of older breastfed infants. Am. J. Clin. Nutr. 2012, 96, 30–35.

4.     Miller, L.; Long, J.; Mondal, P.; Westcott, J.; Islam, M.M.; Ahmed, M.; Ahmed, T.; Krebs, N. Exchangeable Zinc Pool (EZP) size in Bangladeshi toddlers at risk of environmental enteric dysfunction (EED) is not influenced by inflammation (OR07-03-19). Curr. Dev. Nutr. 2019, 3 (Suppl. 1), nzz034.OR07-03-19.

5.     Miller, L.V.; Hambidge, K.M.; King, J.C.; Westcott, J.E.; Krebs, N.F. Predictors of the size of the exchangeable zinc pool differ between children and adults. J. Nutr. 2017, 147, 187–194.

6.     Islam, M.M.; Black R.E.; Krebs N.F.; Westcott J.; Long J.; Islam K.M.; Peerson J.M.; Sthity R.A.; Khandaker A.M.; Hasan M.; El Arifeen S.; Ahmed T.; King J.C.; McDonald C.M. Different doses, forms, and frequencies of zinc supplementation for the prevention of diarrhea and promotion of linear growth among young Bangladeshi children: a six-arm, randomized, community-based efficacy trial. J Nutr. 2022, 152(5), 1306-1315.

Photo credit: World Bank Photo Collection

The Zinc in Powders Trial: more than a "null finding"

The findings from IZiNCG’s Zinc in Powders Trial (ZiPT), conducted in collaboration with icddr,b, are published! ZiPT was a randomized, partially double-blind, controlled, community-based efficacy trial involving 2886 young children in Dhaka, Bangladesh. Children 9-11 months old were randomly assigned to one of six different intervention groups: a standard micronutrient powder (MNP) containing 4.1 mg zinc and 10 mg iron, daily; a high-zinc (10 mg), low-iron (6 mg) MNP, daily; high-zinc low-iron and high zinc, no-iron MNPs on alternating days; dispersible zinc tablet (10 mg), daily; dispersible zinc tablet (10 mg), daily for 2 weeks at enrollment and 12 weeks; and placebo powder, daily. The interventions were provided for 24 weeks, and intensive twice weekly morbidity surveillance occurred over the intervention period.  To summarize the key findings, there were no differences in the incidence or prevalence of diarrhea across intervention groups, and only the high-zinc, low-iron MNP group had a slightly smaller decline in length-for-age z-score compared with the placebo powder group, prompting a corresponding editorial to ask “has zinc lost its shine?”.

ZiPT was designed to narrow a host of evidence gaps around preventive zinc supplementation and MNP formulations. Despite the consistency of evidence for preventive zinc supplementation in reducing the incidence of diarrhea and eliciting small but significant increases in height, no formal recommendations for preventive zinc supplementation exist. MNP programs are reaching over 10 million children globally, and the standard MNP formulation includes 15 micronutrients, including zinc. Hence MNPs are an attractive vehicle for delivery of preventive zinc and other essential vitamins and minerals to young children. At the time ZiPT was designed, there was some limited evidence that MNPs may be associated with an increase in diarrheal incidence and it was hypothesized that the iron content of the MNPs may be a contributing factor. Furthermore, the available evidence indicated that MNPs had minimal effects on child growth, and evidence was emerging that the amount of zinc in the standard MNP formulation (4.1mg) was probably insufficient in settings with a high prevalence of environmental enteric dysfunction (EED). Lastly, the effect of zinc-containing MNPs on serum zinc concentrations was inconclusive.

The findings from ZiPT underscore the reality that micronutrient supplementation is not a “silver bullet” to improving functional child health outcomes, and that several other factors such as EED, premature introduction of complementary food of poor nutritional quality and intrauterine growth restriction may have had a more substantial impact on diarrhea and linear growth in this study population. However, there are several important take-home messages from the ZiPT findings that deserve highlighting.

First, it was reassuring that MNPs did not cause a higher incidence or prevalence of diarrhea, as previously observed in some trials.

Second, the lack of an effect of stand-alone preventive zinc supplementation on rates of diarrhea was somewhat surprising and contrasts much of the literature, particularly given the high burden of diarrheal disease and high compliance to the study interventions in this population. However, a recent trial in Laos among children aged 6–23 months similarly found that neither preventive zinc nor MNPs significantly reduced the incidence or duration of diarrhea. It is possible that therapeutic zinc, received by all children according to WHO guidelines when experiencing a diarrhea episode, may have helped prevent future episodes of diarrhea among children in all groups. It is also possible that the higher doses of therapeutic zinc supplementation may have local, pharmacologic effects on the gastrointestinal tract, thereby reducing the risk of diarrhea in contrast to lower-dose preventive zinc supplementation.

Third, both the daily dispersible zinc tablet and the high-zinc, low-iron MNP formulation, which provided 10mg of zinc, did a remarkable job at improving serum zinc concentrations:  the prevalence of low serum zinc concentrations fell from 48% to 6% and 29% to 12%, respectively. These marked improvements in zinc status in the absence of a response in functional outcomes raise the possibility that the 24-wk duration of ZiPT may have been insufficient to elicit a response in linear growth. However, the modest improvements in linear growth among children in the daily high-zinc, low-iron MNP group compared with the placebo powder group suggests that the zinc content in the standard MNP formulation may need to be increased, at least in populations at risk for EED. 

Where to next? Should MNPs be recommended as an intervention to improve zinc status despite the lack of impact on diarrhea and linear growth? Given the need for targeted interventions to improve zinc intakes of children 6-23 months of age layered on top of population-based strategies such as large-scale food fortification, MNPs remain an attractive vehicle with potential benefits beyond simply delivering additional zinc and other micronutrients. Furthermore, there is no “one size fits all” approach. Ideally, micronutrient delivery approaches need to be designed and tailored according to local dietary practices, micronutrient deficiency prevalence data, and the existence of other interventions. We will be sharing more findings from ZiPT soon. Stay tuned for a deep dive.

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The ZiPT trial is a collaboration between icddr,b, IZiNCG, the University of California, San Francisco, Johns Hopkins University, and the University of Colorado. Funding for the trial was provided from the Bill & Melinda Gates Foundation to IZiNCG.

Zinc fortification reduces the prevalence of zinc deficiency

Two years in the making, IZiNCG’s Zinc Fortification Task Force is pleased to share the publication Effects of Foods Fortified with Zinc, Alone or Cofortified with Multiple Micronutrients, on Health and Functional Outcomes: A Systematic Review and Meta-Analysis.

Why zinc fortification 

Postharvest food fortification is considered a highly cost-effective intervention to improve dietary intake of micronutrients. As such, many countries with a high burden of zinc deficiency could benefit from including zinc in their mandatory fortification standard. 

Why we conducted this review

IZiNCG last reviewed the evidence on zinc fortification in 2009 and concluded that additional information was needed on the efficacy and effectiveness of fortification programs, as only a few studies had found a positive impact on plasma/serum zinc concentrations or functional indicators of zinc status. Although a Cochrane review was conducted in 2016, it was restricted to studies with comparison groups that allowed the independent effect of zinc to be isolated.  Posing this condition meant excluding a large body of literature relevant to the “real world” context of large-scale food fortification yet to be synthesized for zinc-related outcomes.

What we found

Our systematic review synthesizes data from 59 studies which assessed biochemical and health outcomes after the provision of a zinc-fortified food or beverage. Fortification vehicles were primarily cereal grains and products, followed by beverages and condiments. The median dose of zinc provided by fortification across studies was 4.4 mg per day, study durations varied from 1 month to 3 years, and 71% of studies were conducted in low- or lower-middle-income countries.

We found that food fortification with zinc, given alone or with other micronutrients, increased plasma/serum zinc concentrations, with a corresponding 24% and 55% decrease in the prevalence of zinc deficiency in efficacy and effectiveness studies, respectively. We also found that fortification with zinc and other micronutrients may increase child weight, reduce episodes of diarrhea and fever and improve cognitive function. However, fewer studies evaluated these latter outcomes and the effects cannot be solely attributed to zinc. We found no adverse outcomes after food fortification with zinc.

Why this matters

Findings from this systematic review show that fortification with zinc, alone or together with other micronutrients, is an efficacious and effective strategy for reducing the prevalence of zinc deficiency in low- and middle-income countries. Globally, nearly 30 countries have mandated the inclusion of zinc in their wheat flour, maize, and/or rice fortification programs. Yet, preliminary analyses show that another 30 countries could benefit from either including zinc in an existing fortification program, or establishing a new fortification program that includes zinc. We hope these findings will encourage more countries whose populations could benefit from zinc fortification to include zinc as a fortificant in an existing or new program.  

Zinc Fortification Task Force

The IZiNCG Fortification Task Force is made up of representatives from the IZiNCG Steering Committee, the Food Fortification Initiative, the Global Alliance for Improved Nutrition and Nutrition International.

Enablers and barriers of zinc fortification; experience from 10 low- and middle-income countries

The first publication from IZiNCG’s Zinc Fortification Task Force, Enablers and Barriers of Zinc Fortification; Experience from 10 Low- and Middle-Income Countries with Mandatory Large-Scale Food Fortification, is now available in Nutrients.

Food fortification presents an opportunity for enhancing zinc intakes. Despite data suggesting zinc deficiency as being a potential public health problem, only 29 out of 72 low- and middle-income countries with mandatory fortification programs for cereal grains include zinc as a mandatory fortificant.

To guide policy decisions, we investigated the factors enabling and impeding the inclusion of zinc as a fortificant by conducting a series of in-depth interviews with key informants from 10 countries. We learnt that the decision to include zinc was strongly influenced by guidance from international development partners. Enabling factors included the assessment of zinc deficiency, mandatory regional food fortification standards which included zinc, the World Health Organization (WHO) guidelines for zinc fortification, and the low cost of the zinc compound commonly used. Barriers included the absence of zinc from regional fortification standards, limited available data on the efficacy and effectiveness of zinc fortification, and the absence of national objectives related to the prevention of zinc deficiency.

Systematic review coming soon

More evidence about the impact of zinc fortification is available now compared with when many of the countries interviewed the planning of their zinc fortification programs. We have conducted a review of efficacy and effectiveness studies to ascertain the effect of zinc fortification - alone or with multiple micronutrients - on a range of health outcomes. The review has been accepted for publication in Advances of Nutrition.

IZiNCG’s Zinc Fortification Task Force

The objective of Phase 1 of the Task Force has been to assess the efficacy and effectiveness of zinc fortification interventions, and to identify opportunities to enhance impact. We are now moving into Phase 2 - watch this space. Our members represent the Food Fortification Initiative, the Global Alliance for Improved Nutrition, Nutrition International, and the Global Fortification Data Exchange.

Read more about strategies for promoting zinc nutrition here.

Recent publication: Comparison of laboratory instrument types for analysis of plasma or serum zinc concentration

When standardized methods are used for the preparation and analysis of zinc concentrations in plasma and serum, each laboratory instrument type provides similar mean results and similar accuracy and precision. Read more in IZiNCG’s technical brief no. 12 and in this new publication in Biological Trace Element Research.

Although the prevalence of low plasma or serum zinc concentration (PZC) is associated with the prevalence of inadequate zinc intake, numerous other physiological and methodological factors may also influence PZC. IZiNCG Technical Briefs no. 2 and 6, and IZiNCG Practical Tips documents provide guidance on these factors. However, the potential effects of other analytical issues, such as the type of laboratory instrument used, have not been systematically evaluated. 

Three instrument types are commonly used for PZC analysis: atomic absorbance spectrometers (AAS), inductively-coupled plasma optical emission spectrometers (ICP-OES), and ICP mass spectrometers (ICP-MS).  IZiNCG designed a laboratory methods study to assess the accuracy and precision of these instruments for analyzing zinc concentrations in plasma and serum. Seven laboratories in four countries, including two low- to middle-income countries, using nine instruments (4 AAS, 1 ICP-OES, 4 ICP-MS), participated in the study.  All laboratories received a standard set of samples, materials and reagents, and reference methods for sample preparation and analysis. Read more here.

Updated: 31 August 2021

IZiNCG Awarded New Grant from the Bill & Melinda Gates Foundation to Evaluate the Nutritional Impact of Multiply-Fortified Salt Among Women of Reproductive Age in India

IZiNCG is pleased to announce the receipt of a grant from the Bill & Melinda Gates Foundation to evaluate the nutritional impact of quintuply-fortified salt vs. standard iodised salt for the improvement of micronutrient status among non-pregnant women of reproductive age in Punjab, India. This project is a collaborative effort primarily between IZiNCG, the Postgraduate Institute of Medical Education and Research (PGIMER), and the Haryana State Government.

Why women of reproductive age?

Women of reproductive age (WRA) and young children are particularly vulnerable to multiple micronutrient deficiencies, and prevalences of deficiency remain unacceptably high in India. The 2015-2016 National Family Health Survey revealed that over half of non-pregnant WRA in India are anaemic. A recent survey of 866 WRA in Haryana demonstrated the co-occurrence of multiple micronutrient deficiencies: 75% of WRA had iron deficiency, 80% had folate insufficiency, and 82% had vitamin B12 deficiency (unpublished observations).  Although plasma zinc concentrations were not assessed, the high prevalence of stunting and the low level of absorbable zinc in the national food supply are suggestive of a high prevalence of zinc deficiency.

Why zinc, iron, vitamin B12 and folate?

Zinc, iron, vitamin B12, and folate are critically important for several biological processes related to a healthy pregnancy and the optimal growth and development of the foetus and offspring. Thus, deficiencies in these micronutrients are associated with several adverse health outcomes ranging from preterm and small-for-gestational-age births, neural tube defects, stunting and childhood diarrhoea.  These health problems are widespread in India where more than 3.5 million infants are born preterm each year, nearly 10% of children under 5 have had diarrhoea in the previous 2 weeks, and 38% of children under 5 are stunted. A recent analysis estimated that the prevalence of neural tube defects in South Asia is 32 per 10,000 live births. Clearly, there is an urgent need to improve the micronutrient status of Indian WRA and their children.

Photo credit: World Bank Photo Collection

Salt as a novel vehicle for multiple micronutrient fortification

Micronutrient fortification of a staple food or condiment can be an effective strategy for improving the micronutrient status of a population, as the approach is cost-effective, utilises existing delivery systems, can deliver multiple micronutrients simultaneously, and does not require behaviour change by the population.

Salt is an attractive vehicle for multiple micronutrient fortification in India, as it is universally consumed in fairly consistent amounts. India already has excellent coverage of iodized salt and has recently scaled up the distribution of salt fortified with iodine and iron (double-fortified salt).

Extensive research conducted in various settings has shown that double-fortified salt significantly improves iron status in nutritionally vulnerable groups. With new technology developed by the University of Toronto, it is now possible to fortify salt with multiple micronutrients, including zinc, vitamin B12, and folic acid, in addition to iron and iodine.

While salt fortified with encapsulated ferrous fumarate is already being produced at scale and extensively consumed by large populations in India, the planned study proposes to conduct a head-to-head comparison with ferric pyrophosphate likely in combination with citric acid and trisodium citrate (to enhance iron absorption) in an encapsulated medium that is under development and test whether sensory qualities could be improved further.

Plan for the study 

The study will be conducted in two phases. Phase 1 will include formative research involving a dietary assessment to ascertain habitual salt intake and dietary intake of micronutrients; qualitative research to understand salt procurement, storage, and utilization practices; as well as sensory and acceptability testing of the quintuply-fortified salts.

Phase 2 will be a double-blind, randomised controlled efficacy trial where 750 women of reproductive age will be randomised to 1 of 3 groups: 1) quintuply fortified salt with iron in the form of encapsulated ferrous fumarate; 2) quintuply fortified salt with iron in the form of ferric pyrophosphate; and 3) iodised salt. In brief, participating women will be provided with the assigned study salt on a monthly basis for 12 months and blood samples will be taken at baseline, 6 months, and 12 months. The primary outcome will be micronutrient status as measured by the change in biomarkers of zinc, iron, folate, vitamin B12 and iodine status. Secondary outcomes to be assessed include DNA damage, essential fatty metabolism and cognition. 

If proven efficacious, MFS has the potential not only to improve the micronutrient status of WRA, but also lead to improved perinatal outcomes, and better micronutrient status in their offspring. Together with our collaborators, IZiNCG is excited to initiate this study!

Collaborators: PGIMER, Haryana State Government, SWACH Foundation, WHO Southeast Asia, the University of Toronto, Tata Trusts/ The India Nutrition Initiative, ETH Zürich, JVS Foods, St. John’s Research Institute, University of Colorado Denver, University of California Davis, Eurofins, University of Otago, University of Nottingham, VitMin laboratory.

Last edited 24.03.2020.

Read more:

Strategies for promoting zinc nutrition

Zinc supplementation during pregnancy

Zinc Fortification Task Force 

Recent publication: A dynamic model for predicting growth in zinc-deficient stunted infants given supplemental zinc

Zinc deficiency limits infant growth and increases susceptibility to infections, which further compromises growth. Zinc supplementation improves the growth of zinc-deficient stunted infants, but the amount, frequency, and duration of zinc supplementation required to restore growth in an individual child is unknown. A dynamic model of zinc metabolism that predicts changes in weight and length of zinc-deficient, stunted infants with dietary zinc would be useful to define effective zinc supplementation regimens.

A model of zinc metabolism was developed using data on zinc kinetics, tissue zinc, and growth requirements for healthy 9 month old infants. The model suggests that frequent, smaller doses (5–10 mg Zn/d) are more effective for increasing growth in stunted, zinc-deficient 9-mo-old infants than are larger, less-frequent doses.

In the future, the model predictions of zinc supplementation need to be evaluated in homogenous groups of stunted infants with respect to age, sex, and zinc status. That information will improve predicted amounts of supplemental zinc, and it will identify the population subgroups (e.g., infants with plasma zinc and length-for-age z score below defined thresholds) most likely to respond to zinc supplementation.

The publication can be found here.