*4.5. Summary*

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Our results showing increased acidosis in the ALHMF group raise further concerns with use of the ALHMF, as infants with metabolic acidosis may experience altered nutrient metabolism [28,31] and decreased bone mineralization [32], leading to poor growth and osteopenia of prematurity.

Poor growth in the ALHMF group may also be attributed to changes to the nutrient content of the milk caused by acidification as described by Erickson *et al*. [23]. This group reported significant changes in acidified breast milk, including decreased total protein content, lipase activity, and free fatty acids [23]. The nutritional changes in the composition of acidified breast milk documented by Erickson *in vitro* may have led to the *in vivo* growth deficiencies noted in our ALHMF population [23].

#### **5. Strengths and Limitations**

#### *5.1. Strengths*

This study is the first to quantify results of use of ALHMF in a Level IIIc NICU setting. We are uniquely situated to evaluate outcomes of our use of ALHMF in our patient population for several important reasons. First, we initiated utilization of this product on all infants at one time. There was no possibility of crossover product use to decrease the validity of the data. Additionally, we used this product on all infants who would be eligible to receive fortified human milk, as would be expected in a clinical NICU practice. This makes our data very relevant and applicable to clinical NICU settings.

Second, our clinical management of nutrition in this patient population has been published and remains very successful with excellent growth and low baseline rates of NEC. Not only do we manage nutrition care of this population very closely, but we also have a defined protocol in place so that infants (except for fortification method) receive the same nutrition interventions over time regardless of which group, PHMF or ALHMF, they received.

Additionally, our nutrition management with additional protein added to the PHMF group makes the comparison of the two groups more relevant by giving them a more similar nutrient intake at baseline than a comparison of ALHMF and PHMF alone which compares a large difference in delivered protein.

Finally, we have a very detailed nutrition documentation medical record system, Intuacare. This system allows for easy retrieval of detailed nutrition information including daily percentages of breast milk, daily caloric intake, and daily protein intake in g/kg/day. This allows for minimal reporting error in a retrospective study, such as this, and provides an excellent historical representation of each infant's delivered nutrition.

### *5.2. Limitations*

This retrospective review of a clinical trial of a commercially available acidified liquid human milk fortifier has several limitations including the retrospective nature of the study, and a modest sample size, which limits the power of some data points. These limitations were partially reduced by our reliance on electronic documentation for data collection and analysis. All medical documentation remains variable between individuals and we cannot quantify unrecorded data, but the system utilized allowed for complete assessment of all recorded data on each research subject. As with any study evaluating growth, head circumference and length measurements are also variable as length boards were not used and measuring tape placement may vary between nursing staff. Some subjects were discharged prior to 36 weeks EGA, therefore, anthropometric measurements at 36 weeks EGA were not available. Likewise, lab values were also unavailable for these infants and could not be included in data analysis.

Alterations in human milk composition are continuous, so calculated nutrient compositions of fortified human milk may only serve as general estimations for our nutrient comparisons. Standard NICU nutrition practices are followed as consistently as possible, however feeding advancement may remain variable according to infant clinical status. Furthermore, the proportion of feedings as human milk or formula remained variable among each infant. In an ideal study, all enrolled infants would receive human milk only.

Though the incidence of NEC was statistically significant, it was not powered as a primary outcome for this study. We also suspect that diaper dermatitis was under-recorded during this study period, as our clinical experience suggests that diaper dermatitis is infrequently documented in the electronic medical record even when infants experience more serious medical complications. However, perceived worsening skin breakdown in our unit while using the ALHMF prompted development of a unit list of infants with diaper dermatitis. Unfortunately, not all of the infants recorded on the unit list had diaper dermatitis electronically documented as a medical problem. As there was no way to quantify these cases, these select infants were not coded positively for diaper dermatitis in this study. Therefore, our data analysis remains limited.

#### **6. Conclusions**

Use of the new ALHMF resulted in an increase in clinical complications and a decrease in growth as measured in both g/day and g/kg/day. To our knowledge, this is one of the first studies assessing use of the new ALHMF within a high acuity NICU without excluding infants with significant respiratory disease or low five-min APGAR scores. Further research with the ALHMF is needed to compare infant tolerance and outcomes among infants with a variety of gestational ages, weights, and increased clinical acuity.

#### **Acknowledgments**

The authors would like to thank Sarah Kennedy, Ashley Schlaepfer, and Allison Fischer for their contribution to data collection.

#### **Conflicts of Interest**

The authors declare no conflict of interest.

#### **References**

1. American Academy of Pediatrics. *Pediatric Nutrition Handbook*, 6th ed.; American Academy of Pediatrics: Washington, DC, USA, 2009.


© 2014 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution license (http://creativecommons.org/licenses/by/3.0/).

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Reprinted from *Nutrients*. Cite as: Thorisdottir, B.; Gunnarsdottir, I.; Steingrimsdottir, L.; Palsson, G.I.; Thorsdottir, I. Vitamin D Intake and Status in 12-Month-Old Infants at 63–66° N. *Nutrients* **2014**, *6*, 1182-1193.
