# In vitro regeneration of Portulaca grandiflora Hook. and analysis of betalain content within in vivo plants

## TL;DR

This paper describes a method to grow Portulaca grandiflora in the lab and shows how to preserve its natural red-violet pigments for use as a safe colorant.

## Contribution

A reproducible three-step regeneration protocol for Portulaca grandiflora and a method to stabilize its betalain pigments using sodium ascorbate.

## Key findings

- Juvenile leaf explants successfully regenerated shoots using specific plant growth regulators.
- Betalain content in the stem was 26.66 mg/100 g, but degraded by 42.19% within 24 hours.
- Adding 50 mM sodium ascorbate prevented betalain degradation for 24 hours.

## Abstract

Synthetic colors are widely used in the food and cosmetic industries to make products more appealing to consumers. However, the health hazards associated with synthetic colors have prompted their replacement with natural colors. Portulaca grandiflora is a promising candidate for natural color extraction, as it is rich in betalains. This study presents a reliable, reproducible three-step regeneration protocol for this plant and analyzes its betalain content.

In vitro shoot cultures were established on Murashige and Skoog (MS) medium supplemented with different plant growth regulators. Anatomical studies were conducted to determine the stages of shoot primordium development. Betalains were extracted from in vivo plants using 60% methanol and subjected to spectrophotometric analysis. The effect of sodium ascorbate on betalain stability was also evaluated.

Juvenile leaf explants regenerated shoots on MS medium supplemented with 10 μM 6-benzyladenine and 5 μM indole-3-acetic acid. Shoots were multiplied with 20 μM BA (6.25 ± 0.85 shoots/explant), and elongation was achieved with 5 μM gibberellic acid (GA3) (8.2 ± 0.37 shoots/explant). Shoot primordia developed from well-organized meristemoid cells. The betalain content in the stem was 26.66 ± 0.19 mg/100 g, but this pigment degraded within 24 h (42.19% degradation). The addition of 50 mM sodium ascorbate prevented betalain degradation, even after 24 h.

This study reports a regeneration protocol from juvenile leaf explants and demonstrates that betalain stability in the stem can be maintained with 50 mM sodium ascorbate.